NEW RECORDS OF HELMINTHS OF THE JAGUAR IN MEXICO, WITH AN UPDATED LIST OF SPECIES IN THE AMERICAS.

An inventory of parasites infecting the jaguar (Panthera onca) across its distribution range is relevant for the conservation of this threatened big cat. In this study, we report the occurrence of helminths in a jaguar from Mexico using morphological techniques (cleared and stained mounts and scanning electron microscopy) and partial sequences of the 28S ribosomal RNA (28S rRNA) gene and the cytochrome c oxidase 1 mitochondrial (COI) gene. We also provide an updated list of helminth species reported in jaguars in the Americas. Three helminth taxa are identified in the jaguar examined from Mexico: Toxocara cati, Physaloptera sp., and Taenia sp. The new 28S rRNA sequences of To. cati, Physaloptera sp., and Taenia sp. and the COI sequence of Taenia sp. corroborate the identity of the helminths isolated from this host. One hundred and twenty-nine records of helminths parasitizing jaguars from 49 studies up to May 2023 were identified in the Americas. In most of these studies (73.6%), helminths were identified using coproparasitological techniques. Sixteen helminths (7 nematodes, 5 cestodes, 3 acanthocephalans, and 1 trematode) were identified at the species level in free-ranging and captive jaguars. The study demonstrates the value of an integrative taxonomy approach to increase the accuracy of parasite identification in wildlife, especially when helminth specimens are scarce or poorly fixed.

The complete mitochondrial genome of 3 species of allocreadiids (Digenea, Allocreadiidae): characterization and phylogenetic position within the order Plagiorchiida.

Trematodes of the family Allocreadiidae are primarily found in the intestines of freshwater fishes around the world. The family includes 15 genera and c. 130 species. The last 2 decades have witnessed an increase in the genetic library of its species. Molecular data have been crucial for species delimitation and species description within Allocreadiidae and for understanding their evolutionary and biogeographical history and classification. Here, the mitogenomes of 3 species of allocreadiids were obtained using high throughput sequencing methods. Mitogenomes were compared with other members of the order Plagiorchiida to determine their molecular composition, gene rearrangement and phylogenetic interrelationships. The complete circular mitogenomes of Allocreadium lobatum, Creptotrematina aguirrepequenoi and Wallinia mexicana were 14 424, 13 769 and 13 924 bp long respectively, comprising 12 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes and 2 non-coding regions. Gene arrangements were identical to other Xiphidiatan trematodes. Phylogenetic analyses using the mitogenomes revealed Allocreadiidae as a monophyletic group closely related to other members of the suborder Xiphidiata; A. lobatum was yielded as the sister taxon of C. aguirrepequenoi + W. mexicana. Our study increases the complete mitochondrial genome library of trematodes and strengthens our understanding of the phylogenetic relationships and classification of this parasite group.

Dataset of assembly and annotation of the mitogenomes of Triatoma dimidiata and Triatoma huehuetenanguensis captured from Yucatán, México.

Triatoma dimidiata is a species complex, and its members are responsible for the transmission of Trypanosoma cruzi, the causative agent of Chagas disease. We present the assembly and annotation of the mitogenome of the Triatoma dimidiata (Latreille, 1811) and Triatoma huehuetenanguensis Lima-Cordón & Justi, 2019. The mitochondrial genomes were successfully sequenced using the Illumina Nextseq 500 platform, 2×75 cycles, and 5 million reads per sample. Contigs were assembled and annotated using the reference genomes of T. dimidiata and T. huehuetenanguensis available in Genbank (NC_002609 and NC_050325.1, respectively). The mitogenomes of T. dimidiata have lengths of 17,008 bp, while those of T. huehuetenanguensis are 15,910 bp and 15,909 bp. The genome comprises 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes, and a control region. The mitogenomes will be valuable to scholars and students focused on integrative taxonomy, phylogeography, and evolutionary studies of the Triatoma dimidiata complex and the transmission of Chagas diseases.

Description of new species of Trematoda from bats of Southeastern Mexico and a new classification for Brachylecithum rileyi n. comb. (Dicrocoeliidae).

