Ecología

Gerardo A. Salazar ^a, José D. Edquén ^b, Jessy P. Arista ^b, Günter Gerlach ^{c, d}, Elmer Yrigoín ^e, Kely Edquen ^f, Mabel Enco ^f, Elí Pariente ^{g, h}, Manuel Oliva ^h y Lidia I. Cabrera ^a, * 

^a Universidad Nacional Autónoma de México, Instituto de Biología, Circuito Zona Deportiva s/n, Ciudad Universitaria, 04510 Ciudad de México, México

^b Universidad Nacional Toribio Rodríguez de Mendoza de Amazonas, Escuela de Posgrado, Jirón Higos Urco Núm. 342, Barrio de Higos Urco, Chachapoyas, Amazonas, Perú

^c Jardín Botánico de Munich-Nymphenburg, Colecciones de Ciencias Naturales del Estado de Baviera, Menzinger Str. 65, 80638 Munich, Alemania

^d Universidad Nacional Mayor de San Marcos, Museo de Historia Natural, Avenida Arenales 1256, Jesús María, Lima 14, Perú 

^e Universidad Nacional de San Martín, Jirón 20 de Abril, Moyobamba 22001, San Martín, Perú

^f Universidad Católica Sedes Sapientiae, Jirón Santa Cruz cuadra 4 s.n., Sector Nuevo Edén, Nueva Cajamarca, Rioja, San Martín, Perú 

^g Universidad Nacional Toribio Rodríguez de Mendoza de Amazonas, Facultad de Ingeniería y Ciencias Agrarias, Herbario KUELAP, Jirón Higos Urco No. 342, Barrio de Higos Urco, Chachapoyas, Amazonas, Perú 

^h Universidad Nacional Toribio Rodríguez de Mendoza de Amazonas, Instituto de Investigación para el Desarrollo Sustentable de Ceja de Selva, Jirón Higos Urco No. 342, Barrio de Higos Urco, Chachapoyas, Amazonas, Perú 

*Autor para correspondencia: lcabrera@ib.unam.mx (L.I. Cabrera)

Recibido: 10 mayo 2024; aceptado: 27 noviembre 2024

Resumen

Se registra por primera vez la presencia en el Perú de Rodriguezia vasquezii, especie conocida previamente solo de Bolivia, a partir de material recolectado recientemente en 2 áreas naturales protegidas (Área de Conservación Privada La Pampa del Burro, departamento de Amazonas y Bosque de Protección Alto Mayo, departamento de San Martín). Se presenta una descripción basada en el material peruano y se ilustra la especie con fotografías a color tomadas en el hábitat y de detalles de su morfología vegetativa y floral. Se exploran sus relaciones filogenéticas mediante un análisis de secuencias de DNA de un marcador nuclear (ITS) y otro de plastidios (un fragmento del gen matK). Su distribución conocida se muestra en un mapa y se contrastan en forma tabular sus diferencias morfológicas con las de especies más cercanamente relacionadas. Se aclara que el holotipo de R. vasquezii está depositado en SEL, no en MO como indicó el protólogo y se reproduce una fotografía de dicho ejemplar y otra de la ilustración original para facilitar futuras comparaciones.

Palabras clave: Andes; Cordillera Oriental; Herbarium Vasquezianum; Holotipo; Rodriguezia pulchra

© 2025 Universidad Nacional Autónoma de México, Instituto de Biología. Este es un artículo Open Access bajo la licencia CC BY-NC-ND

(http://creativecommons.org/licenses/by-nc-nd/4.0/).

Rodriguezia vasquezii (Orchidaceae: Oncidiinae), an addition to the Peruvian flora

Abstract

Rodriguezia vasquezii, previously known only from Bolivia, is recorded for the first time in Peru on the basis of recently collected material in 2 natural protected areas (Área de Conservación Privada La Pampa del Burro, Department of Amazonas and Bosque de Protección Alto Mayo, Department of San Martín). A description based on the Peruvian material is provided, and the species is illustrated with color photographs in habitat and of details of its vegetative and floral morphology. Its phylogenetic relationships are assessed analyzing DNA sequences of a nuclear marker (ITS) and another from plastids (a fragment of the gene matK). A distribution map shows its known distribution, and its morphological differences from closely related species are contrasted in a table. It is clarified that the holotype of R. vasquezii is housed at SEL, not MO as indicated in the protologue, and a photograph of such specimen and another of the original illustration are reproduced to facilitate future comparisons.

Keywords: Andes; Eastern Cordillera; Herbarium Vasquezianum; Holotype; Rodriguezia pulchra

Introducción 

El género Rodriguezia Ruiz et Pav. pertenece a la subtribu neotropical Oncidiinae (Orchidaceae, Epidendroideae, Cymbidieae) y consiste en ca. 50 especies de plantas epífitas de talla reducida (5-20 cm) que habitan las ramas más externas y delgadas de las copas de los árboles y con frecuencia colonizan especies cultivadas, como cafetos (Coffea L.), guayabos (Psidium L.; Chase et al., 2009) y jícaros o totumos (Crescentia L.; G. Gerlach, obs. pers.). Se distribuye ampliamente en el neotrópico, incluyendo Centroamérica (Nicaragua, Costa Rica, y Panamá), las islas de Barlovento y Sudamérica, de Venezuela al noreste de Argentina, con una especie disyunta en el sudoeste de México (González-Tamayo, 1975; POWO, 2024). Rodriguezia se distingue de otras Oncidiinae epífitas de ramitas por una combinación de caracteres florales, como el labelo oblanceolado y apicalmente bilobado, con una proyección retrorsa basal secretora de néctar que se acumula en un mentón o tubo formado por los sépalos laterales, que están parcial o completamente connados, así como la columna con antera dorsal y un lóbulo falcado a cada lado de la cavidad estigmática (Chase et al., 2009). 

En su catálogo de las angiospermas y gimnospermas de Perú, Brako y Zarucchi (1993) listaron 11 especies de Rodriguezia, incluyendo a R. batemanii Poepp. et Endl., R. caloplectron Rchb.f., R. carnea Lindl., R. chasei Dodson et D.E.Benn., R. ensiformis Ruiz et Pav., R. estradae Dodson, R. fernandezii Dodson et D.E.Benn., R. lanceolata Ruiz et Pav., R. pulchra Løjtnant, R. satipoana Dodson et D.E.Benn., y R. strobelii Garay. Subsecuentemente, Bennett y Christenson (1995) describieron una especie peruana adicional, R. bockiae D.E.Benn. et Christenson (como “bockii”).

Durante el trabajo de campo llevado a cabo por nuestro grupo de investigación, entre agosto de 2021 y diciembre de 2023 en 2 áreas naturales protegidas del norte de Perú, el Área de Conservación Privada La Pampa del Burro y el Bosque de Protección Alto Mayo, fueron registradas varias poblaciones de una orquídea epífita de ramitas cuyas características morfológicas, aunque claramente asignables al género Rodriguezia, no coincidían con ninguna de las especies previamente registradas para el país. Una revisión de la literatura permitió asignarla a Rodriguezia vasquezii Dodson, especie conocida previamente solo de Bolivia (Bock, 1997a; Dodson, 1989; Vásquez et al., 2003b). En el presente trabajo, se documenta a R. vasquezii por primera vez para la flora peruana y se le ilustra con fotografías a color tomadas en el hábitat y de detalles de su morfología vegetativa y floral. También se exploran sus relaciones filogenéticas mediante un análisis de secuencias de DNA de un marcador nuclear (ITS) y otro de plastidios (un fragmento del gen matK), se presenta un mapa de su distribución conocida y se le compara en forma tabular con sus congéneres cercanamente relacionados. Se aclara que el holotipo, indicado en el protólogo como depositado en MO, realmente se encuentra en SEL.

Materiales y métodos

Las 2 áreas naturales protegidas donde se registró la especie se ubican en la vertiente amazónica de la Cordillera Oriental de los Andes, en el norte de Perú (fig. 1).

Figura 1. Distribución conocida de Rodriguezia vasquezii (círculos blancos). Mapa por Elmer Yrigoín.

El Área de Conservación Privada La Pampa del Burro (ACPPB) pertenece al asentamiento Perla del Imaza de la Comunidad Campesina de Yambrasbamba, provincia de Bongará, departamento de Amazonas. Tiene una extensión de 2,776.96 ha y abarca un intervalo altitudinal de 1,750-1,900 m snm (coordenadas aproximadas: 5.58°-5.64° S, 77.91°-77.97° O). Presenta 2 biomas principales, bosque achaparrado o matorral sobre arena blanca y bosque montano húmedo a muy húmedo. Por su parte, el Bosque de Protección Alto Mayo (BPAM) abarca 182,000 ha en la porción noroeste del departamento de San Martín y áreas adyacentes de los departamentos de Amazonas y Loreto, en un intervalo altitudinal de 900 a 4,000 m snm (coordenadas aproximadas: 5.47°-6.18° S, 77.19°-77.76° O). Los biomas principales presentes en el BPAM son el bosque achaparrado sobre arena blanca, bosque montano bajo, bosque pluvial montano y pastizal altoandino o jalca. Ambas áreas protegidas son de gran relevancia para la preservación del hábitat de vertebrados como el mono choro de cola amarilla (Lagothrix flavicauda Humboldt, 1812), el mono nocturno peruano (Aotus miconax Thomas, 1927), el oso de anteojos (Tremarctos ornatus F.G. Cuvier, 1825) y la lechucita bigotona (Xenoglaux loweryi O’Neill et Graves, 1977) (SERNANP, 2019; Shanee et al., 2012). La diversidad vegetal de ambas áreas no ha sido documentada, pero nuestros estudios prospectivos de la riqueza táxica de Orchidaceae en ambas han resultado en el descubrimiento de especies nuevas y nuevos registros para el Perú (e.g., Arista, Hágsater, Santiago, Edquén et al., 2023; Arista, Hágsater, Santiago, Pariente et al., 2023; Edquén et al., 2023, 2024; Salazar et al., 2022).

Figura 2. Rodriguezia vasquezii. A, B, Flores de una planta boliviana sin datos precisos de colecta (Seeger s.n.; fotografía de H.-G. Seeger, cortesía de Botanischer Garten Heidelberg, www.botgart.cos.uni-heidelberg.de); C, D, fotografía y disección floral por K. Senghas de una planta boliviana (S. Leferenz s.n.; BASBG-00123123, cortesía de Herbarien Basel [BAS/BASBG/RENZ] bajo licencia CC-BY 4.0).

La descripción morfológica se basó en material peruano estudiado en fresco, complementado con dibujos y fotografías inéditas de plantas bolivianas cultivadas en Alemania (véase Otros registros; fig. 2) y la literatura (Bock, 1997a; Dodson, 1989). Dos ejemplares de la población del ACPPB y uno del BPAM fueron herborizados y depositados en el Herbario de la Universidad Nacional Toribio Rodríguez de Mendoza de Amazonas (KUELAP). Una planta viva de cada una de esas áreas fue fotografiada in vivo con una cámara digital (figs. 3, 4). Se consultó literatura adicional sobre especies sudamericanas de Rodriguezia para determinar las características morfológicas distintivas de R. vasquezii, principalmente Dodson y Bennett (1989), Bock (1993, 1996, 1997a, b), Dodson (1989, 2004) y Jiménez y Jiménez (2014).

Figura 3. Rodriguezia vasquezii. A, Planta en su hábitat en el puesto de control Venceremos del Bosque de Protección Alto Mayo, Departamento de San Martín, Perú; B, acercamiento de la inflorescencia; C, planta del paraje Río Rojo del Área de Conservación Privada La Pampa del Burro, departamento de Amazonas, Perú; D, acercamiento de una flor; E, fotografía con iluminación al trasluz del tubo formado por los sépalos laterales de una flor, mostrando la presencia de néctar (flecha). Escalas: A, B = 5 cm; C = 3 cm; D, E = 1 cm. Fotografías: Gerardo A. Salazar (A, B, E, de Salazar et al. 10829) y José D. Edquén (C, D, de Arista et al. PA-109).

Figura 4. Rodriguezia vasquezii. A, Planta en floración; B, flor, vista lateral; C, flor, vista dorsal; D, flor, vista frontal; E, disección floral; F, antera, vista lateral (izquierda) y ventral (derecha); G, de izquierda a derecha, columna en vista lateral, dorsal y ventral; H, polinario, vista dorsal (izquierda) y ventral (derecha); I, bráctea floral extendida; J, cápsula antes de la dehiscencia (izquierda) y en plena dehiscencia (derecha). Escalas: A = 5 cm; B-E, J = 1 cm; F, H = 1 mm; G, I = 5 mm. Fotografías: Gerardo A. Salazar (de Salazar et al. 10829).

La identificación del material recolectado se llevó a cabo mediante una revisión de la literatura especializada, incluyendo las descripciones e ilustraciones originales (de existir) de todas las especies sudamericanas de Rodriguezia, así como el examen directo o a través de imágenes digitales de los ejemplares de Rodriguezia disponibles en algunos herbarios peruanos (CUZ, HOXA, KUELAP, USM) y del extranjero (AMES, AMO, BAS, GH, HEID, K, LPB, M, MEXU, MO, NY, QCE, QCNE, SEL, US y W), y la consulta de los registros de tipos disponibles en JSTOR GlobalPlants (https://plants.jstor.org/plants/, acceso 25 de enero, 2024). Cabe resaltar que, pese a una búsqueda llevada a cabo por las curadoras respectivas, no se localizó material tipo de R. vasquezii ni en MO, donde el protólogo indica que fue depositado el holotipo, ni en LPB, donde fue depositado el herbario del colector tras su deceso. Sin embargo, fue posible determinar que el holotipo está ubicado en SEL (fig. 5; véase Comentarios taxonómicos).

Métodos moleculares. Se generaron secuencias de nucleótidos de la región de los espaciadores internos transcritos (ITS) del policistrón ribosomal nuclear, incluyendo el ITS 1, el ITS 2 y el gen 5.8 situado entre ellos, así como un fragmento de 773 pares de bases del gen de plastidios matK, de uno de los ejemplares de la ACPPB (Arista et al. 075). El DNA genómico fue extraído a partir de tejido foliar secado en gel de sílice usando el “Plant DNeasy Mini-kit” (QIAGEN, Hilden, Alemania) de acuerdo con las instrucciones del fabricante. La amplificación de ambos marcadores se llevó a cabo mediante la reacción en cadena de la polimerasa (PCR, por sus siglas en inglés) en reacciones de 25 µL con el “Taq PCR Core Kit” (QIAGEN), agregando 0.25 µl de cada cebador (100 ng/µL), 0.125 µL de polimerasa, 1 µL de una solución acuosa 0.4% de albúmina de suero bovino (BSA) para neutralizar inhibidores potenciales (Kreader, 1996) y, en el caso de la región ITS, 0.5 µL of dimetilsulfóxido (DMSO) para reducir problemas asociados a la estructura secundaria del DNA (Jensen et al., 2010). La región ITS fue amplificada y secuenciada con los cebadores 17SE y 26SE (Sun et al., 1994); para el fragmento de matK se emplearon los cebadores matK-1R_KIM y matK-3F_KIM (Hollingsworth et al., 2011). Las condiciones de la PCR para ambos marcadores consistieron en 2 min de desnaturalización a 94 °C; 28 ciclos con 30 s de desnaturalización a 94 °C, 30 s de hibridación de cebadores a 50 °C y 30 s de extensión a 72 °C; y una extensión final de 7 min a 72 °C. Los productos de la PCR fueron enviados para su secuenciación Sanger al Laboratorio Nacional de Biodiversidad (LaNaBio), Instituto de Biología, Universidad Nacional Autónoma de México. Los cromatogramas fueron editados y ensamblados en Geneious Prime 2023.2.1 (Biomatters Ltd.). Las secuencias fueron depositadas en GenBank (tabla 1).

Análisis filogenético. Las secuencias de ITS y matK de R. vasquezii generadas fueron alineadas visualmente con secuencias de los mismos marcadores de 10 especies adicionales de Rodriguezia descargadas de GenBank (https://www.ncbi.nlm.nih.gov/genbank/; acceso 29 diciembre, 2023) y de Polyotidium huebneri (Mansf.) Garay, esta última incluida para enraizar el árbol a partir de un análisis filogenético previo (Neubig et al., 2012; tabla 1). La reconstrucción filogenética se llevó a cabo bajo el criterio de parsimonia analizando las 2 regiones de DNA concatenadas y usando el programa PAUP* v. 4.0a169 para Microsoft Windows de 64 bits (Swofford, 2023), mediante una búsqueda “branch-and-bound” con adición simple de taxones. El apoyo de las ramas se evaluó con 500 búsquedas de bootstrap bajo las mismas condiciones que el análisis de parsimonia.

Tabla 1

Especies incluidas en el análisis filogenético y números de acceso de las secuencias en GenBank.

Resultados

Análisis filogenético. La matriz alineada consistió en 12 taxones y 1,550 caracteres, de los que 777 correspondieron a la región ITS y 773 al fragmento de matK. Un total de 48 caracteres fueron informativos para la parsimonia. El análisis recuperó un solo árbol máximamente parsimonioso (fig. 6), con una longitud de 115 pasos, CI (excluyendo caracteres no informativos) = 0.84 y RI = 0.95. Dentro de Rodriguezia se recuperaron 2 clados principales con fuerte apoyo bootstrap (BP = 100); uno de ellos consiste en una politomía que incluye a R. lanceolata, R. delcastilloi D.E.Benn. et Christenson, R. satipoana, R. venusta (Lindl.) Rchb.f., R. arevaloi Schltr. y R. batemanii. El otro clado principal incluye a R. leeana Rchb.f. como hermana de un agrupamiento con apoyo marginal (BP = 52) consistente en R. lehmannii Rchb.f. y R. vasquezii como hermanas sucesivas del par de especies fuertemente apoyado (BP = 95) formado por R. pulchra y R. chasei.

Figura 5. Holotipo de Rodriguezia vasquezii (Vásquez 112, SEL).

Descripción

Rodriguezia vasquezii Dodson, Icon. Pl. Trop., ser. 2, 4: t. 370. 1989. Tipo: “Bolivia. Cochabamba: Chapare, Río Limatambo, 1,200 m [sin fecha], R. Vásquez 112” (“holotipo: MO”, datos de acuerdo con el protólogo); “Bolivia. Cochabamba: Chapare, km 115 on the way Cochabamba [to] Villa Tunari, 1,160 m, Feb. 1978, R. Vásquez 112” en la etiqueta, holotipo SEL No. 031758 [imagen digital!]) (figs. 2-5, 7).

Figura 6. Único árbol recuperado por el análisis de parsimonia de la región nuclear ribosomal ITS y un fragmento del gen de plastidios matK de 11 especies de Rodriguezia y Polyotidium huebnerii. Los números bajo las ramas indican valores bootstrap. La escala indica el número de cambios en las ramas del árbol.

Hierba epífita de 10-16 cm de altura incluyendo la inflorescencia. Raíces blancas, glabras, 1.0-1.5 mm de diámetro. Seudobulbos estrechamente agrupados, angostamente elipsoides, lateralmente comprimidos, longitudinalmente plurisulcados, 15-20 × 6-7 mm, parcialmente cubiertos por 1-4 vainas conduplicadas, amarillo verdosas, de 5-17 × 4-5 mm, con limbos foliares, éstos verdes, planos, algo suculentos, rígidamente coriáceos, angostamente elípticos a lanceolados, agudos a obtusos y asimétricos en el ápice, con márgenes translúcidos, 18-60 × 11-20 mm, que con el tiempo se pierden y las vainas se tornan escariosas. Hoja 1 en el ápice del seudobulbo, cortamente peciolada, el pecíolo fuertemente conduplicado de 4-5 ×3.0-3.5 mm, limbo como los de las vainas del seudobulbo pero generalmente de mayor tamaño, 55-85 × 22-27 mm. Inflorescencia originada en la base del seudobulbo maduro, arqueado-péndula, hasta 15 cm de largo, con 3-6 flores, éstas simultáneas, dispuestas dísticamente; pedúnculo subterete, verde con difuminación purpúrea, 4-5 cm de largo, ca. 1.5 mm de diámetro, provisto en la base de 2 brácteas fuertemente conduplicadas, verde pálido, translúcidas, agudas, de ca. 8 mm de largo; racimo hasta ca. 10 cm de largo, con el raquis ligeramente en zigzag. Brácteas florales divergentes del raquis ca. 45°, cóncavas, triangulares, agudas, verde pálido, translúcidas, en ocasiones con una ligera difuminación purpúrea, 5-8 × 5.0-5.5 mm. Flores resupinadas, mirando hacia abajo en un ángulo de ca. 45° respecto de la vertical, sépalos blancos, amarillo pálido o amarillo verdoso, el dorsal con algunas manchas y puntos rojo vino en el interior, pétalos blancos a amarillo verdoso con ápice y márgenes blanquecinos con numerosas manchas irregulares rojo vino, labelo blanco a verde pálido en la base, el resto blanco a amarillo verdoso con manchas irregulares y puntos rojo vino en toda la superficie, menos abundantes en la parte central, columna blanca a amarillo pálido con escasos puntos rojo vino, especialmente debajo del estigma y alas amarillas con difuminación rojo vino. Sépalo dorsal cóncavo, obovado, ampliamente redondeado a truncado, 22-24 × 12-13 mm. Sépalos laterales completamente connados en un sinsépalo que en posición original es tubular, formando un tubo nectarífero retrorso, lateralmente comprimido y ligeramente incurvado cerca del ápice, al extenderse el sinsépalo elíptico, subagudo, 23-25 × 13-15 mm. Pétalos recurvados cerca del ápice, obovados, ampliamente redondeados a truncados y someramente emarginados en el ápice, 25-27 × 14-15 mm. Labelo largamente unguiculado, la uña acanalada, angostamente cuneada, 10-11 mm de largo, ca. 3 mm de ancho a la mitad, lámina del labelo pandurada, más amplia y bilobada arriba de la mitad, los lóbulos semiorbiculares, 21-23 mm de largo, 16-18 mm de ancho en la parte más amplia entre los lóbulos extendidos; centro de la lámina con dos pequeñas quillas; base del labelo con una proyección retrorsa, carnosa, sublinear, aguda, que secreta néctar y está envuelta por el tubo formado por el sinsépalo, ca. 12.5 × 3.5 mm. Columna claviforme, 15-16 mm de largo, ca. 1.5 mm de diámetro a la mitad, ca. 5 mm de diámetro en la porción ensanchada cerca del ápice, base provista a cada lado de un borde membranáceo algo prominente, ápice provisto a cada lado de la cavidad estigmática de una aurícula antrorsa, uncinada, algo membranácea, de color broncíneo con difuminación rojo vino, ca. 3 mm de largo, y una prominencia cónica a digitiforme, rojo vino, a cada lado del clinandrio, aproximadamente a la mitad de la antera. Antera 1-locular, semi-elipsoide, algo comprimida lateralmente, redondeada en la parte proximal (hacia la base de la columna), truncada en la parte distal, ca. 5.5. × 2.5 mm. Polinario ca. 4.5 mm de largo, formado por 2 polinios obovoides, amarillos, duros, ventralmente sulcados, ca. 1.5 mm de largo, con estípite tegular, elíptico, atenuado hacia el ápice y terminado en un viscidio oval, café-anaranjado. Rostelo obsoleto. Estigma suborbicular, ca. 3 mm de diámetro. Ovario angostamente obcónico, subterete, ligeramente arqueado, someramente 6-sulcado, 12-20 mm de largo, ca. 1.5 mm de diámetro en la mitad. Cápsula antes de la dehiscencia elipsoide, verde medio con 3 costillas planas, ca. 25 × 15 mm, con pedicelo de ca. 17 mm de largo y ca. 2.5 mm de diámetro a la mitad, al madurar dehiscente a lo largo de las costillas, con los 3 carpelos recurvados exponiendo las placentas y las semillas.