To date, 23 species of trematodes have been reported in bats from Mexico. However, in some regions of Mexico, such as the Yucatan Peninsula, many species of bats do not have helminthological records. Here, we sampled bats in four localities in Southeastern Mexico from April 2017 to February 2022. Parasites were collected from the intestine of four species of bats: Pteronotus fulvus, Eumops nanus, Noctilio leporinus and Nyctinomops laticaudatus. Conventional morphological techniques and molecular tools with the 28S ribosomal gene were used to describe the helminths. We discovered that our parasites represent three new species and a new combination of trematodes. We found that the new species have morphological differences with their congeneric species, and we complement this information whit molecular data. Furthermore, we found morphological and molecular evidence that places Dicrocoelium rileyi within the genus Brachylecithum. This study points out the importance of comparing morphological and phylogenetic information.

Host-parasite interactions in perpetual darkness: Macroparasite diversity in the cavefish Astyanax mexicanus.

Astyanax mexicanus has repeatedly colonized cave environments, displaying evolutionary parallelisms in many troglobitic traits. Despite being a model system for the study of adaptation to life in perpetual darkness, the parasites that infect cavefish are practically unknown. In this study, we investigated the macroparasite communities in 18 cavefish populations from independent lineages and compared them with the parasite diversity found in their sister surface fish populations, with the aim of better understanding the role that parasites play in the colonization of new environments. Within the cavefish populations, we identified 13 parasite taxa, including a subset of 10 of the 27 parasite taxa known for the surface populations. Parasites infecting the cavefish belong to five taxonomic groups, including trematodes, monogeneans, nematodes, copepods, and acari. Monogeneans are the most dominant group, found in 14 caves. The macroparasites include species with direct life cycles and trophic transmission, including invasive species. Surprisingly, paired comparisons indicate higher parasite richness in the cavefish than in the surface fish. Spatial variation in parasite composition across the caves suggests historical and geographical contingencies in the host-parasite colonization process and potential evolution of local adaptations. This base-line data on parasite diversity in cavefish populations of A. mexicanus provides a foundation to explore the role of divergent parasite infections under contrasting ecological pressures (cave vs. surface environments) in the evolution of cave adaptive traits.

Cathetocephalus badilloi n. sp. (Cestoda: Cathetocephalidea) from the spinner shark Carcharhinus brevipinna off the coast of Yucatan, Mexico.

This study describes a new species belonging to the genus Cathetocephalus from the spinner shark Carcharhinus brevipinna off the coast of Yucatan, Mexico. Cathetocephalus badilloi n. sp. has a soft scolex transversely positioned to the long axis of the strobila. The scolex is composed of an apical pad, papillate band and rugose base. The papillate band has two papillary segments; the upper segment has numerous papillae, positioned one next to the other in an irregular and heterogeneous way with a spongy texture that remains along the band. The lower papillary segment has numerous papillae positioned one next to the other, without space between them. This papillary band presents flattened rectangular papillae with a division located at the top of each the papilla, giving it the appearance of a molar tooth. Using the 28S rDNA gene and the method of Maximum Likelihood (ML) for phylogenetic analysis, we identified the present material as a new species. We did not collect specimens with mature or gravid proglottids, but since the specific identification of the members of this genus is based on the characteristics of the scolex, our proposal of the new species is determined by the morphology of this organ as well as the molecular results obtained.

Exploring Evolutionary Relationships within Neodermata Using Putative Orthologous Groups of Proteins, with Emphasis on Peptidases.

The phylogenetic relationships within Neodermata were examined based on putative orthologous groups of proteins (OGPs) from 11 species of Monogenea, Trematoda, and Cestoda. The dataset included OGPs from BUSCO and OMA. Additionally, peptidases were identified and evaluated as phylogenetic markers. Phylogenies were inferred using the maximum likelihood method. A network analysis and a hierarchical grouping analysis of the principal components (HCPC) of orthologous groups of peptidases were performed. The phylogenetic analyses showed the monopisthocotylean monogeneans as the sister-group of cestodes, and the polyopisthocotylean monogeneans as the sister-group of trematodes. However, the sister-group relationship between Monopisthocotylea and Cestoda was not statistically well supported. The network analysis and HCPC also showed a cluster formed by polyopisthocotyleans and trematodes. The present study supports the non-monophyly of Monogenea. An analysis of mutation rates indicated that secreted peptidases and inhibitors, and those with multiple copies, are under positive selection pressure, which could explain the expansion of some families such as C01, C19, I02, and S01. Whilst not definitive, our study presents another point of view in the discussion of the evolution of Neodermata, and we hope that our data drive further discussion and debate on this intriguing topic.

Survey on helminths of bats in the Yucatan Peninsula: infection levels, molecular information and host-parasite networks.

Helminth species of Neotropical bats are poorly known. In Mexico, few studies have been conducted on helminths of bats, especially in regions such as the Yucatan Peninsula where Chiroptera is the mammalian order with the greatest number of species. In this study, we characterized morphologically and molecularly the helminth species of bats and explored their infection levels and parasite­host interactions in the Yucatan Peninsula, Mexico. One hundred and sixty-three bats (representing 21 species) were captured between 2017 and 2022 in 15 sites throughout the Yucatan Peninsula. Conventional morphological techniques and molecular tools were used with the 28S gene to identify the collected helminths. Host­parasite network analyses were carried out to explore interactions by focusing on the level of host species. Helminths were found in 44 (26.9%) bats of 12 species. Twenty helminth taxa were recorded (7 trematodes, 3 cestodes and 10 nematodes), including 4 new host records for the Americas. Prevalence and mean intensity of infection values ranged from 7.1 to 100% and from 1 to 56, respectively. Molecular analyses confirmed the identity of some helminths at species and genus levels; however, some sequences did not correspond to any of the species available on GenBank. The parasite­host network suggests that most of the helminths recorded in bats were host-specific. The highest helminth richness was found in insectivorous bats. This study increases our knowledge of helminths parasitizing Neotropical bats, adding new records and nucleotide sequences.

Linking phenotypic to genotypic metacestodes from Octopus maya of the Yucatan Peninsula.

Octopus maya is an endemic species of the Yucatan Peninsula. This species sustains the octopus' fishery in the region and is the only cephalopod cultured in Mexico. It is known that O. maya harbor a large richness and abundance of metacestodes that have been tentatively identified by light microscopy alone. Since the larval stages of some orders of marine cestodes lack the taxonomic characteristics shown by the adult stages and on which cestode taxonomy is based, identification down to the species level is often unattainable. Hence, the goal of this study was to characterize the parasites, for the first time, at morphological and molecular levels. A total of 60 octopuses were collected from September to December 2017 from four fishery landing ports in Yucatán: Sisal, Progreso, Dzilam de Bravo, and Rio Lagartos (15 hosts per locality). Morphology of metacestodes was characterized by light and Scanning Electron Microscopy (SEM), while the genes 18S and 28S rDNA were sequenced for molecular characterization. Based on phenotypic characters and molecular data, seven taxa of metacestodes were identified, four of them belonging of order Trypanorhyncha: Eutetrarhynchus sp., Kotorella pronosoma, Nybelinia sp., Prochristianella sp. 1; and the three remaining taxa belonging to the order Onchoproteocephalidea: Acanthobothrium sp., Phoreiobothrium sp., and Prosobothrium sp. This work provides, for the first time, molecular support to the morphological characterization of metacestodes recorded in Octopus maya.

Parasites with zoonotic potential found in commercially important fish in Tamaulipas, Northeastern Mexico.