Resumen taxonómico

Distribución. Conocida únicamente de Bolivia (departamentos de Cochabamba y La Paz) y Perú (departamentos de Amazonas y San Martín; fig. 1).

Hábitat. Epífita, en ramitas cubiertas de musgos de árboles pequeños (3-5 m de altura) al borde del bosque montano húmedo y en matorral con palmas, melastomatáceas y Podocarpus sobre arena blanca, a 1,200-1,850 m snm.

Fenología. Floración de agosto a febrero; fructificación, incluyendo cápsulas en plena dehiscencia, observada a fines de noviembre y principios de diciembre.

Material examinado. Perú. Departamento de Amazonas, provincia de Bongará, distrito de Yambrasbamba, Área de Conservación Privada La Pampa del Burro, comunidad Perla del Imaza, Río Rojo, 1,839 m, matorral con dominancia de palmas, Melastomataceae y Podocarpus, 25-VIII-2021, Arista et al. PA-075 (KUELAP-1174!); mismos datos, 1,875 m, Arista et al. PA-109 (KUELAP-1205!). Departamento de San Martín, provincia Rioja, distrito Pardo Miguel Naranjos, Bosque de Protección Alto Mayo, puesto de control del sector Venceremos, vegetación alterada remanente de bosque montano nublado bajo, 1,662 m, epífita, en colonia de unos 6 individuos, en floración y fructificación, 3-XII-2023, Salazar et al. 10829 (KUELAP-3664!).

Otros registros. Bolivia. Sin localidad precisa, planta cultivada en el Jardín Botánico de Heidelberg, octubre 1996, H.-G. Seeger s.n. (fotografías, reproducidas aquí en la fig. 2A, B); departamento de La Paz, planta cultivada en el Jardín Botánico de Heidelberg, 13 octubre 1990, S. Leferenz s.n. (fotografías y dibujo por K. Senghas reproducidas aquí en la fig. 2C, D); departamento de La Paz, área entre Caravani y Bellavista, ca. 1,200 m, colector y fecha no indicados (ilustración de ese ejemplar en Bock, 1997a). Perú. Departamento de San Martín, provincia Rioja, distrito Pardo Miguel Naranjos, Bosque de Protección Alto Mayo, sector Venceremos, 29 enero 2018, Edquén 565, 566 (fotografías KUELAP!).

Comentarios taxonómicos

El material peruano recolectado en el ACPPB y el BPAM coincide morfológicamente con el holotipo (fig. 5) y la descripción e ilustración originales de R. vasquezii (fig. 7), así como con fotografías e ilustraciones hechas de otras plantas bolivianas cultivadas subsecuentemente en Alemania (Bock, 1997a; véase Otros registros; fig. 2). Cabe destacar que una búsqueda del holotipo de R. vasquezii en MO, donde indicó el protólogo que fue depositado, resultó infructuosa y no hay registros de que un ejemplar haya sido ingresado a la colección (Lauren Rogers, in litt. 08 enero, 2024). De igual manera, una búsqueda en el “Herbarium Vasquezianum”, que albergó el material recolectado por Roberto Vásquez durante sus más de 20 años de exploración y estudio taxonómico de las orquídeas bolivianas (Vásquez et al., 2003a), y actualmente depositado en LPB, tampoco permitió localizar un ejemplar de la colección original (Mónica Moraes, in litt. 1 marzo, 2024). Sin embargo, en LPB hay una copia de la ilustración original montada en una hoja de herbario, que porta 2 etiquetas, una tiene la anotación manuscrita “(Holotipo: SEL. Isotipo: MO. VASQ. FAA)”. A partir de dicha información, fue posible corroborar que existe un ejemplar prensado en SEL, el cual constituye el holotipo. Con el fin de facilitar el reconocimiento de la especie, se reproduce aquí una fotografía de dicho ejemplar (fig. 5), además de la ilustración original por Roberto Vásquez (fig. 7). Es importante notar que la información de la recolecta del holotipo indicada en el protólogo difiere parcialmente, tanto de la etiqueta que porta el holotipo, como de las 2 etiquetas de la hoja de herbario con una copia de la ilustración original depositada en LPB. Por ejemplo, mientras el protólogo indica “Río Limatambo”, la localidad señalada en la etiqueta del holotipo es “km 115 on the way Cochabamna [to] Villa Tunari”. Una de las 2 etiquetas de la hoja con la copia de ilustración en LPB indica también “Río Limatambo” mientras que la otra “Villa Tunari”. Por lo tanto, la información contenida en esas varias etiquetas debe considerarse complementaria a la indicada en el protólogo.

Al describir a Rodriguezia vasquezii, Dodson (1989) la comparó con “R. picta Løjtnant”, sin duda por error, pues este nombre no existe y en realidad debió referirse a R. pulchra Løjtnant, la única especie del género descrita por este último autor (Bock, 1996, 1997a; Løjtnant, 1978). Rodriguezia vasquezii se distingue de otras especies andinas del género por la siguiente combinación de caracteres: seudobulbos estrechamente agrupados; vainas del seudobulbo con lámina similar a la de la hoja apical; flores blancas, amarillo verdosas o amarillas con numerosas manchas irregulares y puntos rojo vino en toda la superficie de los pétalos y el labelo; tubo nectarífero del sinsépalo retrorso, ligeramente incurvado cerca del ápice; y lámina del labelo pandurada, con la mitad apical distintamente más amplia que la basal y netamente bilobada. Las características que permiten diferenciar a R. vasquezii de R. chasei, R. leeana, R. lehmannii y R. pulchra, especies presentes en Perú con las que está cercanamente relacionada (fig. 6), se contrastan en la tabla 2.

Figura 7. Ilustración original de Rodriguezia vasquezii por Roberto Vásquez (publicada en Dodson, C. H. [1989]. Rodriguezia vasquezii Dodson. Icones Plantarum Tropicarum, series II, lám. 370. Reproducida con autorización).

La escasez de registros de R. vasquezii parece sugerir que se trata de una especie poco común. Sin embargo, las localidades conocidas en Bolivia y Perú distan entre sí aproximadamente 1,600 kilómetros en línea recta y las áreas intermedias de la Cordillera Oriental de los Andes tienen amplias extensiones de hábitats potencialmente adecuados para esta especie, que en gran medida no han sido exploradas botánicamente (fig. 1). Por otra parte, las plantas de R. vasquezii, como otras orquídeas epífitas de ramitas de la subtribu Oncidiinae, tienen la capacidad de establecerse tanto en árboles nativos como introducidos en áreas sujetas a diferentes grados de disturbio. La talla reducida de las plantas hace difícil localizarlas entre los musgos en las ramas de sus forófitos, pasando fácilmente desapercibidas para colectores no especialistas. Por lo tanto, es probable que la escasez de registros represente una falta de recolecta en la extensa, topográficamente compleja y relativamente inaccesible vertiente amazónica de la Cordillera Oriental de los Andes. Se sugiere la categoría “Datos insuficientes” (DD; UICN, 2019) mientras no haya estudios que evalúen objetivamente el estatus de riesgo de esta especie.

Tabla 2

Comparación de Rodriguezia vasquezii con especies cercanamente relacionadas de acuerdo con el análisis filogenético de la figura 6.

Agradecimientos

Los autores agradecen el apoyo financiero del Fondo Nacional de Desarrollo Científico y Tecnológico del gobierno peruano al proyecto “Diversidad de las Orchidaceae en el bosque montano del Área de Conservación Privada La Pampa del Burro: integrando la taxonomía tradicional y el código de barras de DNA” (contrato Núm. 058-2021-FONDECYT); al Servicio Nacional de Áreas Naturales Protegidas por el Estado (SERNANP) y el Servicio Nacional Forestal y de Fauna Silvestre (SERFOR) del Perú por las autorizaciones para colecta científica (Núm. 006-2018-SERNANP-BPAM-JBPAM y Núm. AUT-IFL-2021-033-SERFOR, respectivamente); a la comunidad de Perla del Imaza por su apoyo para llevar a cabo el trabajo de campo en el Área de Conservación Privada La Pampa del Burro; a Ivonne Paico Vera, Jefa del Bosque de Protección Alto Mayo, por permitir el acceso al área protegida y apoyar la obtención de la autorización de colecta; a Jorge Alcántara, guardaparque del puesto de control Venceremos del Bosque de Protección Alto Mayo, por su apoyo durante el trabajo de campo; a Irene Bock y Olaf Gruss por su ayuda para obtener literatura; a Jay Pfahl y Andreas Franzke (curador del Botanischer Garten Universität Heidelberg) por información sobre fotografías e ilustraciones de R. vasquezii; a Laura Márquez y Nelly López (LaNaBio) por el apoyo técnico para la secuenciación de DNA; a los curadores de AMES, AMO, BAS, CUZ, GH, HEID, HOXA, K, KUELAP, LPB, M, MEXU, MO, NY, QCE, QCNE, SEL, US, USM por las facilidades para acceder a las colecciones a su cargo, especialmente a Lauren Rogers (MO), Mónica Moraes (LPB) y Bruce Holst (SEL) por su ayuda en localización del material tipo de R. vasquezii y la autorización para reproducir la imagen digital del ejemplar en SEL; a Iván Jiménez por la información proporcionada sobre el material de Roberto Vásquez en LPB; a María Isabel Vásquez por la autorización para reproducir la ilustración original de Rodriguezia vasquezii; a Germán Carnevali, Rodolfo Solano y Lourdes Rico por sus sugerencias constructivas a versiones previas del manuscrito. JDE agradece a CONCYTEC – PROCIENCIA, Programa de Doctorado en Ciencias para el Desarrollo Sustentable (contrato Núm. PE501093193-2024-PROCIENCIA-BM). JPA agradece a CONCYTEC-PROCIENCIA, Programa de Doctorado en Ciencias para el Desarrollo Sustentable (contrato Núm. PE501088645-2024-PROCIENCIA-BM).

Referencias

Arista, J. P., Hágsater, E., Santiago, E., Edquén, J. D., Pariente, E., Oliva, M. et al. (2023a). New and noteworthy species of the genus Epidendrum (Orchidaceae, Laeliinae) from the Área de Conservación Privada La Pampa del Burro, Amazonas, Peru. Phytokeys, 227, 43–87. https://doi.org/
10.3897/phytokeys.227.101907

Arista, J. P., Hágsater, E., Santiago, E., Pariente, E., Oliva, M. y Salazar, G. A. (2023b). Epidendrum edquenii (Laeliinae), a new species from montane wet forest of the Área de Conservación Privada La Pampa del Burro, Amazonas, Peru. Lankesteriana, 23, 409–417. http://dx.doi.org/10.15517/lank.v23i2.56342 http://dx.doi.org/10.15517/lank.v23i2.56342

Bennett, D. E. y Christenson, E. (1995). New species of Peruvian Orchidaceae III. Brittonia, 47, 182–209. https://link.springer.com/content/pdf/10.2307/2806958.pdf

Bock, I. (1993). Die Gattung Rodriguezia Ruiz et Pavón, Teil X. Die Orchidee (Hamburg), 44, 175–178.

Bock, I. (1996). Die Gattung Rodriguezia (Teil XIII). Die Orchidee (Hamburg), 47, 236–239.

Bock, I. (1997a). Die Gattung Rodriguezia Ruiz et Pavon (Teil XIV). Die Orchidee (Hamburg), 48, 1–4.

Bock, I. (1997b). Die Gattung Rodriguezia Ruiz et Pavon, Teil XV. Die Orchidee (Hamburg), 48, 151–154.

Brako, L. y Zarucchi, J. (1993). Catálogo de las angiospermas y gimnospermas del Perú. Monographs in Systematic Botany of the Missouri Botanical Garden Vol. 45. Saint Louis: Missouri Botanical Garden Press.

Chase, M. W., Pridgeon, A. M., Veitch, N. y Grayer, R. J. (2009). Rodriguezia. En A. M. Pridgeon, P. J. Cribb, M. W. Chase y F. N. Rasmussen (Eds.), Genera orchidacearum Vol. 5. Epidendroideae Part 2 (pp. 344–347). Oxford: Oxford University Press.

Dodson, C. H. (1989). Rodriguezia vasquezii Dodson. Icones Plantarum Tropicarum, series II, 4,lám. 370. Saint Louis: Missouri Botanical Garden Press.

Dodson, C. H. (2004). Native Ecuadorian orchids. Volume V. Rodriguezia-Zygosepalum. Sarasota: Dodson Publishing.

Dodson, C. H. y Bennett, D. E. (1989). Orchids of Peru. Icones Plantarum Tropicarum series II, 2, láms. 163–165. Saint Louis: Missouri Botanical Garden Press.

Edquén, J. D., Arista, J. P., Damián, A. y Salazar, G. A. (2023). A new species of Liparis section Decumbentes (Orchida-
ceae, Epidendroideae, Malaxidinae) from the Bosque de Protección Alto Mayo, San Martín, Peru. Phytokeys, 224, 89–99. https://doi.org/10.3897/phytokeys.224.98654 

Edquén, J. D., Yrigoín, E., Salazar, G. A., Hágsater, E., Santiago, E., Cabrera, L. I. et al. (2024). Los géneros de orquídeas del Bosque de Protección Alto Mayo. Guía ilustrada. Lima: SERNANP/ Conservation International-Perú.

González-Tamayo, R. (1975). Rodriguezia dressleriana, una especie nueva del occidente de México. Orquídea (Mexico City), 4, 232–240.

Hollingsworth, P. M., Graham, S. W. y Little, D. P. (2011). Choosing and using a plant DNA barcode. Plos One, 6, e19254. https://doi.org/10.1371/journal.pone.0019254

Jensen, M. A., Fukushima, M. y Davis, R. W. (2010). DMSO and betaine greatly improve amplification of GC-rich constructs in de novo synthesis. Plos One, 5, e11024. https://doi.org/10.1371/journal.pone.0011024

Jiménez, M. M. y Jiménez, M. M. (2014). Orquídeas de Zamora Chinchipe: ecología y descripción. Tomo I: Epidendroideae I, subtribus Zygopetalinae, Stanhopeinae, Coeliopsidinae, Catasetinae y Oncidinae (en parte). Zamora, Ecuador: Studio Creativo Zamorarte.

Kreader, C. A. (1996). Relief of amplification inhibition in PCR with bovine serum albumin or T4 gene 32 protein. Applied and Environmental Microbiology, 62, 1102–1106. https://journals.asm.org/doi/pdf/10.1128/aem.62.3.1102-1106.1996

Løjtnant, B. (1978). Rodriguezia pulchra, eine neue Orchidee aus Ecuador. Die Orchidee (Hamburg), 29, 9–12.

Neubig, K. M., Whitten, W. M., Williams, N. H., Blanco, M. A., Endara, L., Burleigh, J. G. et al. (2012). Generic recircumscriptions of Oncidiinae (Orchidaceae: Cymbidieae) based on maximum likelihood analysis of combined DNA datasets. Botanical Journal of the Linnean Society, 168, 117–146. https://doi.org/10.1111/j.1095-8339.2011.01194.x

POWO (Plants of the World Online). (2024). Facilitado por los Reales Jardines Botánicos, Kew. Recuperado el 01 mayo, 2024 de http://www.plantsoftheworldonline.org/ 

Salazar, G. A., Edquén, J. D. y Trujillo, D. (2022). Liparis inaudita (Orchidaceae, Malaxidinae), a new species from the Bosque de Protección Alto Mayo, San Martín, Peru. Botanical Sciences, 100, 506–514. https://doi.org/10.17129/botsci.2999

SERNANP (Servicio Nacional de Áreas Naturales Protegidas por el Estado). (2019). Alto Mayo. Recuperado el 01 mayo, 2024 de: https://www.gob.pe/institucion/sernanp/informes-publicaciones/1834052-bosque-de-proteccion-alto-mayo

Shanee, N., Shanee, S. y Allgas, N. (2012). Expediente técnico que sustenta el reconocimiento. Área de conservación privada “Pampa del Burro – Yambrasbamba”. Neotropical Primate Conservation (NPC) y Sociedad Peruana de Derecho Ambiental (SPDA).

Sun, Y., Skinner, D. Z., Liang, G. H. y Hulbert, S. H. (1994). Phylogenetic analysis of Sorghum and related taxa using internal transcribed spacers of nuclear ribosomal DNA. Theoretical and Applied Genetics, 89, 26–32. https://doi.org/10.1007/BF00226978

Swofford, D. L. (2023). PAUP*. Phylogenetic analysis using parsimony (*and other methods). Version 4. Sunderland: Sinauer Associates. Disponible en https://paup.phylosolutions.com/

UICN (Unión Internacional para la Conservación de la Naturaleza). (2019). Directrices de uso de las Categorías y Criterios de la Lista Roja de la UICN, ver. 14. Preparado por el Comité de Estándares y Peticiones. Recuperado el 01 mayo, 2024 de: https://www.iucnredlist.org/es/resources/redlistguidelines 

Vásquez, R., Ibisch, P. L. y Gerkmann, B. (2003a). Diversity of Bolivian Orchidaceae —a challenge for taxonomic, floristic and conservation research. Organism’s Diversity and Evolution, 3, 93–102. https://doi.org/10.1078/1439-6092-00061

Vásquez, R., Ibisch, P. L. y Gerkmann, B. (2003b). Preliminary list of Bolivian orchid species. Organism’s Diversity and Evolution, 3 (Electronic suppl.), 1–14.

Ilse E. Segura-Zarzosa ^{a, b}, Hortencia Obregón-Barboza ^c, Gopal Murugan ^c,
Christopher B. Boyko ^{d, e}, Gabino A. Rodriguez-Almaraz ^a, Humberto García-Velazco ^f, Alejandro M. Maeda-Martínez ^c, *

^a Universidad Autónoma de Nuevo León, Facultad de Ciencias Biológicas, Avenida Universidad s/n, Ciudad Universitaria, 66455 San Nicolás de los Garza, Nuevo León, Mexico

^b Universidad Juárez del Estado de Durango, Facultad de Ciencias Biológicas, Centro de Estudios Ecológicos, Avenida Universidad s/n, Fraccionamiento Filadelfia, 35010 Gómez Palacio, Durango, Mexico 

^c Centro de Investigaciones Biológicas del Noroeste, S.C., Calle IPN 195, 23096 La Paz, Baja California Sur, Mexico 

^d American Museum of Natural History, Division of Invertebrate Zoology, Central Park West @79th St., New York, NY 10024, USA 

^e Hofstra University, Department of Biology, 114 Hofstra University, Hempstead, NY 11549, USA

^f Unidad de Educación Media Superior Tecnológica Agropecuaria y Ciencias del Mar, CBTA-198, kilómetro 25.5 carretera Ensenada – La Paz, 22790 Ensenada, Baja California, Mexico

*Corresponding author: almaeda04@cibnor.mx (A.M. Maeda-Martínez)

Received: 18 April 2024; accepted: 22 October 2024

Abstract

As part of a research project on the taxonomy of the woodlice (Oniscidea) of northern Mexico, isopod collections of Mexican academic institutions were examined. For the first time, the main diagnostic morphological traits of 4 native species are documented using SEM micrographs: Alloniscus mirabilis (Stuxberg, 1875) (Alloniscidae) and Littorophiloscia richardsonae (Holmes & Gay, 1909) (Halophilosciidae) from the northern Pacific coast of Baja California, Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) (Armadillidae) from northern central Durango, which is also the first record of this species in Mexico, and Porcellio virgatus (Budde-Lund, 1885) (Porcellionidae) from the northeastern states of Nuevo León and Tamaulipas. The first 2 COI haplotypes (mtDNA) for P. virgatus are reported. Bayesian phylogenetic analysis of COI haplotypes placed P. virgatus in a clade with a Porcellio dilatatus haplotype with high support, forming a group with Porcellio scaber.

Keywords: Alloniscidae; Armadillidae; Halophilosciidae; Porcellionidae; Systematics; Taxonomy

© 2025 Universidad Nacional Autónoma de México, Instituto de Biología. Este es un artículo Open Access bajo la licencia CC BY-NC-ND

(http://creativecommons.org/licenses/by-nc-nd/4.0/).

Revisión taxonómica de cuatro isópodos terrestres (Crustacea: Oniscidea) de México

Resumen

Como parte de un proyecto de investigación sobre taxonomía de las cochinillas (Oniscidea) del norte de México se revisaron colecciones de isópodos en instituciones académicas mexicanas. En este trabajo presentamos una relación taxonómica de 4 especies nativas del norte de México y se documentan por primera vez sus características diagnósticas morfológicas principales con micrografías de MEB: Alloniscus mirabilis (Stuxberg, 1875) (Alloniscidae) y Littorophiloscia richardsonae (Holmes y Gay, 1909) (Halophilosciidae) de la costa norte del Pacífico de Baja California, Venezillo apacheus (Mulaik en Mulaik y Mulaik, 1942) (Armadillidae) de Durango, lo cual representa el primer registro de la especie en México, y Porcellio virgatus (Budde-Lund, 1885) (Porcellionidae) de los estados norteños de Nuevo León y Tamaulipas. Reportamos los primeros haplotipos de COI (ADNmt) para P. virgatus. El análisis filogenético Bayesiano de haplotipos COI colocó a P. virgatus en un clado con Porcellio dilatatus con alto soporte formando un grupo con Porcellio scaber. 

Palabras clave: Alloniscidae; Armadillidae; Halophilosciidae; Porcellionidae; Sistemática; Taxonomía

Introduction

The Oniscidea (Crustacea: Isopoda) from Mexico have received little attention (Souza-Kury, 2000). The last taxonomic revision of terrestrial isopods from this country was published 64 years ago by Mulaik (1960). Jass and Klausmeier (2004) concluded that 86 oniscidean species belonging to 33 genera and 16 families have been recorded in Mexico; 10 of them are considered as exotic species (Garthwaite & Sassaman, 1985; Garthwaite et al., 1995; Mulaik, 1960; Rodríguez-Almaraz et al., 2014; Segura-Zarzosa et al., 2020; Souza-Kury, 2000; Treviño-Flores & Rodríguez-Almaraz, 2012). For the northern part of Mexico (including the states of Aguascalientes, Baja California, Baja California Sur, Chihuahua, Coahuila, Durango, Guanajuato, Jalisco, Nayarit, Nuevo León, San Luis Potosí, Sinaloa, Sonora, Tamaulipas, and Zacatecas) 7 exotic species and only 28 native species have been recorded in this region (Jass & Klausmeier, 2004; Rodríguez-Almaraz et al., 2014; Segura-Zarzosa et al., 2020, 2022). Surprisingly, states with large geographic areas like Chihuahua, Durango, Guanajuato and Zacatecas do not have any native species registered (Jass & Klausmeier, 2004). As part of a research project on the taxonomy of the oniscideans of northern Mexico, we examined 258 isopod collection lots deposited in 4 academic institutions, from which 11 species, belonging to 8 genera and 5 families, were identified. Out of 1,820 specimens, 1,417 belong to 6 exotic species (see Segura-Zarzosa et al., 2020), and 160 specimens belong to 5 native species. The remaining 243 specimens are still under study. The first native species reported from this study, Venezillo stuckchensis (Mulaik, 1960), is endemic to the Baja California Peninsula, and was already redescribed (Segura-Zarzosa et al., 2022). In the present work we give a taxonomic account of 4 native species: Alloniscus mirabilis (Stuxberg, 1875) (Alloniscidae) and Littorophiloscia richardsonae (Holmes & Gay, 1909) (Halophilosciidae) from the northern Pacific coast of Baja California, and Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) (Armadillidae) from Durango, which represents the first record of the species in Mexico. After examination and analysis of molecular data and morphological characteristics, we agree with Schultz’s (1975, 1977, 1982) placement of Porcellionides virgatus (Budde-Lund, 1885) in the genus Porcellio Latreille, 1804. Thus, we report the fourth species, Porcellio virgatus (Budde-Lund, 1885) (Porcellionidae) from the northeastern states of Nuevo León and Tamaulipas with its first 2 identified COI haplotypes (mtDNA). 

Materials and methods

The material examined is deposited in 2 scientific collections housed in the Facultad de Ciencias Biológicas, Universidad Juárez del Estado de Durango (UJED), Gómez Palacio, Durango, and Centro de Investigaciones Biológicas del Noroeste, S.C. (CIB), La Paz, Baja California Sur. Only adult specimens were examined and sorted by morphotypes according to the general shape of the cephalothorax, second antennae, pereon, pleon, uropods and pleotelson. Specimens of each morphotype were differentiated as males and females based on sexual dimorphism (Schmidt, 2002). Specimens were examined with a stereomicroscope and a digital caliper (Mitutoyo 700-113, Kawasaki, Japan) was used to record their total lengths (TL). The identity of the material examined was established following morphological descriptions published by Stuxberg (1875), Budde-Lund (1885), Holmes and Gay (1909), Van Name (1936, 1942), Mulaik and Mulaik (1942), Mulaik (1960), Schultz (1975, 1984), and Taiti and Ferrara (1986). The type material of Venezillo apacheus deposited in the Division of Invertebrates, American Museum of Natural History (AMNH) was also examined. Representative specimens of each species were prepared for SEM examination. They were dissected and dehydrated individually in 100% ethanol baths for 24 h and critical-point dried (Samdri-PVT-3B, Tousimis, Rockville, MD, USA), sputter coated with gold (desk II, Denton Vacuum, Moorestown, NJ, USA), and analyzed with a scanning electron microscope (Hitachi S-300N), at CIB.

Table 1

GenBank accession number and origin of COI sequences of Porcellionidae species used in the genetic distance and phylogenetic analyses.

The classification and order of the families follow Ahyong et al. (2011). The assignation of the species to their respective genera and families follows Schmalfuss and Wolf-Schwenninger (2002), Schmalfuss (2003) and Schmidt and Leistikow (2004). As we mentioned above, we agree with Schultz’s (1975) placement of Porcellionides virgatus in the genus Porcellio. Citations of authorship for species described in Mulaik and Mulaik (1942) are given as “Mulaik in Mulaik and Mulaik, 1942”, rather than “Mulaik and Mulaik, 1942” as is usually seen in publications (e.g., Schmalfuss, 2003; Schultz, 1965). This is because all species in the 1942 paper are listed as being described by “Mulaik” alone, although with no indication of which Mulaik (Stanley or Dorothea) should be so credited.

The taxonomic account includes: name, author, and year of description of the family, genus, species, species synonymy (restricted to Mexican material, except for V. apacheus and P. virgatus where a complete synonymy is provided), diagnosis, and a taxonomic summary including the type locality, distribution in Mexico, general distribution, material examined, and remarks on the material examined. The diagnoses of the species are mainly based on the publications of several authors, which are indicated in each corresponding species account, and are updated according to morphological variations found in the material examined. Types of habitus are based on the eco-morphological categories proposed by Schmalfuss (1984). For each species we present their main diagnostic morphological traits using SEM micrographs. The anatomical terminology used in this work is according to Schmidt (2002, 2003). The section of examined material is organized by state, site, geographical coordinates, collection date, collector’s name, number of males and females examined (with TL range, mean ± 1 sd, between brackets), and catalog code. 

Attempts to obtain DNA from specimens of A. mirabilis, L. richardsonae, and P. virgatus were carried out, but useful genetic material was obtained only from specimens of the latter species, from 2 different localities. Genomic DNA was isolated from pereopods using the Gentra Puregene kit (Qiagen). Fragments of COI were amplified with primers 22F (modified primer of LCO1490) (CAA CAA ATC ATA AAG ATA TTG GAA C) and HCO2198 by adopting the cycling conditions of Tizol-Correa et al. (2009). The sequences were edited in DNA Baser 4.5 (www.dnabaser.com) and aligned in Clustal X (Thompson et al., 1997). Genetic distances (uncorrected p-distance) between the 2 P. virgatus sequences and also among the sequences of the porcellionids used in this study (Table 1) were determined in MEGA X (Kumar et al., 2018). The best nucleotide substitution model determined in jModeltest 2.1.10 (Darriba et al., 2012; Guindon & Gascuel, 2003) was used in the Bayesian phylogenetic analysis with MrBayes v3.2 (Ronquist et al., 2012). COI sequences of other species of Porcellionidae were obtained from GenBank (Table 1). The phylogenetic analysis was run for 10 million generations and a consensus tree was generated after eliminating 0.25 of the trees generated during the analysis. The consensus tree was viewed using the program FigTree v1.4.3.

Results

Descriptions

Order Isopoda Latreille, 1817

Suborder Oniscidea Latreille, 1802

Alloniscidae Schmidt, 2003

Alloniscus Dana, 1854

Alloniscus mirabilis (Stuxberg, 1875)

(Fig. 1A-N)

Rhinoryctes mirabilis Stuxberg, 1875, 51-55 (original description).

Alloniscus mirabilis (Stuxberg, 1875).- Schultz, 1984, 155-160; Schmalfuss, 2003, 10; Jass and Klausmeier, 2004, 4, 6, 18.

For a more comprehensive synonymy see Schultz (1984).

Diagnosis. Habitus type runner. Pleon not narrower than pereon. Dorsal surface covered by small tubercles with triangular setae or scales (Fig. 1A, B, M, N). Cephalothorax approximately 1.3 times as wide as long (Fig. 1A, B); frontal shield (lamina frontalis) with medial prominent quadrangular process (Fig. 1B, D); lateral lobes large and conical (Fig. 1B, D). Compound eyes on sides of cephalothorax, each with 17-20 ommatidia (Fig. 1B, C). First antennae 3-jointed, distal article with subapical and apical aesthetascs, basal article longest and broadest (Fig. 1E). Second antennae not reaching pereon-tergite 3 (Fig. 1A), first article short, second approximately 2 times as long as first (Fig. 1A, D), third and fourth subequal, each a little longer than second, fifth approximately 1.5 times longer than fourth (Fig. 1A, F); flagellum 3-jointed, first article little longer than second, third article lanceolate with apical cone, approximately 1.5 times longer than second; complete flagellum approximately 1.2 times longer than fifth article (Fig. 1F, G). Pereon with pereon-tergite 1 approximately 1.5 longer than rest, which are subequal in size (Fig. 1A), proximal-lateral angles produced forward to surround cephalothorax (Fig. 1A, D) up to anterior margin of compound eyes (Fig. 1A, C), pereon-tergites 1-3 with rounded posterolateral margins, pereon-tergites 4-7 produced posteriorly (Fig. 1A), pereopod 1 with transversal antennal brush composed of groove covered by fringed scales on anterior (frontal) side of carpus (Fig. 1H, I), pereopods 1-3 of male with brush of long setae on ventral side of carpus and merus (Fig. 1H), pereopod 7 with pattern of setae similar in males and females (Fig. 1J). Pleon with pleon-tergites 1 and 2 with lateral parts undeveloped, covered at sides by pereon-tergite 7, pleon-tergites 3-5 broadly expanded laterally, lateral margins forming continuous line with lateral margins of pereon-tergites (Fig. 1K, M), pleopod 1 endopodite (copulatory appendage) elongated with tip slightly divergent (Fig. 1K). Pleotelson (pleon-tergite 6) triangular, of same length or shorter than posterior margins of pleon-tergite 5 (Fig. 1M, N), uropod sympodites with wide lobed posterolateral margins, endopodites inserted in internal margin of sympodites bearing group of long setae on apex, exopodites lanceolate (Fig. 1L, N). Sources: Stuxberg (1875) and Schultz (1984).

Taxonomic summary

Type locality. San Pedro, Los Angeles, California, USA (Schultz, 1984; Stuxberg, 1875).

Distribution in Mexico. Only in northwestern Mexico on the Pacific coast of Baja California Sur in Bahía Magdalena, and in Baja California in Isla Cedros (Schultz, 1984), and Bahía San Quintín (this study).

General distribution. From Pacific Grove, Monterey, California, USA in the north, to Bahía Magdalena, Baja California Sur, Mexico in the south (Jass & Klausmeier, 2004; Schultz, 1984).

Material examined. Bahía Falsa, San Quintín, Baja California, Mexico (30°27’14.34” N, 116°00’06.50” W), on a sand beach, 02-12-2006, coll. A. Maeda, 1 male (8.8 mm), 5 females (8.1-9.7, x 8.8 ± 0.63 s.d. mm TL) (CIB-41B). 

Remarks

As reported by Stuxberg (1875) for the A. mirabilis type specimens from California, the color of the dorsal side of alcohol-fixed specimens examined exhibited a light brown color with a dorsal medial dark line and lateral light and dark spots. Schultz (1984) reported A. mirabilis from Oxnard, California with 12-13 ommatidia, while our specimens have 17-20 ommatidia. 

Armadillidae Brandt in Brandt and Ratzeburg, 1831

Venezillo Verhoeff, 1928

Venezillo apacheus (Mulaik, 1942 in Mulaik & Mulaik, 1942)

(Figs. 2-8)

Cubaris apacheus Mulaik, 1942 in Mulaik and Mulaik, 1942, 8-9 (original description). 

Cubaris apachea Mulaik, 1942.- Van Name, 1942, 313.

Cubaris apacheus Mulaik, 1941 [sic].- Mulaik and Mulaik, 1943, 9.

Venezillo (Venezillo) apacheus (Mul.).- Arcangeli, 1957, 119.

Cubaris apacheus Mulaik and Mulaik, 1942.- Boyko, 1997, 7-8.

Venezillo apacheus (Mulaik, 1942).- Leistikow and Wägele, 1999, 46.

Venezillo apacheus (Mulaik & Mulaik, 1942).- Schmalfuss, 2003, 285.

Figure 1. SEM micrographs of adult male Alloniscus mirabilis (Stuxberg, 1875) from Bahía Falsa, San Quintín, Baja California, Mexico. A, Cephalothorax, second antenna and pereon-tergites in dorsal view; B, cephalothorax in dorsal view; C, left compound eye with ommatidia in lateral view; D, cephalothorax in anteroventral view; E, left first antenna in lateral view; F, left second antenna in ventral view; G, flagellum of left second antenna in ventral view; H, left pereopod 1 in medial view; I, carpal brush of left pereopod 1 in medial view; J, left pereopod 7 in medial view (central appendage in the micrograph); K, pleopods 1-5, endopodites (copulatory appendages) of pleopods 1, uropod sympodites with endopodites and exopodites; L, uropod sympodites with endopodites and exopodites in ventral view; M, pleon-tergites 1-5, triangular pleotelson and uropod sympodites with exopodites in dorsal view; N, pleon-tergite 5, and triangular pleotelson with uropod sympodites and exopodites in dorsal view. Scale bars: A = 3 mm; B = 500 μm; C, G = 300 μm; D, F, H, J, K, L, N = 1 mm; E = 100 μm; I = 200 μm; M = 2 mm.

Figure 2. Adults of Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) from Gómez Palacio, Durango, Mexico. A, C-F are SEM micrographs; B is a stereo light microscope micrograph. A, Habitus of male in right lateral view; B, habitus of female in left lateral view; C, habitus of male in ventral view; D, cephalothorax and pereon-tergite 1 of male in right anterolateral view; E, external surface of frontal shield of male covered by semi-curved scales with scattered tricorn scales; F, external surface of vertex of cephalothorax of male covered by semi-curved scales with scattered tricorn scales. Scale bars: A-C = 2 mm; D = 1 mm; E = 100 μm; F = 200 μm.

Diagnosis. Habitus type endoantennal conglobator (Fig. 2A, B). Color pale yellowish (alcohol preserved specimens). Dorsal surface with scales and tricorn process inserted in diverse positions (Figs. 2E, F; 3E; 7E, F). Cephalothorax approximately 1/3 longer than wide (Fig. 2A-D); frontal shield (lamina frontalis) with proximal lateral antennal lobes for holding proximal portion of second antenna during conglobation (Fig. 2D), posterior margin expanded, remaining a groove behind frontal shield (Figs. 2B, D; 3A). Compound eyes on sides of cephalothorax, each with 8-9 ommatidia in 2 rows (Fig. 3A-D).

Figure 3. SEM micrographs of adult Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) from Gómez Palacio, Durango, Mexico. A, Cephalothorax and pereon-tergite 1 of male in lateral view showing ommatidia of the complex eye; B, left complex eye of male in anterolateral view showing ommatidia; C, right eye of female in lateral view showing ommatidia; D, left eye of female in lateral view; E, frontal shield of male in dorsal view covered with semicircular scales and tricorn scales; F, third article of right first antenna of male with aesthestascs. Scale bars: A, B = 500 µm; C, D, F = 300 µm; E = 100 µm.

First antenna 3-jointed with 13 aesthetascs on distal article (Figs. 2F; 4A), basal article wide and subequal in length as terminal one (Fig. 3A). Second antennae with first article short, second approximately 2 times as long as first, third subequal to second, fourth 1.5 times longer than third, fifth approximately 1.3-1.5 times longer than fourth (Fig. 4B); flagellum 2-jointed, distal article lanceolate, 3 times longer than first, complete flagellum approximately 2/3 of fifth antennal article (Fig. 4B, C). Right and left mandibles with similar pars incisive, lacinia mobilis, and pars molaris (Fig. 5A-D); lateral side of mandibles covered by semi-curved scales with some acute scales (Fig. 5B), pars incisive with 3 round projections, central one largest (Fig. 5C), lacinia mobilis with basal setose lobe (Fig. 5D); second maxilla membranous, laminate, inner lobe smaller, covered by abundant short setae, outer lobe wide, rounded, covered by pubescence; maxilliped palp with 3 articles (Fig. 5E, F). Pereon with pereon-tergite 1 with notch (schisma) along epimeron (Fig. 6A) into which anterior corner of pereon-tergite 2 epimeron fits when animal conglobates; pereon-tergite 2 with large digitiform lobe on anterior ventral side (Fig. 6B); pereon-tergites 3-5 with rounded epimeron (Fig. 2A, B); pereopod 1 with antennal brush composed of groove covered by transversal scales on anterior (frontal) side of carpus, inner side of merus, carpus and propodus with spines; sternite 7 with medial process apically bilobate, covering proximal portion of genital papilla (Fig. 6E, F). Pleon with pleon-tergites (pleon-segments) 1-2 with lateral parts undeveloped and covered at sides by pereon-tergite 7 (Fig. 2A, B); pleon-tergites 3-5 broadly expanded laterally, lateral margins forming continuous line with lateral margins of pereon-tergites (Fig. 2A, B); male genital papilla with lanceolate ventral shield (Fig. 6F); pleopod 1 endopodite (copulatory appendage) elongated with acute apex (Fig. 7A). Pleotelson wide at base, becoming constricted approximately at 3/4 of length, then expanding to truncate caudal margin, dorsomedial surface slightly raised, some specimens with line (somewhat keeled) (Fig. 7A-C); uropod sympodites (uropod protopodites) enlarged and flattened (Fig. 7C), filling space between caudal side of pleon-tergites 5 and lateral side of pleotelson (Fig. 7C, D), exopodites conical and small, inserted on notch in medial margin of sympodites (Fig. 7C, D), endopodites oblong, flat, covered by scales and setae, base covered by basal plate of uropodal sympodite (Fig. 8A-D). Sources: Mulaik in Mulaik and Mulaik (1942) and Van Name (1942).

Taxonomic summary

Type locality. Alice, Brooks County, Texas, USA (Mulaik in Mulaik & Mulaik, 1942).

Type material examined. Holotype male (AMNH 17515) and allotype female (AMNH 17516) with the original label: “Cubaris apacheus M&M, Alice, Brooks Co., Texas, Dec 1939 S-D Mulaik.” Holotype (damaged), ca. 7 mm TL, allotype (partially conglobated), ca. 5 mm TL. Paratypes (AMNH 17517) with the original label: “Cubaris apacheus M&M, Alice, Brooks Co., Texas, Dec 1939 S-D Mulaik.” 12 specimens intact, partly conglobated, mixture of males and females (ca. 4.5-6.5 mm TL), and fragments of 5 specimens.

Figure 4. SEM micrographs of adult Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) from Gómez Palacio, Durango, Mexico. A, Left first antenna of male in lateral view; B, right second antenna of female in dorsal view; C, flagellum of right second antenna of female in dorsal view; D, apex of distal article of flagellum of right second antenna of female in dorsal view; E, labrum and clypeus of male in anteroventral view; F, clypeus and frontal shield of female in anterior view. Scale bars: A = 100 µm; B = 500 µm; C = 200 µm; D = 50 µm; E, F = 300 µm.

Mexican material examined. Gómez Palacio, Durango, Mexico, 23-06-1994, coll. A. Bedolla, 1 male (7.0 mm TL), 2 females (7.6-9.7 mm TL) from the garden of a house (UJED-09).

Distribution in Mexico. Known only from Gómez Palacio, Durango. This is the first record of the species in Mexico, and the first record of a native oniscidean species in the state of Durango.

General distribution. Known from Alice, Kerville, Laredo (Mulaik in Mulaik & Mulaik, 1942), and Jim Wells County (Mulaik & Mulaik, 1943), Texas, USA, and Durango, Mexico.

Remarks

The type material examined fits well the description published by Mulaik in Mulaik and Mulaik (1942). The only difference noted between the type material and the Mexican specimens is that the fifth article of the second antenna is approximately 1.3 times longer than the fourth article in the type specimens, while in the Mexican specimens it is approximately 1.5 times longer than the fourth article (Fig. 4B). According to Mulaik in Mulaik and Mulaik (1942) the most similar species to V. apacheus is V. chamberlini (Mulaik in Mulaik & Mulaik, 1942), a oniscidean from Edinburg, Hidalgo County, Texas, USA, that can be differentiated by having 6 ommatidia, pleon-tergite 5 posterior margins visible in dorsal view, and the inner border of the epimeral schisma not visible in lateral view.

Figure 5. SEM micrographs of female Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) from Gómez Palacio, Durango, Mexico. A, Left mandible in posterior view; B, left mandible in anterior view; C, pars incisus and lacinia mobilis of left mandible, from A; D, pars incisus, lacinia mobilis, and pars molaris of left mandible, from B; E, left maxilliped in anteroventral view; F, left maxilliped palp with 3 articles in anteroventral view. Scale bars: A, B, E = 500 µm; C, F = 100 µm; D = 200 µm.

Halophilosciidae Verhoeff, 1908 (cf. Schmidt, 2003; Schmidt & Leistikow, 2004)

Littorophiloscia Hatch, 1947

Littorophiloscia richardsonae (Holmes & Gay, 1909)

(Fig. 9A-N)

Philoscia richardsonae Holmes and Gay, 1909, 378-379 (original description).