Human population is exposed to numerous parasitic ichthyozoonoses. Although Tamaulipas state (northeastern Mexico) is well known for its fishing and aquaculture industry, there are few reports of this type of zoonosis. Therefore, it is imperative to investigate whether the parasites that affect these fish may represent a zoonotic risk for the inhabitants of the area. The objective of this study was to identify molecular and/or morphologically muscle parasites of fish from coastal locations in Tamaulipas, Mexico, and assess the risk of infection for humans. Between 2017 and 2018, 764 individual fish belonging to 28 species were examined for parasites. Collected worms were processed for their identification using morphological characteristics. In addition, partial sequences of the large subunit (28S) ribosomal RNA gene were obtained from some species to corroborate their identity. Prevalence and mean intensity of all registered infections were calculated. A total of seven species of parasites were found: cestodes (Poecilancistrium caryophyllum), trematodes (Clinostomum tataxumui, Clinostomum cichlidorum), nematodes (Eustrongylides sp., Contracaecum sp.) and pentastomids (Sebekia purdieae, Sebekia sp.). Parasites infected 10 species belonging to different fish families (Ariidae, Centrarchidae, Centropomidae, Cichlidae, Eleotridae, Ictaluridae, Mugilidae and Sciaenidae). Congeneric species of parasites or related to those registered in this study have been identified as zoonotic agents in other regions of the world. Despite the low levels of infection (2.6-16.6% prevalence and 1-5.5 parasites per infected host), there is a latent risk of transmission to humans, so it is recommended to avoid eating raw or undercooked fish meat.

First record of entomopathogenic nematodes from Yucatán State, México and their infectivity capacity against Aedes aegypti.

BACKGROUND: Biological control using entomopathogenic nematodes (EPN) has demonstrated good potential to contribute to the integral control of mosquito larvae, which as adults are vectors of diseases such as Dengue fever, Zika and Chikungunya. However, until now there are no records of the presence of EPN or their killing capacity in Yucatán state, southern México. The objectives of the current study were: (1) to report the entomopathogenic nematodes present in Yucatán soils and (2) to determine the killing capacity of the most frequent and abundant EPN against Aedes aegypti mosquito larvae and the microbial community developed by Ae. Aegypti exposed to this EPN. METHODS: The nematodes were collected by the insect trap technique using the great wax moth Galleria mellonella. Internal transcribed spacer (ITS), 28S gene of ribosomal DNA and phylogenetic analyses were performed to identify the EPN. For the bioassay, four concentrations of the most frequent and abundant EPN were tested: 1,260:1 infective juveniles (IJs) per mosquito larvae, 2,520 IJs:1, 3,780 IJs:1 and 5,040 IJs:1. High-throughput sequencing of the 16S rRNA gene was used to identify bacterial amplicon sequences in the mosquito larvae infected with EPN. RESULTS: Six isolates of Heterorhabditis were recovered from 144 soil samples. Heterorhabditis indica (four isolates) was the most frequent and abundant EPN, followed by Heterorhabditis n. sp. (two isolates). Both nematodes are reported for the first time for Yucatán state, Mexico. The concentration of 2,520 IJs:1 produced 80% of mosquito larvae mortality in 48 h. Representative members of Photorhabdus genus were numerically dominant (74%) in mosquito larvae infected by H. indica. It is most likely that these bacteria produce secondary toxic metabolites that enhance the mortality of these mosquito larvae.

Exploring the genetic structure of Parastrigea diovadena Dubois and Macko, 1972 (Digenea: Strigeidae), an endoparasite of the white ibis, Eudocimus albus, from the Neotropical region of Mexico.