Philoscia richardsonae Holmes and Gay, 1909.- Mulaik, 1960, 158.

Littorophiloscia richardsonae (Holmes & Gay, 1909).- Leistikow and Wägele, 1999, 18; Souza-Kury, 2000, 245; Schmalfuss, 2003, 133; Jass and Klausmeier, 2004, 3, 10; Taiti and Ferrara, 1986, 1350-1354.

For a more comprehensive synonymy see Taiti and Ferrara (1986).

Diagnosis. Habitus type runner (Fig. 9A). Pleon abruptly narrower than pereon. Color brownish with spots on dorsal and ventral sides of body, pereon-tergites with 3 longitudinal dark lines, 1 medial and 2 lateral. Dorsal surface with short minute scale-spines, margins with line of small spines (Fig. 9B). Cephalothorax 2 times as wide as long (Fig. 9B), vertex convex (Fig. 9D), frontal shield prominent, rounded with posteromedial border concave (supra-antennal line) (Fig. 9D), lateral lobes small, subtriangular (Fig. 9B). Compound eyes at anterolateral sides of cephalothorax, each with 12-15 ommatidia (Fig. 9C). First antennae 3-joined with aesthetascs on distal article, basal article longest, broadest (Fig. 9E). Second antennae long, approximately 1/2 of total length, first article short, second and third 2 times as long as first, fourth 2 times longer than third, fifth 1.5 times longer than fourth (Fig. 9F); flagellum 3-joined, first and third articles subequal, slightly longer than second (Fig. 9G). Pereon with pereon-tergite 1 broadest (Fig. 9A), pereon-tergites 3-7 with rectangular lateral borders, pereopod 1 antennal brush with transversal scales on anterior (frontal) side of carpus (Fig. 9H, I). Pleon with pleon-tergites 3-5 posterolateral margins strongly produced and curved backwards, margins with line of small scale-spines (Fig. 9J, M); male pleopod 1 endopodite (copulatory appendage) elongated with acute apex (Fig. 9J); pleopods 1-5 with triangular exopodites (Fig. 9J). Pleotelson triangular with rounded tip and concave lateral borders, margins with line of small scale-spines (Fig. 9M, N); uropod sympodites as long as wide, complete structure of endopodites approximately 2/3 as long as exopodites, in dorsal view endopodites approximately 1/3 as long as exopodites, with long apical setae, inserted in sympodites before exopodites, exopodites long, subconical, lateral margin nearly straight, medial margin somewhat convex (Fig. 9J, L). Sources: Holmes and Gay (1909), Van Name (1936) and Taiti and Ferrara (1986).

Figure 6. SEM micrographs of male Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) from Gómez Palacio, Durango, Mexico. A, Cephalothorax and pereon-tergite 1 with notch (schisma) along epimeron, pereon-tergite 2 with large digitiform lobe on anterior ventral side; B, lateral margins of pereon-tergites 1-4, and left pereopod 2 in ventral view; C, left pereopod 1 in anterior view; D, propodus and dactylus of left pereopod 1 in anterior view; E, sternite 7 with medial bilobate process covering proximal portion of genital papilla in posteroventral view; F, genital papilla with lanceolate ventral shield in posteroventral view. Scale bars: A = 1 mm; B, C = 500 µm; D-F = 200 µm.

Taxonomic summary

Type locality. San Diego, California, USA (Holmes & Gay, 1909).

Material examined. Bahía Falsa, San Quintín, Baja California (30°27’14.34” N, 116°00’06.50” W), on a sand beach, 02-12-2006, coll. A. Maeda, 8 males (4.8-8.3 mm, x 6.9 ± 1.02 s.d. mm TL), 10 females (4.7-8.0 mm, x 6.6 ± 0.94 s.d. mm TL) (CIB-42B).

Distribution in Mexico. Isla Cedros, Baja California (Mulaik, 1960; Taiti & Ferrara, 1986), and Bahía Falsa, San Quintín, Baja California, a new locality record for the species. 

General distribution. From northwestern Mexico to Vancouver Island, Canada (Garthwaite et al., 1985; Schmalfuss, 2003; Taiti & Ferrara, 1986).

Figure 7. Adults of Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) from Gómez Palacio, Durango, Mexico. A, C, D-F are SEM micrographs; B, is a stereo light microscope micrograph. A, Pleon-tergites 3-5, pleotelson and uropod sympodites of male in posterior view; B, pleon-tergites 1-5, pleotelson and uropod sympodites of male in posterodorsal view; C, pleotelson and uropod sympodites with exopodites of female in posterior view; D, left uropod exopodite in sympodite from C; E, dorsal surface of female pleotelson; F, dorsal surface of pleotelson with semicircular scales and tricorn scales from E. Scale bars: A, C = 500 µm; D = 300 µm; E = 100 µm; F = 50 µm.

Remarks

Taiti and Ferrara (1986) reviewed Littorophiloscia and recognized 15 species, 2 of them recorded in Mexico: L. richardsonae from Baja California and L. tropicalis Taiti and Ferrara, 1986 from Yucatán. Taiti and Ferrara (1986) reported specimens of L. richardsonae up to 7 mm of length with 12 or 13 ommatidia and pereopod 1 with an antennal brush with transversal scales on the anterior (frontal) side of the carpus and propodus; our examined specimens have a mean TL of 6.9 mm for males, 6.6 mm for females, the number of ommatidia varied from 13 to 15, and the antennal brush is only on the carpus (Fig. 9H, I).

Figure 8. SEM micrographs of adults of Venezillo apacheus (Mulaik in Mulaik & Mulaik, 1942) from Gómez Palacio, Durango, Mexico. A, Pleotelson and uropod sympodites with endopodites of male in ventral view; B, pleotelson and uropod sympodites with endopodites of female in ventral view; C, uropod endopodites of male in ventral view; D, uropod endopodites of female in ventral view. Scale bars: A, B = 500 µm; C, D = 100 µm.

Porcellionidae Brandt in Brandt and Ratzeburg, 1831

Porcellio Latreille, 1804 

Porcellio virgatus (Budde-Lund, 1885)

(Fig. 10A-L)

Metoponorthus virgatus Budde-Lund, 1879, 4 (nomen nudum).

Metoponorthus virgatus Budde-Lund, 1885, 182 (original description).

Metoponorthus virgatus Budde-Lund, 1885.- Richardson, 1905, 630; Mulaik, 1960, 174.

Porcellionides virgatus (Budde-Lund, 1885).- Van Name, 1936, 241; 1940, 137; 1942, 327; Mulaik and Mulaik, 1942, 7; 1943, 7; Leistikow and Wägele, 1999, 36; Souza-Kury, 2000, 245; Jass and Klausmeier, 2004, 5. 

Porcellionides mulaiki Van Name, 1936, 522.- Van Name, 1940, 137.

Porcellio virgatus (Budde-Lund, 1885).- Schultz, 1975, 185-193; 1977, 152; 1982, 2, 5, 6, 20-21.

“Porcellionides” virgatus (Budde-Lund, 1885).- Schmalfuss, 2003: 214.

Diagnosis. Habitus type runner. Color brown, mottled with light spots, 2 light lateral and 1 medial lines. Body surface with small tubercles and rounded tricorn scales (Fig. 10A, B, D, L). Cephalothorax approximately 2 times as wide as long (Fig. 10B), vertex with convex medial part (Fig. 10B), frontal shield with small rounded lateral lobes (Fig. 10B), posterior frontal line forming slightly medial convex margin (Fig. 10B, D), supra antennal line V-shaped absent (Fig. 10D). Compound eyes each with 19-30 ommatidia (Fig. 10C). First antennae 3-joined; basal article longest and broadest, second short, third with subapical and apical aesthetascs (Fig. 10F). Second antennae with first article short, second 2 times as long as first, third subequal to second, fourth 1.5 times longer than third, fifth approximately 1.5 times longer than fourth (Fig. 10G); flagellum 2-joined, proximal article shorter or subequal to distal article, distal article lanceolate with apical cone (Fig. 10H). Maxilliped palp with 3 articles (Fig. 10E). Pereon-tergites 1-7 with a well marked dorsal transverse furrow (Fig. 10A); pereon tergite 1 with projected anterolateral sides not reaching cephalothorax anterior margin, 1.5 times longer than pereon-tergites 2-7 (Fig. 10A); sternite 7 without medial process (Fig. 10I). Pleon not abruptly narrower than pereon (Fig. 10K); pleon-tergites with acute lateral margins posteriorly directed (Fig. 10K); male pleopod 1 endopodites (copulatory appendages) lanceolate (Fig. 10I); pleopods 2-5 with exopodites semi-quadrangular (Fig. 10I). Pleotelson triangular with rounded tip (Fig. 10K, L); uropod sympodites twice as long as wide, as long as exopodites, endopodites thin, lightly flattened, reaching tip of exopodites, exopodites subconical (Fig. 10J). Sources: Budde-Lund (1885), Richardson (1905), Van Name (1936), Mulaik (1960), and Schultz (1982).

Figure 9. SEM micrographs of male Littorophiloscia richardsonae (Holmes & Gay, 1909) from Bahía Falsa, San Quintín, Baja California, Mexico. A, Cephalothorax with right antenna and pereon-tergites 1-6 in dorsal view; B, cephalothorax in dorsal view; C, left compound eye with ommatidia in anterolateral view; D, cephalothorax in anterior view (left second antenna removed); E, left first antenna in anterior view; F, right second antenna in dorsal view; G, flagellum of right second antenna in dorsal view; H, right pereopod 1 in medial view; I, carpus with brush, protopod and dactyl of pereopod 1 in medial view (from H); J, pleon-tergites 1-5 with pleopods, copulative appendages of pleopod endopodites, uropods sympodites with endopodites and exopodites in ventral view; K, endopodites of uropod sympodites in ventral view; L, right uropod sympodite with exopodite in posteroventral view; M, pleon-tergites 2-5, triangular pleotelson and uropod sympodites with endopodites and exopodites in posterodorsal view; N, triangular pleotelson in dorsal view. Scale bars: A = 2 mm; B, G, I = 300 μm; C, E = 100 μm; D, F, H, L, M = 500 μm; J = 1 mm; K, N = 200 μm. 

Figure 10. SEM micrographs of male Porcellio virgatus (Budde-Lund, 1885) from San José, Gómez Farías, Tamaulipas, Mexico. A, Cephalothorax and pereon-tergites 1-7 in dorsal view; B, cephalothorax in dorsal view; C, right compound eye in dorsal view; D, cephalothorax, first antennae, clypeus, and labrum in ventral view; E, maxillipeds in ventral view; F, left first antenna with aesthetascs in ventral view; G, second antenna in medial view; H, flagellum of second antenna in medial view; I, posterior part of body showing pereon-tergite 7, pleon-tergites 3-5, and pleopods 1-5 in ventral view; J, left uropod with sympodite, endopodite, and exopodite in ventral view; K, posterior part of body showing pereon-tergite 7, pleon-tergites 1-5, and pleotelson in dorsal view; L, pleon-tergite 5, and triangular pleotelson in dorsal view. Scale bars: A, I, K = 2 mm; B, E, H, J = 500 μm; C = 200 μm; D, L = 300 μm; F = 100 μm; G = 1 mm.

Taxonomic summary

Type locality. New Orleans and Florida, USA (Budde-Lund, 1885).

Material examined. Nuevo León: Cola de Caballo, Santiago (25°26’12.18” N, 100°09’12.68” W), under stones, 06-05-2006, coll. A. Maeda, 2 females (5.4-6.8 mm) (CIB-49B); Parque Chipinque, Km 2, San Pedro Garza García (25°36’38.1” N, 100°21’20.8” W), under stones, 06-05-2006, coll. A. Maeda, 1 male (11.0 mm) (CIB-30B1). Tamaulipas: San José, Gómez Farías (23°05’23.1” N, 99°06’40.1” W), natural zone below stones, 30-07-2006, coll. A. Maeda, 3 males (6.8-9.6 mm, mean 7.9 ± 1.21 s.d. mm TL), 2 females (7.5-7.6 mm) (CIB-09B). 

Haplotypic identity. COI haplotypes were obtained from a specimen from Chipinque, Nuevo León (GenBank accession number OK376218), and a specimen from San José, Tamaulipas (GenBank accession number OK376217). The genetic distance between these haplotypes is 2.5% (Table 2). Prior to this study there was no record of DNA sequences for Porcellionides virgatus/Porcellio virgatus in GenBank. The Bayesian analysis placed these haplotypes in a well supported clade within the genus Porcellio with high support rather than with species in the genus Porcellionides (Fig. 11).

Distribution in Mexico. Santiago and San Pedro Garza García, Nuevo León, and San José, Tamaulipas. Mulaik (1960) reported Porcellio virgatus (as Porcellionides virgatus) from Villagrán, Tamaulipas, and Tepic, Nayarit. 

Table 2

Genetic p-distance (%) among COI sequences (623 bp) of Porcellionidae species used in the phylogenetic analysis.

Figure 11. Bayesian consensus tree of phylogenetic relationships of Porcellio virgatus (Budde-Lund, 1885) COI haplotypes from Nuevo León and Tamaulipas, Mexico, with selected GenBank haplotypes of Porcellionides and Porcellio species. GenBank sequences of palaemonids Periclimenes imperator Bruce, 1967 (= Zenopontonia rex [Kemp, 1922]) (DecaNet eds., 2024), and Periclimenes rathbunae Schmitt, 1924 were used as outgroup.

General distribution. Budde-Lund (1885), Van Name (1936), Mulaik and Mulaik (1942, 1943), and Mulaik (1960) reported the species (as Porcellionides virgatus)from the USA in Florida, Louisiana, Mississippi and Texas. Schultz (1982) reported Porcellio virgatus from North Carolina and Georgia, USA.

Remarks

The species has been cited as belonging to Metoponorthus Budde-Lund, 1879(Mulaik, 1960; Richardson, 1905), and to Porcellionides (the senior synonym, see Schmalfuss & Ferrara, 1978) (Jass & Klausmeier, 2004; Leistikow & Wägele, 1999; Souza-Kury, 2000; Van Name, 1936, 1940, 1942). Schmalfuss (2003) listed it as “Porcellionides” virgatus, indicating that the species does not belong to this genus. Schultz (1975, 1977, 1982) was the first carcinologist to recognize the species as Porcellio virgatus from Georgia and North Carolina, USA. The morphological similarity between Porcellio and Porcellionides has been, to date, an issue in recognizing them as separate genera (Dimitriou et al., 2018; Mattern, 2003; Mulaik, 1960; Schmalfuss & Ferrara, 1978; Van Name, 1936). Dimitriou et al. (2018) analysed 2 mitochondrial and 3 nuclear genes and found evidence against the monophyly of both Porcellionidae and Porcellio. The genetic distances they found were considerably high between Porcellionidae genera; e.g. for COI the variation was between 16.9-50.3% (Dimitriou et al., 2018). Mattern (2003) applied small subunit rRNA analyses to study the molecular phylogeny of Oniscidea and found that the genera Porcellio and Porcellionides are not monophyletic groups. He reported that Porcellio scaber Latreille, 1804 is more closely related to Porcellionides sexfaciatus Budde-Lund, 1885 than to Porcellio spinicornis Say, 1818, the latter branching as the sister group of Porcellionides pruinosus (Brandt, 1833). However, considering Mattern’s (2003: Fig. 2) molecular phylogenies, it can be concluded that Porcellio scaber and Porcellionides pruinosus represent 2 different lineages. Herein, we report the first COI haplotypes of the species obtained from specimens from Tamaulipas and Nuevo León. Our molecular analysis is not intended as a revision of the phylogenetic systematics of the group but rather to provide the first information on the molecular identity and the generic placement of these species. The results showed that the genetic distance of our haplotypes is larger when compared with haplotypes of Porcellionides (19.40 to 21.19%) than with haplotypes of Porcellio (15.57 to 19.58%) (Table 2). Besides, the Bayesian analysis placed our haplotypes in a clade with a specimen of Porcellio dilatatus Brandt in Brandt and Ratzeburg, 1831 haplotype with high nodal support, forming a group with Porcellio scaber (Fig. 11). Our specimens fit the diagnostic morphological features of the genus Porcellio proposed by Schmalfuss and Ferrara (1978): 2 pairs of pseudotracheae, unable to conglobate, pereon tergite 1 with caudally concave epimera, eyes with more than 20 ommatidea. The typical supra antennal line V-shape showed by most species of Porcellionides (Schmalfuss & Ferrara, 1978) is absent in our specimens (Fig. 10D) and in specimens examined by Van Name (1936, Fig. 135). Our specimens have the pereon-tergites 1-7 with a well marked dorsal transverse furrow (Fig. 10A), as figured for Porcellionides virgatus by Van Name (1936, Fig. 135). Mulaik (1960) recorded specimens of P. virgatus from Nayarit and Tamaulipas with 21 ommatidia and 10 mm TL. The specimens examined herein have 6.8-11.00 mm TL for males and 5.4-7.6 mm TL for females, with 19-30 ommatidia. The molecular data and morphological characteristics mentioned support Schultz’s assignation of the species as Porcellio virgatus.

Discussion

According to Schultz (1984), 3 Alloniscus species are recognized in North America, A. perconvexus Dana, 1854, known from Laguna Beach, California, USA to Vancouver Island, British Columbia, Canada; A. mirabilis recorded from Magdalena Bay, Mexico to the Pacific Grove, California; and A. thalassophilus Rioja, 1963 known from Isla de Ixtapan, Zihuatanejo, Guerrero, Mexico. The presence of Alloniscus in Mexico was first recorded by Mulaik (1960) who reported A. perconvexus based on specimens from Bahía Magdalena, Baja California Sur, and Isla Cedros and Bahía San Quintín, Baja California. Schultz (1984) mentioned that A. mirabilis and A. perconvexus are similar species, but can be separated by the uropod sympodite which is larger in A. mirabilis resulting in a broadened, extended posterolateral margin (Schultz, 1984, Fig. 3B, H) (Fig. 1L-N), and by the cephalothorax anterolateral lobes, which are well developed and extending far beyond the eyes in A. mirabilis (Schultz, 1984, Fig. 3D, E) (Fig. 1B, D), but small and well defined in A. perconvexus (Schultz, 1984, Fig. 1D). Schultz (1984) argued that Mulaik’s (1960) A. perconvexus illustrations actually represented A. mirabilis. We disagree with Schultz’s (1984) statement because the uropod sympodite illustrated by Mulaik (1960, Fig. 175) shows a straight posterolateral margin typical of A. perconvexus as also figured by Schultz (1984, Fig. 2B), while A. mirabilis has a wide-lobed posterolateral margin (Schultz, 1984, Fig. 3B, H) (Fig. 1L-N). Thus, we agree with Jass and Klausmeier (2004) that these 2 species are native to the northwest Mexican Pacific coast. As with all Mexican oniscideans, the populations of these similar species deserve more detailed morphological and genetic studies, including examination of material from their type localities.

The record of V. apacheus, formerly known only from Texas, USA, increases the number of species of Venezillo in Mexico from 18 (Jass & Klausmeier, 2004; Schmalfuss, 2003) to 19. Venezillo tanneri (Mulaik in Mulaik & Mulaik, 1942) also has been recorded in both USA (Texas) and Mexico (Nuevo León) (Mulaik & Mulaik, 1942; Schultz, 1965). In the southern USA Venezillo arizonicus (Mulaik in Mulaik & Mulaik, 1942) has also been recorded from Robles, Nogales, Olberg and Rock Springs, Arizona, and V. chamberlini (Mulaik in Mulaik & Mulaik, 1942) from Edinburg, Texas; both species are likely also inhabiting northern Mexico (Mulaik, 1960).

The genus Littorophiloscia is not well known in Mexico. Taiti and Ferrara (1986) recognized the presence of L. richardsonae from Baja California and L. tropicalis Taiti and Ferrara, 1986 from the Yucatán. In this study, L. richardsonae was recorded as co-occurring with A. mirabilis on a sand beach of Bahía Falsa in San Quintín, Baja California, sharing a habitat under marine macroalgae.

In Mexico, the genus Porcellio is now represented by the exotic P. laevis and P. scaber, and the putative natives P. scabriusculus Mulaik, 1960 (Jass & Klausmeier, 2004; Schmalfuss, 2003; Segura-Zarzosa et al., 2020), and P. virgatus (this study). Mulaik (1960) described and figured P. scabriusculus as very different from P. virgatus, with 9 ommatidea and spinous tubercles on the dorsal side of the cephalotorax, pereion terguites, and pleon terguites (except the pleotelson). Schmalfuss (2003) suggested that P. scabriusculus is probably not a member of the genus Porcellio. Mulaik (1960) mentioned his decision to keep Porcellionides saussurei (Dollfus, 1896) (a species known from Córdova, and San Luis Potosí, Mexico) in this genus until the Porcellio–Porcellionides complex is clarified. As mentioned above, these morphologically  similar genera are not monophyletic groups (Dimitriou et al., 2018; Mattern, 2003) and delimiting them is an important unsolved problem that limits the understanding of the systematics of the species involved. 

Acknowledgments

We thank an anonymous reviewer and Fernando Álvarez for helpful corrections to improve this article. Ariel A. Cruz Villacorta, Laboratorio de Microscopía Electrónica, CIB, provided expert assistance in the production of SEM images, and Gerardo Rafael Hernández García, Departamento de Extensión y Divulgación Científica, CIB edited the SEM plates and cladogram. Ilse E. Segura-Zarzosa received a doctoral fellowship from Consejo Nacional de Ciencia y Tecnología (No. 551538).

References

Ahyong, S. T., Lowry, J. K., Alonso, M., Bamber, R. N., Boxshall, G. A., Castro, P. et al. (2011). Subphylum Crustacea Brünnich, 1772. In Z. Q. Zhang (Ed.), Animal biodiversity: an outline of higher-level classification and survey of taxonomic richness. Zootaxa, 3148, 165–191. http://dx.doi.org/10.11646/zootaxa.3148.1.33 

Arcangeli, A. (1957). I generi Diploexochus, Venezillo, Paramardillo [sic] (crostacei isopodi terrestri). Bollettino dell’Istituto e Museo di Zoologia dell’Università di Torino (1955-56), 5, 101–142.