Parastrigea diovadena Dubois and Macko, 1972, is an allogenic trematode species that infects the intestine of white ibis. This widely distributed Neotropical species has been studied poorly, and nothing is known about its population genetic structure. In the current study, we attempt to fill this gap for the first time and to explore the genetic diversity in P. diovadena populations from three biogeographic provinces (Sierra Madre Oriental, Sierra Madre Occidental, and Sierra Madre del Sur) in the Neotropical region of Mexico. Newly generated sequences of the internal transcribed spacers (ITS) from ribosomal DNA and cytochrome c oxidase subunit 1 (cox 1) from mitochondrial DNA were compared with sequences available from the GenBank data set. Phylogenetic analyses performed with the ITS and cox 1 data sets using maximum likelihood and Bayesian inference unequivocally showed that new sequences of P. diovadena recovered from the white ibis formed a clade with other sequences of specimens previously identified as P. diovadena. The intraspecific genetic divergence among the isolates was very low, ranging from 0 to 0.38% for ITS and from 0 to 1.5% for cox 1, and in combination with the phylogenetic trees confirmed that the isolates belonged to the same species. The cox 1 haplotype network (star-shaped) inferred with 62 sequences revealed 36 haplotypes. The most frequent haplotype (H3, n = 18) corresponded to specimens from all the populations (except Tecolutla, Veracruz). In addition to the common haplotype, we identified four other shared haplotypes (H2, H9, H12, and H14) and 31 unique haplotypes (singlets). In addition, high haplotype diversity (Hd = 0.913), low nucleotide diversity (Pi = 0.0057), and null genetic differentiation or population structure (Fst = 0.0167) were found among the populations from the three biogeographic provinces. The results suggest that the biology of the definitive host has played a key role in the population genetic structure of Parastrigea diovadena in the Neotropical region of Mexico.

Morphological and molecular identification of helminths of the greater bulldog bat Noctilio leporinus (Quiroptera: Noctilionidae) from Campeche, Mexico.

Surveys on parasites of bats from the Americas have been conducted, but information on helminths is still scarce, especially in the Neotropical region. In Mexico, there are species of bats that lack of a record for helminth species, such as members of the family Noctilionidae. The present study describes for the first time the helminths of Noctilio leporinus in Campeche, Mexico. In 2017, six specimens of N. leporinus were studied for helminths. The species identification of helminths was based on morphological studies and molecular analysis of fragments of the 28S rDNA. All bat specimens were infected for at least one helminth species. Three helminth taxa were identified: the trematode Pygidiopsis macrostomum, and the nematodes Tricholeiperia cf. proencai, and Heligmonellidae gen. sp. The morphological identification of P. macrostomum was confirmed by sequence analysis of 28S rDNA gene. The phylogeny of P. macrostomum grouped our sequence with other sequences of the same species collected in Brazil. The phylogenetic tree of Heligmonellidae gen. sp. indicated that the helminth belongs to clade formed by the species Odilia bainae, Nippostrongylus magnus and Nippostrongylus brasiliensis of the family Heligmonellidae. The phylogenetic analysis of the 28S sequences of T. cf. proencai did not show any similarity or close affinity with nematodes from which that gene has been sequenced to date. The findings of the present study increase the number of helminth species parasitizing bats in Mexico.

A new species of Neoheterobothrium Price, 1943 (Monogenea, Diclidophoridae) from Syacium papillosum (Linnaeus) (Pleuronectiformes, Paralichthyidae) in the Yucatan Shelf, with notes on the validity of the subfamilies in the Diclidophoridae.

Neoheterobothrium papillosum n. sp. (Monogenea: Diclidophoridae) is described based on specimens collected from the gills of the dusky flounder Syacium papillosum (Linnaeus) (Paralichthyidae) in the Yucatan Shelf. The new species can be differentiated from other congeneric species by the following characteristics: (1) number of hooks in the genital atrium (8 hooks), (2) number of testes (20-34), (3) pharynx size, (4) peduncle length, and (5) egg size. Neoheterobothrium papillosum n. sp. resembles Neoheterobothrium syacii; however, detailed morphological analysis allowed for their separation into two different species. Sequences of the nuclear ribosomal DNA partial 28S and ITS1 were obtained and compared with available sequences of other species and genera of the family Diclidophoridae from GenBank. A phylogenetic analysis was conducted, including 33 sequences, 18 of which represented 17 species and 11 genera of the family Diclidophoridae. The phylogenetic tree showed that Diclidophoridae is a monophyletic family and species of Diclidophora were grouped as the sister group of Neoheterobothrium papillosum n. sp. Our phylogenetic analyses were consistent with Mamaev's hypothesis, who divided Diclidophoridae into Diclidophorinae and Choricotylinae and concluded that Neoheterobothrium was a genus belonging to Diclidophorinae.

Morphological and molecular identification of hymenolepidid cestodes in children and synanthropic rodents from rural Mexico.