Boyko, C. B. (1997). Catalog of recent type specimens in the Department of Invertebrates, American Museum of Natural History. IV. Crustacea: Isopoda. American Museum Novitates, 3217, 1–39. 

Brandt, J. F., & Ratzeburg, J. C. T. (1831). Isopoda. Gleichfüßler. In Medizinische Zoologie oder getreue Darstellung und Beschreibung der Thiere die in der Arzneimittellehre in Betracht kommen, in systematischer Folge herausgegeben, 2, 70–84.

Brandt, J. F. (1833). Conspectus monographiae crustaceorum oniscidorum Latreillii. Bulletin de la Société Impériale des naturalistes de Moscou, 6, 171–193. 

Bruce, A. J. (1967). Notes on some Indo-Pacific Pontoniinae III-IX. Descriptions of some new genera and species from the western Indian Ocean and the South China Sea. Zoologische Verhandelingen, 87, 1–73.

Budde-Lund, G. (1879). Prospectus generum specierumque Crustaceorum Isopodum Terrestrium. Copenhagen. 10 pp.

Budde-Lund, G. (1885). Crustacea Isopoda terrestria per familias et genera et species descripta. Copenhagen: Nielsen & Lydiche. https://doi.org/10.5962/bhl.title.109769 

Dana, J. (1854). Catalogue and descriptions of Crustacea collected in California by Dr. John L. LeConte. Proceedings of the Academy of Natural Sciences of Philadelphia, 7, 175–177.

Darriba, D., Taboada, G. L., Doallo, R., & Posada, D. (2012). jModelTest2: more models, new heuristics and parallel computing. Nature Methods, 9, 772. https://doi.org/10.1038/nmeth.2109

DecaNet eds. (2024). DecaNet. Zenopontonia rex (Kemp, 1922). Accessed through: World Register of Marine Species. https://www.marinespecies.org/aphia.php?p=taxdetails&id=871470 

Dimitriou, A. C., Taiti, S., Schmalfuss, H., & Sfenthourakis, S. (2018). A molecular phylogeny of Porcellionidae (Isopoda, Oniscidea) reveals inconsistencies with present taxonomy. In E. Hornung, S. Taiti, & K. Szlavecz (Eds.), Isopods in a changing World. Zookeys, 801, 163–176. https://doi.org/10.3897/zookeys.801.23566 

Dollfus, A. (1896). Sur les Crustacés Isopodes terrestres du Mexique. Bulletin de la Societe Zoologique de France, 21, 46–49. https://doi.org/10.5962/bhl.part.18717 

Garthwaite, R. L., Hochberg, F. G., & Sassaman, C. (1985). The occurrence and distribution of terrestrial isopods (Oniscoidea) on Santa Cruz Island with preliminary data for the other California islands. Bulletin of Southern California Academy of Sciences, 84, 23–37.

Garthwaite, R. L., Lawson, R., & Sassaman, C. (1995). Population genetics of Armadillidium vulgare in Europe and North America. Crustacean Issues, 9, 145–199. 

Garthwaite, R., & Sassaman, C. (1985). Porcellionides floria, new species, from North America: provinciality in the cosmopolitan isopod Porcellionides pruinosus (Brandt). Journal of Crustacean Biology, 5, 539–555. https://doi.org/10.2307/1547923 

Guindon, S., & Gascuel, O. (2003). A simple, fast, and accurate algorithm to estimate large phylogenies by maximum likelihood. Systematic Biology, 52, 696–704. https://doi.org/10.1080/10635150390235520 

Hatch, M. (1947). The Chelifera and Isopoda of Washington and adjacent regions. University of Washington Publications in Biology, 10, 155–274. 

Holmes S. J., & Gay, M. E. (1909). Four new species of isopods from the coast of California. United Proceedings of the United States National Museum, 36, 375–379. https://doi.org/10.5479/si.00963801.36-1670.375 

Jass, J., & Klausmeier, B. (2004). Terrestrial isopod (Crustacea: Isopoda) atlas for Mexico. Milwaukee Public Museum Contributions in Biology and Geology, 100, 1–77.

Kemp, S. (1922). Notes on Crustacea Decapoda in the Indian Museum, XV. Pontoniinae. Records of the Indian Museum, 24, 113–288.

Kumar, S., Stecher, G., Li, M., Knyaz, C., & Tamura, K. (2018). MEGA X: Molecular evolutionary genetics across computing platforms. Molecular Biology and Evolution, 35, 1547–1549. https://doi:10.1093/molbev/msy096 

Latreille, P. A. (1802). Histoire naturelle générale et particulière, des crustacés et des insectes. Vol. 3. F. Dufart,Paris.
https://www.biodiversitylibrary.org/page/24884921

Latreille, P. A. (1804). Histoire naturelle generale et particulière, des crustacés et des insectes. Vol. 7. F. Dufart, Paris.
https://www.biodiversitylibrary.org/page/24882991 

Latreille, P. A. (1817). Les crustacés, les arachnides et les insectes. Vol. 3. Le règne animal distribuè d’après son organisation, pour servir de base a l’histoire naturelle des animaux et d’introduction a l’anatomie compare. Deterville, Paris.

Leistikow, A., & Wägele, J. W. (1999). Checklist of the terrestrial isopods of the new world (Crustacea, Isopoda, Oniscidea). Revista Brasileira de Zoologia, 16, 1–72. https://doi.org/10.1590/s0101-81751999000100001 

Mattern, D. (2003). New aspects in the phylogeny of the Oniscidea inferred from molecular data. In S. Sfenthourakis, P. B. De Auraujo, E. Hornung, H. Schmalfuss, S. Taiti, & K. Szlávecz (Eds.), The biology of terrestrial isopods V, Crustaceana Monographs, 2 (pp. 23–37). Koninklijke Brill NV, Leiden. 

Mulaik, S. (1960). Contribución al conocimiento de los isópodos terrestres de México (Isopoda, Oniscoidea). Revista de la Sociedad Mexicana de Historia Natural, 21, 79–292. https://doi.org/10.18268/bsgm2009v61n2a15  

Mulaik, S., & Mulaik, D. (1942). New species and records of American terrestrial isopods. Bulletin of the University of Utah, 32, 1–11. 

Mulaik, S., & Mulaik, D. (1943). New Texas terrestrial isopods with notes on other species. Bulletin of the University of Utah, 34, 1–15.

Richardson, H. (1905). A monograph on the isopods of North America. Bulletin of the United States National Museum, 54, 1–727. https://doi.org/10.5479/si.03629236.54.i 

Rioja, E. (1963) [1964]. Estudios carcinológicos XXXVI. Descripción y algunos datos morfológicos de Alloniscus thalassophilus n. sp. (Isopoda, Oniscidae) del piso supralitoral de las costas mexicanas del Pacífico. Anales del Instituto de Biología, Universidad Nacional Autónoma de México, 34, 285–306.

Rodríguez-Almaraz, G. A., Ortega-Vidales, V., & Treviño-Flores, J. A. (2014). Macrocrustáceos del Parque Nacional Cumbres de Monterrey, México: distribución y estado de conservación. Revista Mexicana de Biodiversidad, 85, 276–293. https://doi.org/10.7550/rmb.34967v

Ronquist, F., Teslenko, M., van der Mark, P., Ayres, D. L., Darling, A., Höhna, S. et al. (2012). MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Systematic Biology, 61, 539–542.
https://doi.org/10.1093/sysbio/sys029 

Say, T. (1818). An account of the Crustacea of the United States (concluded). Journal of the Academy of Natural Sciences of Philadelphia, 1, 423–441.

Schmalfuss, H. (1984). Eco-morphological strategies in terrestrial isopods. Symposia of the Zoological Society of London, 53, 49–63. 

Schmalfuss, H. (2003). World catalog of terrestrial isopods (Isopoda: Oniscidea). Stuttgarter Beiträge zur Naturkunde, Serie A, 654, 1–341. 

Schmalfuss, H., & Ferrara, F. (1978). Terrestrial isopods from West Africa. Part 2: Families Tylidae, Ligiidae, Trichoniscidae, Styloniscidae, Rhyscotidae, Halophilosciidae, Philosciidae, Platyarthridae, Trachelipidae, Porcellionidae, Armadillidiidae. Monitore Zoologico Italiano. N. S. Supplemento, XI, 15–97. https://doi.org/10.1080/03749444.1978.10736575 

Schmalfuss, H., & Wolf-Schwenninger, K. (2002). A bibliography of terrestrial isopods (Crustacea: Isopoda: Oniscidea). Stuttgarter Beiträge zur Naturkunde, Serie A (Biologie), 639, 1–120. https://doi.org/10.18476/sbna.v9.a3

Schmidt, C. (2002). Contribution to the phylogenetic system of the Crinocheta (Crustacea, Isopoda). Part 1. (Olibrinidae to Scyphacidae s. str.). Mitteilungenausdem Museum für Naturkunde in Berlin, Zoologische Reihe, 78, 275–352. https://doi.org/10.1002/mmnz.4850780207  

Schmidt, C. (2003). Contribution to the phylogenetic system of the Crinocheta (Crustacea, Isopoda). Part 2. (Oniscoidea to Armadillidiidae). Mitteilungenausdem Museum für Naturkunde in Berlin, Zoologische Reihe, 79, 3–179.
https://doi.org/10.1002/mmnz.20030790102  

Schmidt, C., & Leistikow, A. (2004). Catalogue of genera of the terrestrial Isopoda (Crustacea: Isopoda: Oniscidea). Steenstrupia, 28, 1–118.

Schmitt, W. L. (1924). The macruran, anomuran and stomatopod Crustacea. Bijdragen tot de kennis der fauna van Curaçao. Resultaten eener reis van Dr. C.J. van der Horst in 1920. Bijdragen tot de Dierkunde, 23, 61–81.

Schultz, G. A. (1965). Terrestrial isopods from caves and mines in Texas and northern Mexico with a description of Venezillo tanneri (Mulaik and Mulaik) allotype. The Texas Journal of Science, 17, 101–109.

Schultz, G. A. (1975). Terrestrial isopod crustaceans from coastal sites in Georgia. Bulletin of the Georgia Academy of Science, 34, 185–194.

Schultz, G. A. (1977). Terrestrial isopod crustaceans (Onisco-idea) from SI. Catherines Island, Georgia. Georgia Journal of Science, 35, 151–158.

Schultz, G. A. (1982). Terrestrial isopods (Crustacea: Isopoda: Oniscoidea) from North Carolina. Brimleyana, 8, 1–26. 

Schultz, G. A. (1984). Four species of Alloniscus Dana, 1854, from the west coast of North America and Hawaii (Isopoda, Oniscoidea). Crustaceana, 47, 149–167.

Segura-Zarzosa, I. E., Rodríguez-Almaraz, G. A., Obregón-Barboza, H., Murugan, G., Treviño-Flores, J. A., & Maeda-Martínez, A. M. (2020). New records of exotic species of Oniscidea (Crustacea: Isopoda) from northern Mexico. Revista Mexicana de Biodiversidad, 91, e913098. https://doi.org/10.22201/ib.20078706e.2020.91.3098

Segura-Zarzosa, I. E., Maeda-Martínez, A. M., Murugan, G. & Obregón-Barboza, H. (2022). Redescription of Venezillo stuckchensis (Crustacea: Oniscidea: Armadillidae), a terrestrial isopod from the Baja California Peninsula, Mexico. Revista Mexicana de Biodiversidad, 93, e934028. https://doi.org/10.22201/ib.20078706e.2022.93.4028 

Souza-Kury, L. (2000). Oniscidea. In J. E. L. Bousequets, E. G. Soriano, & N. Papavero (Eds.), Biodiversidad, taxonomía y biogeografía de artrópodos de México: hacia una síntesis de su conocimiento, Vol. II (pp. 239–246). Ciudad de México: Universidad Nacional Autónoma de México.

Stuxberg, A. (1875). Om Nord-Amerikas Oniscider. Ofversigt af kongliga Vetenskaps Akademiens Förhandlingar (Stockholm), 2, 43–64. 

Taiti, S., & Ferrara, F. (1986). Taxonomic revision of the genus Littorophiloscia Hatch, 1947 (Crustacea, Isopoda, Oniscidea) with description of six new species. Journal of Natural History, 20, 1347–1380. https://doi.org/10.1080/00222938600770911 

Thompson, J. D., Gibson, T. J., Plewniak, F., Jeanmougin, F., & Higgins, D. G. (1997). The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Research, 25, 4876–4882. https://doi.org/10.1093/nar/25.24.4876  

Tizol-Correa, R., Maeda-Martínez A. M., Weekers, P. H. H., Torrentera, L., & Murugan, G. (2009). Biodiversity of the brine shrimp Artemia from tropical salterns in southern México and Cuba. Current Science, 96, 81–87. https://doi.org/10.1651/s-2691.1 

Treviño-Flores, J. A., & Rodríguez-Almaraz, G. (2012). Primeros registros de Porcellio laevis y Porcellio scaber (Crustacea: Oniscidea) del noreste de México. In M. López-Mejía, & L. M. Mejía-Ortiz (Eds.), La carcinología en México: el legado del Dr. Alejandro Villalobos 30 años después (pp. 13–21). Universidad de Quintana Roo. Cozumel, Quintana Roo, Mexico.

Van Name, W. G. (1936). The American land and freshwater isopod Crustacea. Bulletin of the American Museum of Natural History, LXXI, 1–535. 

Van Name, W. G. (1940). A supplement to the American land and fresh-water isopod Crustacea. Bulletin of the American Museum of Natural History, LXXVII, 109–142.

Van Name, W. G. (1942). A second supplement to the American land and fresh-water isopod Crustacea. Bulletin of the American Museum of Natural History, LVIII, 299–329.

Verhoeff, K. W. (1908). Uber Isopoden. 12. Aufsatz. Nene oniscoidea aus Mittel und Südeuropa und zur Klärung einiger bekannter Formen. Archiv für Naturgeschichte, 74, 163–198. 

Verhoeff, K. W. (1928). Über einige Isopoden der Zoologischen Staatssammlung in München. Zoologischer Anzeiger, 76, 25–36, 113–123.

Alejandro Valdez-Mondragón *, Samuel Nolasco-Garduño

Centro de Investigaciones Biológicas del Noroeste, S.C., Programa Académico de Planeación Ambiental y Conservación, Laboratorio de Aracnología, Km. 1 Carretera a San Juan de La Costa “El Comitán”, 23205 La Paz, Baja California Sur, Mexico

*Corresponding author: lat_mactans@yahoo.com.mx (A. Valdez-Mondragón)

Received: 11 March 2024; accepted: 29 October 2024

Abstract

We describe the morphological and genetic variation in Ixchela azteca Valdez-Mondragón & Francke (Pholcidae) from central Mexico, based on morphological and DNA barcoding evidence. Species delimitation employing the barcoding locus included 3 methods under the corrected genetic p-distances neighbor-joining (NJ) criteria: 1) Assemble Species by Automatic Partitioning (ASAP), 2) General Mixed Yule Coalescent model (GMYC), and 3) Bayesian Poisson Tree Processes (bPTP). The genetic analyses found a genetic p-distance of 3.5% between 2 populations of I. azteca. The molecular methods and morphology were not congruent in delimiting and recognizing 2 possible different species. Instead, 2 allopatric populations of I. azteca are recognized. Also, an updated taxonomic revision of I. azteca is included. In conclusion, the incongruence between the molecular delimitation methods for species delimitation and the morphological, genetic, ecological, and biogeographic evidence maintain I. azteca as a widespread species across central Mexico, with allopatric populations in temperate and semiarid regions.

Keywords: Species delimitation; Integrative taxonomy; CO1; Ecology; Biogeographic provinces

© 2025 Universidad Nacional Autónoma de México, Instituto de Biología. Este es un artículo Open Access bajo la licencia CC BY-NC-ND

(http://creativecommons.org/licenses/by-nc-nd/4.0/).

Ixchela azteca (Araneae: Pholcidae), una especie de araña ampliamente distribuida en el centro de México: ¿diversidad subestimada o variación morfológica y genética?

Resumen

Describimos la variación morfológica y genética en Ixchela azteca Valdez-Mondragón y Francke (Pholcidae) del centro de México, basado en evidencia morfológica y código de barras de DNA. La delimitación de especies con el locus de código de barras incluyó 3 métodos bajo el criterio de distancias genéticas p corregidas “neighbor-joining” (NJ): 1) “Assemble Species by Automatic Partitioning” (ASAP), 2) “General Mixed Yule Coalescent model” (GMYC), and 3) “Bayesian Poisson Tree Processes” (bPTP). Los análisis genéticos entre 2 poblaciones de I. azteca encontraron una distancia genética p de 3.5%. Los métodos moleculares y morfología no fueron congruentes para delimitar y reconocer 2 posibles especies, pero sí se reconocen 2 poblaciones alopátricas de I. azteca. Adicionalmente, se incluye una revisión taxonómica actualizada de I. azteca.En conclusión, la incongruencia de los métodos de delimitación molecular para la delimitación de especies en combinación con la evidencia morfológica, genética, ecológica y biogeográfica, corrobora a I. azteca como una especie ampliamente extendida en la región central de México, con poblaciones alopátricas en zonas templadas, pero también en regiones semiáridas.

Palabras clave: Delimitación de especies; Taxonomía integrativa; CO1; Ecología; Provincias biogeográficas

Introduction

The spider family Pholcidae is the ninth-largest spider family in the world and the most diverse within the Synspermiata clade (WSC, 2025). Commonly known as cellar spiders or daddy long-legs spiders, Pholcidae is currently composed of 97 genera and 1,901 species (WSC, 2025). This family is among the most diverse and abundant web-building spiders in the world, mainly occurring in tropical and subtropical forests and with a high number of synanthropic and troglobite species (Huber, 2000, 2011, 2018; Valdez-Mondragón, 2010). 

The genus Ixchela Huber, 2000 (subfamily Modisiminae) currently contains 22 described species (Valdez-Mondragón, 2013, 2020; Valdez-Mondragón & Francke, 2015; WSC, 2025). The genus is composed of relatively large pholcid spiders mainly distributed in Mexico and Central America. These spiders inhabit temperate climate zones in pine, oak, or pine-oak forests at moderately high elevations (1,000-2,950 m) (Valdez-Mondragón, 2013, 2020; Valdez-Mondragón & Francke, 2015). The monophyly of the genus is supported based on morphological and molecular evidence (CO1+16S), with the genus’ divergence time estimated to have occurred in the late Miocene (Valdez-Mondragón & Francke, 2015).

Although pholcid spiders generally have a relatively uniform and conservative somatic morphology, their genitalia (palps and epigyna) and secondary sexual characters (male chelicerae) are highly informative for morphological identification at species level. These morphological features are a source of important and robust evidence and are commonly used for species delimitation in most groups of spiders (Huber, 2003). Spider genitalia usually show little intraspecific, but conspicuous interspecific variation (Eberhard, 1985; Eberhard et al., 1998). However, in certain groups with highly conservative morphology, or species with a wide geographic distribution, species delimitation and identification require additional diagnostic features to differentiate underestimated species diversity. In contrast, other species of spiders have wide geographic variation and even show intrasexual dimorphism within males or females. In the last 4 decades, research has focused on understanding the variation among individuals, leading sexual variation to be understood as the differences exhibited not only between males and females but also among individuals of the same sex within a species (Andersson, 1994; Barraclough, 1995; Gross, 1996; Huber, 2021).

The mitochondrial cytochrome c oxidase subunit 1 (CO1) DNA barcode gene has emerged as the standard animal barcoding marker for many taxa, including spiders (Astrin et al., 2006; Correa-Ramírez et al., 2010; Graham et al., 2015; Hamilton et al., 2011, 2014, 2016; Hebert, Cywinska et al., 2003; Hebert, Ratnasingham et al., 2003; Hebert et al., 2004; Ortiz & Francke, 2016; Planas & Ribera, 2015; Nolasco & Valdez-Mondragón, 2022a; Tahami et al., 2017). The integrative taxonomy approach is generally accepted as being more effective in the diagnosis, delimitation, and description of species or even to understand the variation in different populations within species.

Ixchela azteca Valdez-Mondragón & Francke, 2015from central Mexico and Ixchela abernathyi (Gertsch, 1971) from northeast Mexico represent the species with the widest distributions in comparison with other species of the genus, mainly in temperate regions. However, Valdez-Mondragón and Francke (2015) reported body size variation for populations of I. azteca from temperate regions, bringing into question if this is merely geographic variation in the same species or is evidence of the presence of different species not yet described. Because of that, morphological and molecular evidence are needed to test if I. azteca has a widespread distribution in temperate and semiarid habitats or represents different species.

In this study, we described based on morphological evidence and DNA barcoding (CO1) the variation of different populations of I. azteca including 3 populations from temperate regions of central Mexico and a population from a semiarid habitat in Guanajuato state, Mexico. In addition, an updated taxonomic revision of I. azteca is included herein, including new records for the species.

Materials and methods

The specimens used in this study are deposited in the Colección Nacional de Arácnidos (CNAN), Instituto de Biología, Universidad Nacional Autónoma de México (IBUNAM), Mexico City; and the Colección Aracnológica (CARCIB), Centro de Investigaciones Biológicas del Noroeste, CIB, La Paz, Baja California Sur, Mexico. The specimens from Guanajuato, Mexico are deposited at the CARCIB. Morphological structures, such as female epigyna and male palps, were dissected in ethanol (80%) and cleared in potassium hydroxide (KOH-10%). Habitus, chelicerae, palps, and epigyna were placed on 96% gel alcohol to facilitate positioning and covered with a thin layer of liquid ethanol (80%) to minimize diffraction during photography. The analyzed morphological structures were female epigyna and male palps, dorsal and ventral habitus, and male chelicerae.