Hymenolepidid cestodes of synanthropic rodents represent a risk for public health. In order to describe the occurrence of hymenolepidids in children and the role of rodents as a potential source of infection, we conducted a morphological and molecular survey on cestodes in two rural villages from Yucatan, Mexico. One hundred and thirty-five stool samples from children (64 from Paraíso and 71 from Xkalakdzonot), 233 Mus musculus (159 from Paraíso and 74 from Xkalakdzonot) and 125 Rattus rattus (7 from Paraíso and 118 from Xkalakdzonot) were analyzed for the presence of cestodes. Three hymenolepidid species were identified morphologically: Hymenolepis nana in 7.8% of children from Paraíso, Hymenolepis microstoma in 4.4% of M. musculus from Paraíso and Hymenolepis diminuta in 15.3% of R. rattus from Xkalakdzonot. The molecular characterization and phylogenetic analysis based on mitochondrial cytochrome c subunit 1 (CO1) gene and ribosomal internal transcribed spacer 1 (ITS1) region, confirmed the identity of the three cestodes isolated from Yucatan. Phylogeny of the CO1 gene identified intraspecific genetic differences within H. nana ranging from 0 to 5%, in H. microstoma from 0 to 0.4%, and in H. diminuta ranged from 0 to 6.5% which suggests, the presence of complex species within H. nana and H. diminuta infecting humans and rodents, as reported by other authors. Based on the morphological and molecular results, and the epidemiological evidence, infections with H. nana suggest a non-zoonotic transmission; however, the presence of H. microstoma and H. diminuta in synanthropic rodents serve as a possible source for human infection.

Metazoan parasite infracommunities of the dusky flounder (Syacium papillosum) as bioindicators of environmental conditions in the continental shelf of the Yucatan Peninsula, Mexico.

BACKGROUND: We assessed metrics of the metazoan parasite infracommunities of the dusky flounder (Syacium papillosum) as indicators of aquatic environmental health of the Yucatan Shelf (YS) prior to oil extraction. We sampled the dusky flounder and its parasites along the YS, mostly during the 2015 north wind season (November-April). Our aims were: (i) to determine whether the parasite infracommunity metrics of S. papillosum exhibit significant differences among YS subregions; (ii) to determine whether the probability of the occurrence of its parasite species and individuals were affected by environmental variables, nutrients, heavy metals and hydrocarbons at the seascape level; and (iii) to determine whether there were statistical differences between the parasite infracommunity metrics of S. papillosum from YS and those of Syacium gunteri from the Campeche Sound. Multivariate statistical analyses and generalised additive models (GAMs) were used to examine the potential statistical associations between the contaminants, environmental variables and parasite community metrics, and the maximum entropy algorithm (MaxEnt) was used to characterise the habitat's suitability for the parasite's probability of occurrence. RESULTS: We recovered 48 metazoan parasite species from 127 S. papillosum, with larval cestodes and digeneans being the most numerically-dominant. Multivariate analyses showed significant differences in parasite infracommunity metrics among Western YS, Mid YS and Caribbean subregions, with the latter being the richest in species but not in individuals. The GAM and MaxEnt results indicated a negative effect of top predators (e.g. sharks and rays) removal on parasite metrics. The parasite infracommunities of S. papillosum were twice as rich in the number of species and individuals as those reported for S. gunteri from the Campeche Sound. CONCLUSIONS: The significant differences among subregions in parasite metrics were apparently due to the interruption of the Yucatan current during the north wind season. The fishing of top predators in combination with an influx of nutrients and hydrocarbons in low concentrations coincides with an increase in larval cestodes and digeneans in S. papillosum. The dusky flounder inhabits a region (YS) with a larger number of metazoan parasite species compared with those available for S. gunteri in the Campeche Sound, suggesting better environmental conditions for transmission in the YS.

Taxonomic status of Rhabdochona ictaluri (Nematoda: Rhabdochonidae) based on molecular and morphological evidence.