A Zeiss Axiocam 506 color camera attached to a Zeiss AXIO Zoom.V16 stereoscope was used to photograph specimen structures. For measurements, the specimens were observed using a Zeiss Discovery V8 stereoscope. All measurements are in millimeters (mm). The distribution map was generated using QGIS v. 2.18.17 “Las Palmas”. Photographs and a map were edited using Adobe Photoshop CS6. Morphological abbreviations follow Valdez-Mondragón (2013, 2020) and Valdez-Mondragón and Francke (2015). Abbreviations: ALE, anterior lateral eyes; AME, anterior median eyes; E, embolus; FAC, frontal apophysis of chelicerae; MSE, median septum of epigynum; PAB, prolateroventral apophysis of bulb; PLE, posterior lateral eyes; PME, posterior median eyes; PP, pore plates; PR, procursus; SAC, sclerotized apophysis of chelicerae; VPP, ventrobasal protuberance of procursus.

The DNA barcoding locus was obtained from a total of 59 specimens belonging to 19 species of Ixchela previously described by Valdez-Mondragón (2013, 2020) and Valdez-Mondragón and Francke (2015), and combined with available CO1 sequences, including I. azteca and a single outgroup (Physocyclus dugesi Simon) for the genetic p-distances analysis. Species used in the molecular analyses are listed in Supplementary material (S1). Physocyclus dugesi (subfamily Arteminae) was used only to root the tree. The comparison among sequences of I. azteca and the rest of the species’ sequences was to analyze the intra and interspecific genetic variation. For samples of I. azteca,1 sequence (CNAN Ara-0160) from Km 46 highway Toluca-Valle de Bravo, Estado de México (type locality of the species); 2 sequences (CNAN Ara-0158 and Ara-0159) from Cueva del Fraile, Gustavo A. Madero, Mexico City; and 2 sequences (CNAN Ara-0169 and Ara-0170) from Cueva del Diablo, Tepoztlán, Morelos were used for the analysis. Specimens used in this study, including GenBank accession numbers and locality information are listed in S1.

DNA was isolated from legs (1 complete leg per sample), and extractions were performed with a Qiagen DNeasy Tissue Kit following a modified protocol described in Valdez-Mondragón and Francke (2015) and Valdez-Mondragón (2020). DNA fragments corresponding to 658 bp of the mitochondrial cytochrome c oxidase subunit 1 (CO1) gene were amplified using the same primers (LCO1490 and HCO2198) as in Valdez-Mondragón and Francke (2015) and Valdez-Mondragón (2020). Amplifications were carried out using a Veriti 96 Well Thermal Cycler (Applied Biosystems) following the modified protocol of Valdez-Mondragón and Francke (2015) and Valdez-Mondragón (2020). PCR products were checked to analyze length and purity on 1% agarose gels with a marker of 100 bp and purified directly from the QIAquick PCR Purification kit of Qiagen. DNA extraction and PCR amplification were performed at Laboratorio de Biología Molecular, Laboratorio Regional de Biodiversidad y Cultivo de Tejidos Vegetales (LBCTV), Instituto de Biología, Universidad Nacional Autónoma de México (IB-UNAM), Tlaxcala. Bi-directional Sanger sequencing was performed at the Laboratorio de Secuenciación Genómica de la Biodiversidad y la Salud (LANABIO), IB-UNAM, Mexico City. The generated sequences were saved in FASTA format. All newly generated sequences were deposited in GenBank with the accession numbers OR106002-OR106006 (see S1).

Sequences were aligned using the default gap opening penalty of 1.53 in the online MAFFT platform (multiple alignment using fast fourier transform), version 7 (Katoh & Toh, 2008) using the following alignment strategy: auto (FFT-NS-2, FFTNS-I, or L-INS-i; depending on data size). Inspection and editing of sequences and alignments was done using GENEIOUS version 10.2.6 (Kearse et al., 2012) and BioEdit version 7.0.5.3 (Hall, 1999). The matrix obtained from the multiple sequence alignments was then used for subsequent analyses.

To assess species delimitation with the barcoding locus, we first conducted a phenetic analysis with the neighbor joining (NJ) method to visualize the uncorrected p-distances using the program MEGA v. 7.0 (Tamura et al.,2007). We subsequently used the reconstructed NJ tree to carry out analyses with the following 3 methods: 1) Assemble Species by Automatic Partitioning (ASAP) (Puillandre et al., 2021): we ran this analysis on the online platform (https://bioinfo.mnhn.fr/abi/public/asap/) with the parameters suggested by Nolasco and Valdez-Mondragón (2022a); 2) General Mixed Yule Coalescent model (GMYC) (Pons et al., 2006): this analysis was performed using the GMYC web server (https://species.h-its.org/gmyc/); and 3) Bayesian Poisson Tree Processes (bPTP) (Kapli et al., 2017; Zhang et al., 2013): this method was conducted using the web server (https://species.h-its.org/ptp/).

p-distances under neighbor joining (NJ) 

The genetic distances tree was reconstructed with MEGA v. 7.0 (Tamura et al.,2007), using the following commands: No. replicates = 1,000, bootstrap support values = 1,000 (significant values ≥ 50%), substitution type = nucleotide, model = p-distance, substitution to include = d: transitions + transversions, rates among sites = gamma distributed with invariant sites (G+I), missing data treatment = pairwise deletion.

Automatic Barcode Gap Discovery (ABGD): this method is implemented to find gaps in genetic divergence, considering that the intraspecific genetic variation is smaller than the interspecific divergences. First, the method generates a prior data partition of the data into putative species (initial partitions, IP). Then, these initial partitions are recursively partitioned until there is no further partitioning of the data (recursive partitions, RP). ABGD analyses were carried out in the online platform (https://bioinfo.mnhn.fr/abi/public/abgd/) using the following options: K2P distances non-corrected, pmin = 0.001, pmax = 0.1, steps = 10, relative gap width (X) = 1, nb bins = 20. Assemble Species by Automatic Partitioning (ASAP): this is an ascending hierarchical clustering method, merging sequences into groups that are successively further merged until all sequences form a single group. A partition is the equivalent to each sequence merge step, then, the software analyzes all partitions and scores the most probable groups on the tree (Puillandre et al., 2021). ASAP analyses were run in the online platform (https://bioinfo.mnhn.fr/abi/public/asap/) using Kimura (K80) distance matrices and configurated with following parameters: substitution model = p-distances, probability = 0.01, best scores = 10, fixed seed value = -. General Mixed Yule Coalescent (GMYC): this species delimitation method applies single (Pons et al., 2006) or multiple (Monaghan et al., 2009) time thresholds to delimit species in a maximum likelihood context, using ultrametric trees (Ortiz & Francke 2016). To generate the ultrametric tree, phylogenetic analyses were carried out in BEAUti and BEAST v.1.10.4 software (Drummond et al., 2012) using a coalescent (constant population) tree prior. Independent log normal clock uncorrelated was applied with its respective evolutionary model and substitution rates. The models of sequence evolution were selected using jModelTest version 2.1.10 using the Akaike information criterion (AIC) (Posada & Buckley, 2004). The models selected for CO1for each partition block were GTR+G+I (1^st and 2^nd codon positions) and GTR+G (3^rd codon position). For the analyses, 40 million iterations were run 5 times, independently. The program Tracer 1.6 (Rambaut & Drummond, 2003) was used to evaluate the convergence values. Tree annotator v. 2.6.0 (BEAST package) was used to construct maximum clade credibility trees, after discarding the first 25% of generations (“burnin”) of the 5 independent runs. Finally, the GMYC method was implemented in the web service (https://species.h-its.org/gmyc/), which uses the original R implementation of the GMYC model (Fujiwasa & Barraclough, 2013). Bayesian Poisson Tree Processes (bPTP): this species delimitation method is similar to GMYC, but it does not use an ultrametric tree as input. This is because the models of speciation rate are implemented directly using the numbers of substitutions calculated from the branch lengths. The Bayesian and maximum likelihood variants were carried out on the online version (https://species.h-its.org/ptp/), with the following options: rooted tree, MCMC = 1,000,000, thinning = 100, burn-in = 0.1, seed = 123. The trees obtained in all analyses were edited with iTOL online version (https://itol.embl.de/) (Letunic & Bork 2021) and Photoshop CS6. We used the congruence integration criteria to delimit species, which is based on the correspondence among the different molecular methods to generate high support for a species hypothesis (DeSalle et al., 2005, Hamilton et al. 2011, Navarro-Rodríguez & Valdez-Mondragón, 2020, 2024; Nolasco & Valdez-Mondragón, 2022a; Valdez-Mondragón, 2020).

The approaches for DNA barcoding tree-based delimitation explicitly use the phylogenetic species concept, where the analysis recognizes a monophyletic cluster by searching differential intra and inter-specific branching patterns (Ortiz & Francke, 2016). 

Results

According to the morphology of primary (male palps and female epigyna) and secondary (male chelicerae) sexual structures, the populations of Ixchela azteca showed geographical variation in morphological traits (Fig. 1). The males of the 3 populations have the same shape of the palp (Figs. 1, 8, 9; Valdez-Mondragón & Francke, 2015: Figs. 38, 39); however, there are differences in the shape of the male chelicerae (Figs 1, 4, 6, 7; Valdez-Mondragón & Francke, 2015: Figs. 36, 37, 40) and female epigyna (Figs. 10-13; Valdez-Mondragón & Francke, 2015: Fig. 36). The populations from the temperate region of Cueva del Fraile, Mexico City (CNAN Ara-0158 and Ara-0159), the population from the type locality (Estado de México, CNAN Ara-160), and the population from Cueva de Diablo, Morelos (CNAN Ara-0169 and Ara-0170) have the frontal apophysis of chelicerae (FAC) shorter (Valdez-Mondragón & Francke, 2015: Fig. 40) than the males from the population of the dry region of Guanajuato, which have a longer and conical FAC (Fig. 7). The female epigynum from Guanajuato is square-shaped in ventral view (Fig. 10), whereas in the other 3 populations from temperate region of central Mexico (Mexico City, Estado de México and Morelos) it is more trapezoidal (Valdez-Mondragón & Francke, 2015: Fig. 42). In frontal view, the conical projection in the population from Guanajuato is smaller (Fig. 12) than the one observed in females from central Mexico (Valdez-Mondragón & Francke, 2015: Fig. 44). Finally, females from the 3 populations of the temperate region of central Mexico have a longer and straighter epigynum in lateral view (Valdez-Mondragón & Francke, 2015: Fig. 41) than the population from the dry region of Guanajuato, which have a shorter and curved epigynum (Fig. 13). The dorsal pattern of the carapace is similar between populations of I. azteca,with lateral spots on the carapace and a dark region along the fovea (Fig. 5; Valdez-Mondragón & Francke, 2015: Fig. 35). In comparison with the rest of the species of the genus, I. azteca so far is the only species with morphological variation in the primary (male palps and female epigyna) and secondary (male chelicerae) sexual structures.

The CO1 matrix included 59 terminals (including 1 outgroup) and 601 aligned positions. The uncorrected CO1 average distances between the populations of I. azteca from temperate regions and the dry region of Guanajuato was 3.5% (Table 1), whereas the average genetic distances among the remaining species of Ixchela was 12.1% (Table 1). Bootstrap support for the populations from Guanajuato was 100% (Fig. 1, red branch), whereas the group with the populations from temperate regions in central Mexico had a bootstrap support of 59% (Fig. 1, green branch). 

Molecular methods for species delimitation

In the maximum likelihood (ML) phylogram obtained with CO1 (Fig. 2), most of the species assignments based on the current taxonomy were congruent with all species delimitation methods, except for the specimens assigned to I. azteca (Fig. 2). The GMYC model recovered a total of 21 species, and the 2 clades of I. azteca (Fig. 2, green and red branches) were recovered as a single species exclusive of I. azteca CNAN-Ara0169 and Ara0170, which were recognized as a different species (Fig. 2). The ASAP method recognized a total of 18 species, recovering both clades of I. azteca as a single species (Fig. 2). Finally, the bPTP method recovered a total of 25 species of Ixchela under both variants (BI and ML); however, similar to the GMYC analysis, the specimens CNAN-Ara0169 and Ara0170 were recognized as a different species from I. azteca (Fig. 2). The first clade of I. azteca (Fig. 2, green branch)did not receive significant bootstrap support (58%), a result also observed in the NJ analysis, whereas the clade from Guanajuato (Fig. 2, red branch) was supported by a high bootstrap value (100%). The I. azteca samples CNAN-Ara0169 and Ara0170 had a high bootstrap support of 99% for their sister relationship (Fig. 2).

Family Pholcidae C. L. Koch, 1850

Genus Ixchela Huber, 2000

Diagnosis and description. For an updated diagnosis and description of the genus, see Valdez-Mondragón (2013, 2020) and Valdez-Mondragón and Francke (2015).

Taxonomic summary

Type species. Coryssocnemis furcula F. O. Pickard-Cambridge, 1902, by original designation of Huber (2000). Type locality: female holotype from Tecpam, región de los Altos (cerro Tecpam, Departmento Chimaltenango), Guatemala, around 2,300 m, Otto Stoll. Godman & Salvin Coll., in BMNH (F. O. Pickard-Cambridge, 1902; Huber, 1998).

Composition. Twenty-two species. See the World Spider Catalog (2025) for the specific list of species. 

Distribution. Ixchela is widely distributed from northeastern Mexico to Nicaragua. Although specimens from Nicaragua were not examined in this work, Huber (2000) examined 1 male of an undescribed species from this country deposited in the Museo Entomológico Nicaraguense (León, Nicaragua). 

Ixchela azteca Valdez-Mondragón & Francke, 2015

I. azteca Valdez-Mondragón & Francke, 2015: 29, f. 1-6, 18-44 (Description ♂, ♀).

Figs. 3-13, 16-19 

Diagnosis and description. See Valdez-Mondragón and Francke (2015).

Taxonomic summary

Type material. Mexico: Estado de México: ♂ holotype (CNAN T0763) [26 August 2011; A. Valdez, J. Mendoza, D. Barrales, R. Monjaraz, E. Miranda Cols.] from Km 46 highway Toluca-Valle de Bravo (19°15’21.6” N, 100°04’00.8” W; 2,315 m asl) (examined). Paratypes: 1 f# (CNAN T0764); 1 ♀, 3 juv. (CNAN T0765), same data as holotype (examined). See Valdez-Mondragón and Francke (2015) for the complete list of material examined from Estado de México, Guerrero, Mexico City, Michoacán, and Morelos.

Table 1

Average genetic p-distance under NJ among the different species of Ixchela analyzed with CO1. The value in bold indicates the average p-distance between the populations from temperate regions (A) and the population from the dry region in Guanajuato (B) of I. azteca (3.5%) (bold). The average genetic p-distance under NJ between the species of Ixchela was 12.1%.

Figure 1. Neighbor-joining tree constructed with p-distances of the CO1 barcode sequences from different specimens and species of Ixchela. Branch colors indicate species, red and green branches and asterisks represent populations of Ixchela azteca. Male chelicera, male palp, and female epigyna are shown to illustrate the variation at species level and within I. azteca. Numbers on branches indicate bootstrap support values (> 50%).

Figure 2. Maximum likelihood tree of Ixchela constructed with CO1. Bar colors represent putative species in the tree and in the columns, which represent the different species delimitation methods. Red and green branches and asterisks represent populations of Ixchela azteca. Numbers below the columns represent the species recovered in each species delimitation method (not considering Physocyclus dugesi). Numbers above branches represent bootstrap support values for ML (> 50%). Column abbreviations: morphology (M); GMYC with single (SN) and multi (MT) thresholds; ASAP; bPTP with maximum likelihood (ML) and Bayesian inference (BI) variants.

Figures 3-9. Ixchela azteca Valdez-Mondragón & Francke, 2015 from Cortazar, Guanajuato, Mexico. Male: 3-4, habitus, lateral and dorsal views, respectively; 5, carapace and chelicerae, frontal view; 6-7, chelicerae, frontal and left lateral views, respectively; 8-9, left palp, prolateral and retrolateral views, respectively. E: Embolus, PAB: prolateroventral apophysis of bulb, PR: procursus, SAC: sclerotized apophysis of chelicerae, VPP: ventrobasal protuberance of procursus. Scale bars: Figs. 6, 7 = 0.5 mm, Figs. 5, 8, 9 = 1 mm, Figs 3, 4 = 2 mm.

New material examined. Mexico: Guanajuato: 1 ♂, 2 ♀, 2 juv. (CARCIB) [14 October 2021; A. Valdez, A. Juárez, L. Cabrera, S. Nolasco Cols.] from 7.5 km southeast of Cortazar (20°25’31.1” N, 100°54’51.8” W; 2,160 m asl), Municipality Cortazar (night collecting). 

Morphological variation. Male specimens from Cueva del Diablo (Morelos, Mexico) and from Gruta de Tziranda were notably smaller than specimens from the type locality and the other localities, including the new specimens from Guanajuato. Female specimens from Grutas de Tziranda (Michoacán, Mexico) and road to Cueva del Fraile (Mexico City, Mexico) were notably smaller than specimens from the other localities, including the new specimens from Guanajuato.

Figures 10-13. Ixchela azteca Valdez-Mondragón & Francke, 2015 from Cortazar, Guanajuato, Mexico. Female epigynum: 10, ventral view; 11, dorsal view; 12, frontal view; 13, left lateral view. MSE: Median septum of epigynum, PP: pore plates. Scale bars: 0.5 mm.

Specimens from Cueva del Diablo have paler coloration on carapace and legs than specimens from other localities. The opisthosomal coloration can vary from grey, pale grey, blue, or pale blue, potentially related to the prey consumed. Males:Cueva del Diablo (Morelos, Mexico) (n= 4), tibia I: 10.87-12.50 (x= 11.74). Cueva del Diablo, La Peña (Estado de México, Mexico) (n= 1): tibia I: 18.12. Cueva de Peña Blanca (Estado de México, Mexico) (n= 2), tibia I: 13.75, 14.00. Gruta de Tziranda (Michoacán, Mexico) (n= 1): tibia I: 11.00. Road to Cueva del Fraile and Cueva del Fraile, respectively (Mexico City, Mexico) (n= 2): tibia I: 9.50, 12.25. 7.5 km southeast of Cortazar (Guanajuato, Mexico) (n= 1): tibia I: 13.63. Females:Cueva del Diablo (Morelos, Mexico) (n= 7): tibia I: 7.50-16.00 (x= 12.77). Km 34 highway Toluca-Zitácuaro (Estado de México, Mexico) (n= 3): tibia I: 6.60-12.5 (x = 9.50). 5 km W of Casahuates (Guerrero, Mexico) (n= 3): tibia I: 9.37-15.25 (x= 11.95). El Naranjo (Michoacán, Mexico) (n= 2): tibia I: 9.50, 11.75. 7 km SE of Ciudad Hidalgo (Michoacán, Mexico) (n= 2): tibia I: 9.7, 11.37. Gruta de Tziranda (Michoacán, Mexico) (n= 4): tibia I: 7.10-10.2 (x= 8.70). Road to Cueva del Fraile (Mexico City, Mexico) (n= 2): tibia I: 10.12, 11.87. 7.5 km southeast of Cortazar (Guanajuato, Mexico) (n = 2): tibia I: 9.87, 11.87 (x = 10.87).

Figures 14-19. Habitat and live specimens of I. azteca. 14-15, Thorny scrub forest at 2,160 m asl. from 7.5 km southeast of Cortazar, Municipality Cortazar, Guanajuato, Mexico (red arrow indicates the microhabitat where the specimens were collected); 16-17, live male specimens; 18-19, live female specimens.

Natural history. Valdez-Mondragón and Francke (2015) reported that the specimens from Estado de México and Guerrero were collected on their sheet webs in oak-pine and pine forests, inside cavities on walls along road-cuts in wet and shaded areas covered with roots. The male collected in the Instituto de Biología, UNAM was walking on a wall inside a classroom. The specimens from Cueva del Fraile, Mexico City, and Gruta de Tziranda, Michoacán were collected on their sheet webs inside the caves, close to the walls. Specimens collected outside the Cueva del Fraile were among boulders in shady moist areas, whereas specimens from Cueva del Diablo, Morelos were collected in the cave entrance and inside the cave, where humidity was c.70% and temperatures cold. These specimens were collected on their sheet webs, and it was very common to find prey remains in their webs, mainly large leafcutter ants of the genus Atta (subfamily Myrmicinae). The specimens from Cortazar, Guanajuato, were collected in a disturbed thorny scrub forest (Figs. 14, 15) with columnar cacti and nopales (Opuntia sp.). These specimens were observed on their sheet webs built among walls of big rocks (Figs. 14, 15 red arrow, 16-19). 

Figure 20. Distribution records of I. azteca in the Chihuahuan Desert biogeographic province (Southern Semiarid Elevations ecoregion) and in the Transmexican Volcanic Belt biogeographic province (Temperate Mountains ecoregion). Map by Mayra Cortez Roldán.

Distribution. Mexico: Estado de México, Guanajuato, Guerrero, Mexico City, Michoacán, Morelos (Fig. 20).

Discussion

The evolutionary aspects of morphological variation, including dimorphism and polymorphism within and among sexes, has been widely studied in several animal groups. Also, assessing species boundaries is a central debate in modern systematics, particularly in cases where morphology presents wide variations or lacks information at deep levels. The use of DNA barcoding approaches has been applied in modern systematics in 2 different ways: first, to distinguish among described species (equivalent to species identification or species diagnosis), and second, to discover new species (incorporating species delimitation methods and species descriptions) (DeSalle et al., 2005). In this way, another taxonomic task is to establish the limits between species or identify those species with wide morphological and genetic variation among populations.

Regarding the genus Ixchela,molecular methods for species delimitation have been used in parallel with morphological features to obtain the most robust evidence for species delimitation and diagnoses. Valdez-Mondragón (2020) described I. panchovillai and I. zapatai using 4 molecular species delimitation methods and morphology. All molecular methods were congruent with the morphology of primary and secondary sexual features (male palps, male chelicerae, and female epigyna). However, in the case of I. azteca,despite the population from Guanajuato (Fig. 2, green branch) being genetically divergent from the populations from central Mexico (Fig. 2, red branch) by a p-distance value of 3.5% (Table 1), the molecular species delimitation methods were not congruent with morphology, recovering the 2 clades as a single species. Previous studies on species delimitation in pholcid spiders have usually found congruence between species limits hypothesized a prioribased on morphology and those recovered using mitochondrial markers (Nolasco & Valdez-Mondragón, 2022a; Valdez-Mondragón, 2020). However, similar to I. azteca, morphological variation of sexual structures within the same species has been previously reported in other pholcid spiders. Huber and Pérez-González (2001) reported a case of morphological variation in the female genitalia of 3 species in the genus Ciboneya Huber & Pérez-González, 2001, which even have different female morphs in the same population. Valdez-Mondragón (2010) described discontinuous interspecific variation in the epigyne shape in females of Physocyclus enaulus Crosby, 1926 with 3 distinct morphotypes. Such variation has been observed in insular pholcids as Huber and Acurio (2021) reported for Aymaria conica (Banks, 1902), with wide morphological variation in the female´s genitalia. In the case of I. azteca, the epigyna of the population from Guanajuato is wider and more curved than in the populations from central Mexico, which are longer and less curved. Males of the different I. azteca populations also showmorphological variation, mainly in the chelicerae, a robust and very important feature at species level in the family Pholcidae. Males from Guanajuato have longer apophyses than males from central Mexico. 

Pholcid spiders show a great variety of morphological features that allow for delimitation and diagnosis to the species level in most cases. Primary sexual structures such as male palps and female genitalia, as well as secondary sexual structures such as male chelicerae, are important and useful characters for identifying and diagnosing pholcids species (Huber et al., 2018; Nolasco & Valdez-Mondragón, 2022b; Valdez-Mondragón, 2020). However, as Huber and Pérez-González (2001) suggest, it is possible that taxonomists are strongly biased against discovering genitalic variation such as dimorphisms or even polymorphisms, simply because the genitalia have traditionally been used with an overwhelming priority for species delimitation and identification. This is why taxonomic descriptions should (ideally) be based on a large series of specimens to analyze both intra- and inter-specific morphological and genetic variation. This practice also allows us to identify cases of morpho-clines, dimorphisms, or even polymorphisms.

Although the genetic p-distance under CO1 between both populations of I. azteca was 3.5% (Table 1), this mitochondrial marker is susceptible to over- and underestimating diversity in some cases; therefore, the limits interpretation must be considered with caution (Astrin et al., 2006; Ortiz & Francke, 2016). Additional informative mitochondrial (e.g., 16S) or even nuclear (e.g., ITS1, ITS2, or 28S) markers, as well as further morphological evidence, would help to obtain better resolution of species hypotheses (Astrin et al., 2006; Agnarsson, 2010; Navarro-Rodríguez & Valdez-Mondragón, 2020; Nolasco & Valdez-Mondragón, 2022a; Ortiz & Francke, 2016; Planas & Ribera, 2015; Valdez-Mondragón et al., 2019).

For the most part, the natural distribution of the genus Ixchela is in temperate climate zones, principally in pine, oak, or pine-oak forests, between 1,000-2,950 m of elevation (Valdez-Mondragón, 2013). However, species such as Ixchela juarezi Valdez-Mondragón, 2013have been recorded in thorny scrub forest. The population of I. azteca from Cortazar, Guanajuato, was similarly collected in a disturbed thorny scrub forest in the Chihuahuan Desert biogeographic province. Although intensive collecting efforts have targeted low elevations, Ixchela seems to occur only above 1,000 m asl mainly in temperate climate regions (Valdez-Mondragón, 2013, 2020; Valdez-Mondragón & Francke, 2015). Species such as I. juarez and I. azteca that are found in arid and semiarid elevations might represent relict species or populations resulting from vicariant events, and the temperate mountains likely served as a conduit for dispersal. Allopatric populations of I. azteca may be a product of the repeated appearance and disappearance of orographic barriers and habitat fragmentation across a complex landscape formed in response to past orogenic events and climate oscillations, mainly in the Transmexican Volcanic Belt (TVB) biogeographic province (transition zone between the Nearctic and Neotropical regions), as was demonstrated by Valdez-Mondragón and Francke (2015) in the genus Ixchela.

Based on the available evidence, we propose that the allopatric populations of I. azteca assessed herein belong to the same species considering the following: 1) although morphological variation is evident, this variation could be influenced by environmental effects, genetic mutations, developmental perturbations, or even geographic variation as was suggested in other groups by Debat and David (2001); 2) although 2 clades of I. azteca were recovered under ML with a genetic p-distance of 3.5%, the populations from central Mexico present a low bootstrap support value (59%) and 2 subclades were found; 3) incongruence among molecular species delimitation methods was observed with regards to I. azteca, with some methods recovering both clades as a single species; and 4) the population from Guanajuato was collected in a thorny scrub forest (Chihuahuan Desert province-Southern Semiarid Elevations ecoregion), while the rest of the specimens from central Mexico where collected in temperate pine-oak forests (TVB province-Temperate Mountains ecoregion) (Fig. 20). We suggest that the observed morphological and genetic variation might be due to the geographic variation of the species.

In conclusion and considering the suggestions of Carstens et al. (2013), while probing different methods or lines of evidence is necessary for properly implementing species delimitation, it is better to be conservative about species delimitation when the information and results are incongruent to avoid over-splitting species.

Acknowledgments

The first author thanks SEP-Conahcyt for financial support of the project of Basic Science (Ciencia Básica) 2016, No. 282834. We also thank Edmundo González Santillán (curator) and Oscar F. Francke (ex-curator) of the Colección Nacional de Arácnidos (CNAN), Instituto de Biología, UNAM, for providing specimen loans. We are grateful to Laura Márquez Valdelamar for help in the molecular sequencing of the samples, to Brett O. Butler for the English language review of the manuscript, and to the reviewers for their comments and suggestions that improved the manuscript. We thank Carlos Palacios-Cardiel (Technician of the Centro de Investigaciones Biológicas del Noroeste) for his support with storage of the material at the CARCIB. Specimens were collected under the Scientific Collector Permit FAUT-0309 from the Secretaría de Medio Ambiente y Recursos Naturales provided to Alejandro Valdez Mondragón.

References

Agnarsson, I. (2010). The utility of ITS2 in spider phylogenetics: Notes on prior work and an example from Anelosimus. The Journal of Arachnology, 38,377–382. https://doi.org/10.1636/B10-01.1 

Andersson, M. (1994). Sexual selection. Princeton, NJ: Princeton University Press. 

Astrin, J. J., Huber, B. A., Misof, B., & Klütsch, C. F. C. (2006). Molecular taxonomy in pholcid spiders (Pholcidae: Ara-
neae): evaluation of species identification methods using CO1 and 16S and rRNA. Zoologica Scripta, 35, 441–457. https://doi.org/10.1111/j.1463-6409.2006.00239.x

Barraclough, T. G., Harvey, P. H., & Nee, S. (1995). Sexual selection and taxonomic diversity in passerine birds. Proceedings of the Royal Society of London, Series B, 259, 211–215. https://doi.org/10.1098/rspb.1995.0031 

Carstens, B. C., Pelletier, T. A., Reid, N. M., & Satler, J. (2013). How to fail at species delimitation. Molecular Ecology, 22,4369–4383. https://doi.org/10.1111/mec.12413 

Correa-Ramírez, M. M., Jiménez, M. L., & García-De León, F. J. (2010). Testing species boundaries in Pardosa sierra (Araneae: Lycosidae). Journal of Arachnology, 38, 538–554. https://doi.org/10.1636/Sh09-15.1 

DeSalle, R., Egan, M. G., & Siddall, M. (2005). The unholy trinity: taxonomy, species delimitation and DNA barco-ding. Philosophical Transactions of the Royal Society, London, Ser. B, 360, 1905‒1916. https://doi.org/10.1098/rstb.2005.1722 

Debat, V., & David, P. (2001). Mapping phenotypes: canalization, plasticity and developmental stability. Trends in Ecology and Evolution, 16,555‒561. https://doi.org/10.1016/S0169-5347(01)02266-2 

Drummond, A. J., Suchard, M. A., Xie, D., Rambaut, A. (2012). Bayesian phylogenetics with BEAUti and the BEAST 1.7. Molecular Biology and Evolution, 29, 1969–1973. https://doi.org/10.1093/molbev/mss075 

Eberhard, W. G. (1985). Sexual selection and animal genitalia. Cambridge, MA: Harvard University Press. https://doi.org/10.4159/harvard.9780674330702 

Eberhard, W. G., Huber, B. A., Rodríguez, R. L., Briceno, R. D., Salas, I., & Rodríguez, V. (1998). One size fits all? Relationships between the size and degree of variation in genitalia and other body parts in twenty species of insects and spiders. Evolution, 52, 415–431. https://doi.org/10.1111/j.1558-5646.1998.tb01642.x 

Fujisawa, T., & Barraclough, T. G. (2013). Delimiting species using single-locus data and the Generalized Mixed Yule Coalescent approach: a revised method and evaluation on simulated data sets. Systematic Biology, 62, 707–724. https://doi.org/10.1093/sysbio/syt033 

Graham, M. R., Hendrixson, B. E., Hamilton, C. A., & Bond, J. E. (2015). Miocene extensional tectonics explain ancient patterns of diversification among turret-building tarantulas (Aphonopelma mojave group) in the Mojave and Sonoran deserts. Journal of Biogeography, 42, 1052–1065. https://doi.org/10.1111/jbi.12494 

Gross, M. R. (1996). Alternative reproductive strategies and tactics: diversity within sexes. Trends in Ecology &Evolution, 11, 92–98. https://doi.org/10.1016/01695347(96)81050-0 

Hall, T. A. (1999). BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series, 41, 95–98.

Hamilton, C. A., Formanowicz, D. R., & Bond, J. E. (2011). Species delimitation and phylogeography of Aphonopelma hentzi (Araneae, Mygalomorphae, Theraphosidae): Cryptic diversity in North American tarantulas. Plos One, 6, e26207. https://doi.org/10.1371/journal.pone.0026207 

Hamilton, C. A., Hendrixson, B. E., Brewer, M. S., & Bond, J. E. (2014). An evaluation of sampling effects on multiple DNA barcoding methods leads to an integrative approach for delimiting species: A case study of the North American tarantula genus Aphonopelma (Araneae, Mygalomorphae, Theraphosidae). Molecular Phylogenetics and Evolution, 71, 79–93. https://doi.org/10.1016/j.ympev.2013.11.007 

Hamilton, C. A., Hendrixson, B. E., & Bond, J. E. (2016). Taxonomic revision of the tarantula genus Aphonopelma Pocock, 1901 (Araneae, Mygalomorphae, Theraphosidae) within the United States. Zookeys, 560, 1–340. https://doi.org/10.3897/zookeys.560.6264 

Hebert, C. A., Ball, S. L., & Dewaard, J. R. (2003). Biological identifications through DNA barcodes. Proceedings Bio-logical Sciences, 270,313–321. https://doi.org/10.1098/rspb.2002.2218 

Hebert, P. D. N., Ratnasingham, S., DeWaard, J. R. (2003). Barcoding animal life: Cytochrome c oxidase subunit 1 divergences among closely related species. Proceedings of the Royal Society of London, Ser. B, 270, S96–S99. https://doi.org/10.1098/rsbl.2003.0025 

Hebert, P. D. N., Penton, E. H., Burns, J. M., Janzen, D. H., & Hallwachs, W. (2004). Ten species in one: DNA barcoding reveals cryptic species in the Neotropical skipper butterfly Astraptes fulgerator. Proceedings of the National Academy of Sciences of the United States of America, 101, 14812‒14817. https://doi.org/10.1073/pnas.0406166101 

Huber, B. A. (2000). New World Pholcid Spiders (Araneae: Pholcidae): a revision at generic level. Bulletin of the American Museum of Natural History, 254, 1–348. http://dx.doi.org/10.1206/0003-0090(2000)254<0001:NWPSAP>2.0.CO;2 

Huber, B. A. (2003). Rapid evolution and species-specificity of arthropod genitalia: fact or artifact? Organisms Diversity
& Evolution, 3, 63–71. https://doi.org/10.1078/1439-6092-00059 

Huber, B. A. (2011). Phylogeny and classification of Pholcidae (Araneae): an update. Journal of Arachnology, 39, 211–222. https://doi.org/10.1636/CA10-57.1 

Huber, B. A. (2018). Cave-dwelling pholcid spiders (Araneae, Pholcidae): a review. Subterranean Biology, 26, 1–18. http://doi.org/10.3897/subtbiol.26.26430 

Huber, B. A. (2021). Beyond size: sexual dimorphisms in pholcid spiders. Arachnology, 18, 656–677. https://doi.org/10.13156/arac.2020.18.7.656 

Huber, B. A., & Pérez G. A. (2001). A new genus of pholcid spiders (Araneae: Pholcidae) endemic to western Cuba, with a case of female genitalic dimorphism. American Museum Novitates, 3329, 1–23. https://doi.org/10.1206/00030082(2001)329<0001:ANGOPS>2.0.CO;2 

Kapli, P., Lutteropp, S., Zhang, J., Kobert, K., Pavlidis, P., & Stamatakis, A. (2017). Multi-rate Poisson tree processes for single-locus species delimitation under maximum likelihood and Markov chain Monte Carlo. Bioinformatics, 33, 1630–1638. https://doi.org/10.1093/bioinformatics/btx02 

Katoh, K., & Toh, H. (2008). Recent developments in the MAFFT multiple sequence alignment program. MAFFT version 7. Briefings in Bioinformatics, 4, 286–298. https://doi.org/10.1093/bib/bbn013

Kearse, M., Moir, R., Wilson, A., Stones-Havas, S., Cheung, M., & Sturrock, S. (2012). Geneious Basic: an integrated and extendable desktop software platform for the organization and analysis of sequence data. Bioinformatics, 28, 1647–1649. https://doi.org/10.1093/bioinformatics/bts199 

Letunic, I., & Bork, P. (2021). Interactive Tree of Life (iTOL) v5: an online tool for phylogenetic tree display and annotation. Nucleic Acids Research, 49, 293–296. https://doi.org/10.1093/nar/gkab301 

Monaghan, M. T., Wild, R., Elliot, M., Fujisawa, T., Balke, M., & Inward, D. J. (2009). Accelerated species inventory on Madagascar using coalescent-based models of species delineation. Systematic Biology, 58,298–311. https://doi.org/10.1093/sysbio/syp027 

Navarro-Rodríguez, I., & Valdez-Mondragón, A. (2020). Description of a new species of Loxosceles Heineken & Lowe (Araneae, Sicariidae) recluse spiders from Hidalgo, Mexico, under integrative taxonomy: morphological and DNA barcoding data (CO1+ITS2). European Journal of Taxonomy, 704, 1–30. https://doi.org/10.5852/ejt.2020.704 

Navarro-Rodríguez, C. I., & Valdez-Mondragón, A. (2024). Violins we see, species we don’t… Species delimitation of the spider genus Loxosceles Heineken & Lowe (Araneae: Sicariidae) from North America using morphological and molecular evidence. Zootaxa, 5428, 527–548. https://doi.org/10.11646/zootaxa.5428.4.4

Nolasco, S., & Valdez-Mondragón, A. (2022a). To be or not to be… Integrative taxonomy and species delimitation in the daddy long-legs spiders of the genus Physocyclus (Araneae, Pholcidae) using DNA barcoding and morphology.
Zookeys, 1135, 93–118. https://doi.org/10.3897/zookeys.1135.94628

Nolasco, S., & Valdez-Mondragón, A. (2022b). Four new species of the spider genus Physocyclus Simon, 1893 (Araneae: Pholcidae) from Mexico, with updated taxonomic identification keys. European Journal of Taxonomy, 813, 173–206. https://doi.org/10.5852/ejt.2022.813.1739 

Ortiz, D., & Francke, O. F. (2016). Two DNA barcodes and morphology for multi-method species delimitation in Bonnetina tarantulas (Araneae: Theraphosidae). Molecular Phylogenetics and Evolution, 101, 176‒193. https://doi.org/10.1016/j.ympev.2016.05.003 

Planas, E., & Ribera, C. (2015). Description of six new species of Loxosceles (Araneae: Sicariidae) endemic to the Canary Islands and the utility of DNA barcoding for their fast and accurate identification. Zoological Journal of the Linnean Society, 174, 47–73. https://doi.org/10.1111/zoj.12226 

Pons, J., Barraclough, T. G., & Gómez-Zurita, J. (2006). Sequence based species delimitation for the DNA taxonomy of undescribed insects. Systematic Biology, 55, 595–609. https://doi.org/10.1080/10635150600852011 

Posada, D., & Buckley, T. R. (2004). Model selection and model averaging in phylogenetics: advantages of the AIC and Bayesian approaches over likelihood ratio tests. Systematics Biology, 50, 580–601. https://doi.org/10.1080/10635150490522304 

Puillandre, N., Brouillet, S., & Achaz, G. (2021). ASAP.
Assemble species by automatic partitioning. Molecular Ecology Resources, 21, 609–620. https://doi.org/10.1111/1755-0998.13281

Rambaut, A., & Drummond, A. J. (2003). TRACER, MCMC trace analysis tool. Version 1.6. Institute of Evolutionary Biology, University of Edinburgh, Edinburgh, Department of Computer Science, University of Auckland, Auckland.

Ronquist, F., & Huelsenbeck, J. P. (2003). MRBAYES 3: Bayesian phylogenetic inference under mixed models. Bioinformatics, 19, 1572‒1574. https://doi.org/10.1093/bioinformatics/btg180 

Tahami, M. S., Zamani, A., Sadeghil, S., & Ribera, C. (2017). A new species of Loxosceles Heineken & Lowe, 1832 (Araneae: Sicariidae) from Iranian caves. Zootaxa, 4318, 377–387. https://doi.org/10.11646/zootaxa.4318.2.10 

Tamura, K., Dudley, J., Nei, M., & Kumar, S. (2007). MEGA7: Molecular Evolutionary Genetics Analysis (MEGA) software version 4.0. Molecular Biology and Evolution, 24, 1596–1599. https://doi.org/10.1093/molbev/msm092 

Valdez-Mondragón, A. (2010). Revisión taxonómica del género de arañas Physocyclus Simon, 1893 (Araneae: Pholcidae), con la descripción de especies nuevas de México. Revista Ibérica de Aracnología, 18, 3–80.

Valdez-Mondragón, A. (2013). Taxonomic revision of the spider genus Ixchela Huber, 2000 (Araneae: Pholcidae), with description of ten new species from Mexico and Central America. Zootaxa, 3608, 285–327. http://dx.doi.org/10.11646/zootaxa.3608.5.1 

Valdez-Mondragón, A. (2020). COI mtDNA barcoding and morphology for species delimitation in the spider genus Ixchela Huber (Araneae: Pholcidae), with the description of two new species from Mexico. Zootaxa, 4747, 54–76. https://doi.org/10.11646/zootaxa.4747.1.2 

Valdez-Mondragón, A., & Francke, O. F. (2015). Phylogeny of the spider genus Ixchela Huber, 2000 (Araneae: Pholcidae) based on morphological and molecular evidence (CO1 and 16S), with a hypothesized diversification in the Pleis-
tocene. Zoological Journal of the Linnean Society, 175, 20‒58. https://doi.org/10.1111/zoj.12265 

Valdez-Mondragón, A., Navarro-Rodríguez, C. I., Solís-Catalán K. P., Cortez-Roldán, M. R., & Juárez-Sánchez, A. R. (2019). Under an integrative taxonomic approach: the description of a new species of the genus Loxosceles (Araneae, Sicariidae) from Mexico City. Zookeys, 892, 93–133. https://doi.org/10.3897/zookeys.892.39558 

World Spider Catalog (WSC). (2025). World Spider Catalog. Version 25.5. Natural History Museum Bern. http://wsc.nmbe.ch

Zhang, J., Kapli, P., Pavlidis, P., & Stamatakis, A. (2013). A general species delimitation method with applications to phylogenetic placements. Bioinformatics, 29, 2869–2876. https://doi.org/10.1093/bioinformatics/btt499

Maria Alejandra Villegas-Ojeda ^{a, b}, Guido Fernández-Marinone ^b, Mariana Jofré ^b, Cynthya González ^c, *

^a Universidad Nacional de San Luis – Consejo Nacional de Investigaciones Científicas y Técnicas, Almirante Brown 907, 5700 San Luis, Argentina

^b Universidad Nacional de San Luis, Facultad de Química, Bioquímica y Farmacia, Área de Biología, Departamento de Biología, Ejército de los Andes 950, segundo piso, 5700 San Luis, Argentina 

^c Universidad Nacional del Nordeste – Consejo Nacional de Investigaciones Científicas y Técnicas, Centro de Ecología Aplicada del Litoral, Ruta Provincial Nº 5 Km 2,5, 3400 Corrientes, Argentina 

*Corresponding author: cynthyaelizabethgonzalez@gmail.com (C. González)

Received: 16 October 2023; accepted: 2 October 2024

Abstract

The studies of helminth parasites in Argentine amphibians have focused on the Dry and Humid Chaco ecoregions; in the Dry Chaco the reports are from the Chaco and Formosa provinces. The aim of this work was to report the first record of Aplectana hylambatis (Baylis, 1927) Travassos, 1931 (Nematoda: Cosmocercidae) from San Luis province and the southernmost record for the Dry Chaco ecoregion. Specimens of Rhinella arenarum (Hensel, 1867) (n = 6), Leptodactylus mystacinus (Burmeister, 1861) (n = 13), and Odontophrynus cf. asper (n = 6) were analyzed. Specimens of A. hylambatis were collected from the large intestine of these hosts and were studied using light and scanning electron microscopy (SEM). In addition, relevant features are described, including the number and arrangement of caudal papillae, and of mamelon-like cuticular protuberances associated to the vulva. The morphological characteristics are compared and discussed in relation to those of specimens collected from other hosts in other localities of the country. The presence of A. hylambatis in localities of the Dry Chaco represents an expansion of the geographic range of this cosmocercid and the first record in L. mystacinus and O. cf. asper from Argentina.

Keywords: Aplectana hylambatis; Bufonidae; Cosmocercidae;Dry Chaco; Leptodactylidae; Odontophrynidae; Parasitic helminths

© 2025 Universidad Nacional Autónoma de México, Instituto de Biología. Este es un artículo Open Access bajo la licencia CC BY-NC-ND

(http://creativecommons.org/licenses/by-nc-nd/4.0/).

Primer reporte de Aplectana hylambatis (Nematoda: Cosmocercidae) en anfibios
de la provincia de San Luis, Argentina 

Resumen

Los estudios de helmintos parásitos de anfibios argentinos se han centrado en las ecorregiones del Chaco Seco y Húmedo; en el Chaco Seco los reportes se realizaron en las provincias de Chaco y Formosa. El objetivo de este trabajo fue reportar el primer registro de Aplectana hylambatis (Baylis, 1927) Travassos, 1931 (Nematoda: Cosmocercidae) para la provincia de San Luis y el registro más austral para la ecorregión del Chaco Seco. Se analizaron ejemplares de Rhinella arenarum (Hensel, 1867) (n = 6), Leptodactylus mystacinus (Burmeister, 1861) (n = 13) y Odontophrynus cf. asper (n = 6). Los ejemplares de A. hylambatis fueron colectados del intestino grueso de estos hospedadores y se estudiaron mediante microscopía óptica y electrónica de barrido (MEB). Se describen características relevantes, como el número y disposición de las papilas caudales, y el número y disposición de las protuberancias cuticulares tipo mamelón asociadas a la vulva. Los caracteres morfológicos encontrados se comparan y discuten con los especímenes colectados en otros hospedadores de diferentes localidades del país. La presencia de A. hylambatis en localidades del Chaco Seco representa una ampliación del rango geográfico de este cosmocércido y el primer registro para L. mystacinus y O. cf. asper de Argentina.

Palabras clave: Aplectana hylambatis; Bufonidae; Cosmocercidae; Chaco Seco; Leptodactylidae; Odontophrynidae; Helmintos parásitos

Introduction 

In 1927, Baylis described Oxysomatium hylambatis parasitizing the large intestine of Leptopelis aubryi (Duméril, 1856) from Macenta, Guinea, Africa (Baylis, 1927). In that work, the author established a synonymy between the genera Oxysomatium and Aplectana; however, he did not question the validity of the genus Aplectana, stating that it included species without a gubernaculum (called at that time “accessory piece”).

Later, Travassos (1931) published a review of the family Cosmocercidae (Railliet, 1916) Travassos, 1925, where he established that the division between the 2 genera occurs mainly by the female reproductive system, which is prodelphic in the genus Aplectana and amphidelphic in Oxysomatium. In this way, it reproduces the description of the specimens made by Baylis (1927), but places them under the genus Aplectana due to the anterior location of the reproductive system of females.

Over the years, both genera have been studied in numerous works around the world. In some of them, these taxa were considered synonymous (Fotedar, 1960; Gutiérrez, 1945; Harwood, 1930; Hsü & Hoeppli, 1933; Skrjabin, 1910, 1951; Walton, 1940, 1941), whereas in others, they were separated on the basis of the size and shape of gubernaculum and spicules, the presence or absence of lateral alae, the number of cephalic papillae, the position of the vulva, and the stage of the eggs in utero (Bravo-Hollis, 1943), or on the basis of the presence or absence of gubernaculum in males, and prodelphic or amphidelphic condition in females (Ballesteros-Márquez, 1945).

Baker (1980) redescribes A. hylambatis (Baylis, 1927) Travassos, 1931 based on specimens collected from Sclerophrys mauritanica (Schlegel, 1841) (= Bufo mauritanicus), Leptopelis aubryi (Duméril, 1856), and Rhinella achalensis (Cei, 1972) (= Bufo achalensis). Subsequently, Baker and Vaucher (1986) synonymized Oxysomatium bonariensis (Gutiérrez, 1945) collected from Rhinella arenarum (Hensel, 1867) (= Bufo arenarum) in Argentina, and A. pudenda (Masi Pallarés and Maciel, 1974) collected from Rhinella diptycha (Cope, 1862) (= Bufo paracnemis), Rana pacybrachion (Steffen, 1815) (= Leptodactylus latrans), Boana raniceps (Cope, 1862), and Boana punctata (Schneider, 1799) (= Hyla punctata) in Paraguay, with A. hylambatis.

In South America, this cosmocercid has a wide geographic and host range; it was found in Argentina, Ecuador, Guyana, Paraguay, Peru, Brazil, and Uruguay (Campião et al., 2014; González, Hamann et al., 2021). In Argentina it is the nematode species of amphibians with the widest geographic distribution. There are records in 7 provinces, namely Buenos Aires, Córdoba, Corrientes, Chaco, Formosa, Salta, and San Juan, where it has been found parasitizing anurans of the families Bufonidae, Ceratophryidae, Hylidae, Leptodactylidae, Microhylidae, and Odontophrynidae (Draghi et al., 2015; González et al., 2019; González, Hamann et al., 2021).

Rhinella arenarum (Anura: Bufonidae) is distributed in Argentina, Bolivia, Brazil, and Uruguay (Frost, 2023). It is characterized by having terrestrial habits, reproducing in temporary or semi-permanent lentic and lotic environments where it deposits strings of pigmented eggs, and having a generalist diet (Babini et al., 2015; Bionda et al., 2011; de Oliveira et al., 2017; Quiroga et al., 2009). Leptodactylus mystacinus (Burmeister, 1861) (Anura: Leptodactylidae) is distributed in Argentina, Bolivia, Brazil, Paraguay, and Uruguay (Frost, 2023). It is a terrestrial anuran, with fossorial characteristics; it breeds on foam nests built in temporary or semi-permanent lentic and lotic environments, and has a generalist diet (Babini et al., 2015; De-Carvalho et al., 2008; Heyer et al., 2010; López et al., 2005). These amphibian species are categorized as “Not Threatened” according to Vaira et al. (2012) and as “Least Concern” according to IUCN (2023). Finally, the genus Odontophrynus Reinhardt and Lütken, 1862 (Anura: Odontophrynidae), which includes 11 species, is distributed in southern and eastern South America (Frost, 2023). Recently, Argentinean species of this genus were revised by Rosset et al. (2022). These authors synonymized Odontophrynus americanus (Duméril and Bibron, 1841) with O. occidentalis, and established its distribution area, restricted to the center, west and south of the country, in the ecoregions of Monte, Dry Chaco, Patagonian steppe, and in the coastal dunes of the Humid Pampa ecoregion. On the other hand, these authors also proposed to name the cryptic diploid and tetraploid species currently grouped as the Odontophrynus americanus species group, as Odontophrynus asper (Philippi, 1902), with distribution in extensive areas of central and northern Argentina, Uruguay, southern Paraguay, southeastern Brazil, and probably also Bolivia (Rosset et al., 2022).

Studies on parasitic helminths in these 3 hosts, both in the larval and adult stages, refer to descriptions of new species, intraspecific variations, or extensions of host or geographic range (Campião et al., 2014; da Graça, et al., 2017; da Silva et al., 2018; González et al., 2019; González, Duré et al., 2021; González, Hamann et al., 2021; Queiroz et al., 2020; Ramallo et al., 2020). Throughout its geographic range, the helminth fauna of R. arenarum has been studied in Argentina, Brazil, and Uruguay (Campião et al., 2014). Specifically, in Argentina, A. hylambatis has been reported from Buenos Aires and San Juan provinces; Rhabdias elegans Gutiérrez, 1945 from Buenos Aires and Tucumán provinces; Borrellostrongylus platensis Gutiérrez, 1945 (= Parabatrachostrongylus platensis Tantalean and Naupay, 1974) from Buenos Aires province; Oswaldocruzia proencai Ben Slimane and Durette-Desset, 1995, Ophidascaris sp., Aplectana tarija Ramallo, Bursey and Goldberg, 2007, and Cosmocercoides lilloi Ramallo, Bursey and Goldberg, 2007 from Salta province; and Bufonerakis andersoni Baker, 1980 has been found but its locality was not reported (González and Hamann, 2015; González et al., 2019; Ramallo et al., 2020). For L. mystacinus, previous reports about its helminths have been made in Brazil and Paraguay (Campião et al., 2014); so far, no helminths have been found in this amphibian in Argentina. The previous records of helminths that have been made in the country for amphibians of the genus Odontophrynus include the nematodes R. elegans, Cosmocerca podicipinus Baker and Vaucher, 1984, and C. parva Travassos, 1925, and the digeneans Echinostomatidae gen. sp., and Travtrema aff. stenocotyle for O. asper (= O. americanus) from Corrientes province; A. hylambatis and Falcaustra sanjuanensis González, Sanabria and Quiroga, 2013 for O. americanus (= O. occidentalis) from San Juan province, and A. hylambatis for O. lavillai Cei, 1985 from Formosa province (Campião et al., 2014; González, Hamann et al., 2021; Piñeiro-Gómez et al., 2023).

The aim of this work was to report the first record of the cosmocercid A. hylambatis from the province of San Luis, Argentina, in the anurans R. arenarum, L. mystacinus, and O. cf. asper.

Materials and methods

San Luis, Argentina is part of the Dry Chaco ecoregion, and has xerophytic deciduous shrub vegetation with patches of Larrea (Morello et al., 2012). The sampled sites were located on the Central Sierras of San Luis. In the extreme west, these mountain ranges constitute an orographic barrier to humid winds from the east, which causes greater rainfall on the eastern slopes and drier climates to the west (Torrella & Adámoli, 2006). From November 2019 to February 2023, through active sampling techniques of direct visual recording and auditory detection (Vaira et al., 2021), 25 anuran amphibians of 3 species, belonging to 3 families, were captured: R. arenarum (Bufonidae) (n = 6), L. mystacinus (Leptodactylidae) (n = 13), and O. cf. asper (Odontophrynidae) (n = 6) (Table 1). Secretaría de Medio Ambiente, San Luis Province, authorized the capture of the specimens for this study (Resolution N°295-PRN-2022). 

All organ systems were examined in the captured specimens, using helminthological techniques according to the criteria of Goater and Goater (2001) and González and Draghi (2021). Nematodes found in the large intestine, were counted, fixed with a hot 10% formalin solution, preserved in 70% alcohol, cleared with Amann’s lactophenol and then, were morphometrically analyzed. Illustrations were made using a Leica DM2500 optical microscope with the aid of a drawing tube. For examination with scanning electron microscope (SEM), nematodes were dehydrated through a series of ethanol and acetone, and critical point-dried using CO₂. The specimens were coated with gold-palladium and examined with a JEOL JSM-5800 scanning electron microscope. Measurements along the text are given in micrometers unless otherwise indicated and are presented as the mean ± 1 standard deviation, with the minimum and maximum value in parentheses. Nematodes were deposited in the Helminthological Collection of Museo de la Plata, La Plata, Argentina (MLP-He). The anuran specimens are stored in the Universidad Nacional de San Luis Herpetology Collection (CH-UNSL), San Luis, Argentina.

Results

Of the total amphibians analyzed, 11 were parasitized (total prevalence: 44%). In these, 338 specimens belonging to the species Aplectana hylambatis (Baylis, 1927) Travassos, 1931 (Nematoda, Cosmocercidae) were found (accession numbers: MLP-He 8044, 5 males and 5 females from O. cf. asper; MLP-He 8045, 5 males and 5 females from L. mystacinus; MLP-He 8046, 5 males and 5 females from R. arenarum). 

Description

Aplectana hylambatis (Baylis, 1927) Travassos, 1931 [= Oxysomatium bonariensis

Gutiérrez, 1945; = Aplectana pudenda Masi Pallarés and Maciel, 1974] 

(Figs. 1-3)

Table 1

Sample size (n), sex (F: females; M: males), collection date and accession numbers for 3 amphibian species from San Luis Province, Argentina.

General morphology (based on 82 adult specimens). Small nematodes, cuticle with transverse striations evenly distributed along the body. Lateral alae present in both sexes beginning at the level of the nerve ring and ending at the level of the anus in females and at the level of the first pairs of precloacal papillae in males. Triangular mouth surrounded by 3 lips, 1 dorsal with 2 cephalic papillae, and 2 ventral lips each with a papilla and an amphid (Fig. 1A). Well-differentiated esophageal bulb with chitinous valves. Very visible excretory pore, pre-bulbar and fringed (Fig. 2A). Prodelphic females with a highly developed ovijector (Fig. 2D, E); thin-membrane, morulated eggs and larvated eggs at the level of the vulva. Males with gubernaculum and equal spicules with characteristic articulation. The morphometric characters of males and females collected are provided in Table 2. 

Male (based on 40 adult specimens). Specimens with 3 typical groups of caudal papillae present in this species, precloacal, adcloacal, and postcloacal, with a large medial unpaired papilla anterior to the cloaca. In the analyzed specimens, thirteen pairs of caudal papillae were counted with the following arrangement: 4 pairs of ventral precloacal papillae; 5 pairs adcloacal (2 pairs ventrolateral, 3 pairs in anterior lip of the cloaca) and 1 unpaired papilla on anterior lip of the cloaca; 4 pairs of postcloacal papillae (2 pairs ventrolateral at the middle tail; 2 pairs close to its posterior end) (Figs. 1B-D; 2B, C). 

Table 2

Morphometric characters (mean ± SD and Min.-Max.) of males and females of Aplectana hylambatis collected in the 3 anuran species from San Luis Province, Argentina.

Figure 1. Aplectana hylambatis parasite of anurans from San Luis province, Argentina. A, Anterior end, apical view; B, male, posterior end, ventral view; C, detail of cloacal region, latero-ventral view; D, detail of paired papillae, latero-ventral view; E-F, female, mamelon-like cuticular, latero-ventral view. Abbreviations: a: amphid; ad: adcloacal papillae; fc: cuticular fringe; up: unpaired papilla; po: postcloacal papillae; pr: precloacal papillae; * cephalic papillae; closed arrows indicate paired papillae. A-D from O. cf. asper; E, F from L. mystacinus.

Figure 2. Aplectana hylambatis parasite of anurans from San Luis province, Argentina. A, Male, anterior end, ventral view; B, male, posterior end, ventral view; C, male, posterior end, lateral view; D, ovijector and 2 mamelon-like cuticular protuberances anterior to the vulva, ventral view; E, ovijector and 2 mamelon-like cuticular protuberances, 1 anterior and 1 posterior to the vulva, ventral view; F, female, posterior end, lateral view; G, gubernaculum, ventral view; H, spicules and gubernaculum, ventral view. Scale: A-D: 200 μm; E: 100 μm; F: 200 μm; G: 50 μm; H: 100 μm). A-C from R. arenarum; D-F from L. mystacinus; G, H from O. cf. asper.

Spicules and gubernaculum did not show morphological variation between specimens collected in different amphibian hosts and localities. Spicules showed the characteristic structure described for the species, long and chitinised, with a capitulum in the proximal end, and a pointed distal end that, when the spicules are outside the body, have a sickle or hockey stick appearance (Fig. 2H). Gubernaculum has an oval shape, chinitised, concave ventrally, with thickened edges, tapering towards the distal end (Fig. 2G).

Female (based on 42 adult specimens). Regarding the mamelon-like cuticular protuberances, most of the specimens analyzed showed between 1 large mamelon or 2 smaller mamelon-like protuberances located on the anterior lip of the vulva. However, in an analyzed specimen collected in L. mystacinus, 2 mamelon-like cuticular protuberances were observed, but with a different disposition than the one previously detailed; in this case, one protuberance was located anterior to the vulva and the other, posterior to it (Figs. 1E, F; 2D-F). 

Some specimens collected from R. arenarum (2 couples) and from L. mystacinus (2 couples) were found in copulation (Fig. 3A). In these cases, a detailed examination of these specimens showed that only the right spicule enters the female reproductive system. In it, the distal end presented the typical articulation described in previous works. This articulation is in the elbow of the ovijector. The spicule that remains inside the male reproductive system was fully stretched. Although the specimens were clarified, it was not possible to identify the gubernaculum in the specimens in copulation (Fig. 3B).

Figure 3. Aplectana hylambatis parasite of anurans from San Luis province, Argentina. A, Couples in copulation; B, detail of the arrangement of the spicules at the copulation time. Scale: A: 400 μm; B: 100 μm. A, B from L. mystacinus.

Discussion

The genus Aplectana is characterized by having lateral alae and somatic papillae in both sexes. Males lack plectanes or rosettes at the posterior end. Females have numerous thin-shelled eggs in the uterus and have prodelphic reproductive system (Gibbons, 2010). These characters were easily observable in the specimens studied here. The species of the genus Aplectana are grouped mainly according to the presence or absence of gubernaculum. Other features that distinguish these helminths are the number and arrangement of caudal papillae, and the size and morphology of the gubernaculum and spicules (Baker, 1980; Piñeiro-Gómez et al., 2017).

Morphological characters such as number and arrangement of caudal papillae in males, and mamelon-like cuticular protuberances on the anterior lip of the vulva in females, are presented in Table 3, and compared to those previously found in other hosts and other localities in Argentina. Regarding the number and arrangement of the caudal papillae, the males collected from the 3 host species presented a total of 13 pairs with the formula: 4:5:4+1. Compared with previous studies, only the specimens found in T. typhonius had a lower number of pairs of caudal papillae (12) (Draghi et al., 2015). Thirteen pairs of caudal papillae were also found in P. santafecinus and R. major from Taco Pozo (Table 3); however, their arrangement presented differences (P. santafecinus: 3:5:5; R. major: 4:4:5 vs. 4:5:4 in the present study). Finally, in males collected from other amphibians, a greater number of pairs of cloacal papillae were observed (up to 16 pairs in D. muelleri) (González et al., 2019) (Table 3).

Table 3

Some morphological characters of males and females of Aplectana hylambatis of the present study compared with specimens collected in different hosts in Argentina. n.e. = not specified. 

The mamelon-like cuticular protuberances are structures that were not included in the original description of the species by Baylis (1927); however, subsequent studies have detailed their number and arrangement in specimens collected from different hosts and locations (Baker, 1980; González et al., 2019; Gutiérrez, 1945). In this study, these structures of the females varied between 1 and 2, and were located on the anterior lip of the vulva. This character of the females was like that found in the specimens studied in R. arenarum from La Plata, R. major from Concepción del Bermejo and Las Lomitas, and L. bufonius from Las Lomitas (Table 3) (González et al., 2019). However, one female presented a protuberance posterior to the vulva; and to our knowledge this had not been reported until now. Finally, of all the morphological characters studied, mamelon-like cuticular protuberances varied the most among hosts and localities (González et al., 2019; Gutiérrez, 1945) (Table 3). 

In reference to the specimens in copulation, they were found with the typical arrangement of the male coiled by its posterior end to the female at the level of the vulva and forming an angle of approximately 90 degrees between them. In nematode mating, the spicules play an active role in copulation, while the gubernaculum, which is typically not everted during this process, acts as a guide for the spicules and prevents them from piercing the wall of the spicular pouch and cloaca when pressure is exerted on them (Chitwood & Chitwood, 1974).

In the 4 couples studied, it was observed that the right spicule is inserted into the vagina of the female and the articulation of the distal part is curved at the level of the ovijector’s elbow, while the left spicule remains fully elongated in the male without showing the articulation. Regarding this, the spicules serve to keep the vulva and vagina open. In trichostrongylids, the branching of the spicular lamina and the formation of twisted spicules seem to be adapted to this activity; in taxa with unequal spicules, the short spicule opens the vulva and the proximal region of the vagina, while the long one ensures sperm progression (Chitwood & Chitwood, 1974). In reference to the gubernaculum, unfortunately, we could not observe its location in the specimens studied. Thus, it is necessary to find new mating specimens to elucidate the position it occupies during copulation.

Intraspecific variations in parasitic nematodes of amphibians have been recorded for different families such as Molineidae (Skrjabin & Schulz, 1937) Durette-Desset and Chabaud, 1977, Onchocercidae (Leiper, 1911), Pharyngodonidae (Travassos, 1919), and Cosmocercidae (González et al., 2019; Kirillova et al., 2021; Kuzmin et al., 2023; Rhoden & Bolek, 2011; Vhora & Bolek, 2013; Willkiens et al., 2023). Specifically, in A. hylambatis, previous studies carried out in different hosts from different locations of the Neotropical realm showed variations in 2 of the characters studied here, namely number and arrangement of mamelon-like cuticular protuberances and number and arrangement of cloacal papillae, in addition to variations in the total length of individuals of both sexes (Baker, 1980; González et al., 2019; Gutiérrez, 1945; Masi-Pallarés & Maciel, 1974). In this regard, intraspecific variation may be attributed to a number of factors related to the host, such as size, age, physical condition, metabolic rate, and food supply, and factors related to the parasite, such as the time of penetration (first, second or subsequent infection) and the crowding of the specimens, or related to different study techniques, in addition to ecological and geographic factors (Baker, 1980; Chitwood, 1957; Perkins et al., 2011; Vhora & Bolek, 2013). 

The studies of helminth parasites of Argentine amphibians have focused on the Dry and Humid Chaco ecoregions; specifically, in the Dry Chaco the reports have been made in the northern provinces of the country, Chaco and Formosa. Until now, the southernmost records in this ecoregion correspond to Concepción del Bermejo in Chaco province (González, Duré et al., 2021; González, Hamann et al., 2021). In this way, the present study expands the knowledge on parasitic helminths from the South American Chaco by providing the southernmost record for the Dry Chaco ecoregion. Furthermore, it represents the first record of A. hylambatis in L. mystacinus and O. cf. asper from Argentina.

Acknowledgments

Financial support was received from Secretaría de Ciencia y Tecnología, Universidad Nacional de San Luis-UNSL (grant number PROICO 02-1520 to M.B.J.), from Universidad Nacional del Nordeste-UNNE (grant number PI21Q001 to C.E.G.), and from CONICET (PIP 11220200101582CO to C.E.G.). M.A. Villegas Ojeda was supported by Doctoral scholarship of CONICET. 

References

Babini, M. S., Salas, N. E., de Lourdes-Bionda, C., & Martino, A. L. (2015). Implicaciones de la urbanización en la presencia, distribución y ecología reproductiva de la fauna de anuros de una ciudad del área central de Argentina. Revista Mexicana de Biodiversidad, 86, 188–195. https://doi.org/10.7550/rmb.43684