The genus Rhabdochona includes more than 100 species infecting freshwater fishes in all zoogeographical regions of the world. In Mexico, 12 nominal species of Rhabdochona have been recorded. Of these, Rhabdochona ictaluri was originally described as a parasite of endemic catfishes of the family Ictaluridae; however, the species was later considered on morphological grounds as a junior synonym of Rhabdochona kidderi. In this study, newly sampled specimens of R. ictaluri were obtained from the type host and type locality and were used to perform a detailed morphological analysis and molecular phylogenetic inferences through one mitochondrial and two nuclear genes; data were used in an integrative taxonomy context to test the taxonomic status of R. ictaluri. This approach proved to be very useful to confirm the validity of this species, and robust species limits were established between these two putative species considering morphology, molecular data, host association, and biogeography.

Description of two new species of allocreadiid trematodes (Digenea: Allocreadiidae) in middle American freshwater fishes using an integrative taxonomy approach.

Integrative taxonomy uses several sources of information to establish more robust species delimitation criteria. In this study, we followed that approach to describe two new species of allocreadiid trematodes from freshwater fish distributed in locations across Middle America. The new species were first recognized by using sequences of the domains D1-D3 of the ribosomal 28S rRNA gene and then, morphological data (light and scanning electron microscopy (SEM)), host association, and geographical distribution were considered as additional sources of information to complement the species delimitation and description. Auriculostoma tica n. sp. was found in the intestine of Gymnotus maculosus Albert and Miller in Costa Rica, while Wallinia anindoi n. sp. was found in the intestine of Astyanax aeneus (Günther) in Oaxaca and Chiapas, Mexico, as well as in a location of Guatemala. A phylogenetic analysis combining newly generated sequences of the two new species along with those available for other allocreadiids in GenBank, revealed that isolates of each species are reciprocally monophyletic, and also their interrelationships: Au. tica n. sp. is a sister taxon of Auriculostoma totonacapanensis Razo-Mendivil et al., 2014, and W. anindoi n. sp. is a sister taxon of Wallinia brasiliensis Dias, et al., 2018. Genetic distances for the 28S rRNA gene were estimated among the American species of allocreadiids and further supported the validity of the new species.

Phylogenetic Position of Pseudosellacotyla lutzi (Freitas, 1941) (Digenea: Cryptogonimidae), A Parasite of Hoplias malabaricus (Bloch) in South America, through 28S rDNA Sequences, and New Observations of the Ultrastructure of Their Tegument.

The freshwater fish digenean Pseudosellacotyla lutzi ( Freitas, 1941 ) Yamaguti, 1954 has had an unsettled taxonomic history, and has at various times been classified as a member of Nanophyetidae, Heterophyidae, Microphallidae, Faustulidae, and Cryptogonimidae. Nine individual specimens of the trahira, Hoplias malabaricus (Bloch, 1794), were sampled in the Paraná River basin, Paraná State, Brazil; 22 specimens of P. lutzi were collected. One specimen of P. lutzi was used to obtain a sequence of the domains D1-D3 of the 28S rRNA gene, and to perform a phylogenetic analysis to assess their position and classification within Plagiorchiida. The resulting tree unequivocally shows that the species, along with acanthostomines, belong to the Cryptogonimidae, corroborating recent findings based on the morphology of the cercariae, and in the characteristics of the life cycle. In addition, the study of the ultrastructure of the tegumental spines through scanning electron microscopy allowed us to characterize them as pectinate spines possessing 3 to 8 digitiform projections at their distal end and extending from the anterior to the posterior extremity of the body. This study also provides the first molecular data for a cryptogonimid from South America.

Filling the gaps in the classification of the Digenea Carus, 1863: systematic position of the Proterodiplostomidae Dubois, 1936 within the superfamily Diplostomoidea Poirier, 1886, inferred from nuclear and mitochondrial DNA sequences.

The Diplostomida Olson, Cribb, Tkach, Bray & Littlewood, 2003 is the less diverse order of the two orders within the subclass Digenea Carus, 1863 and is currently classified into three superfamilies, i.e. Brachylaimoidea Joyeux & Foley, 1930, Diplostomoidea Poirier, 1886, and Schistosomatoidea Stiles & Hassall, 1898. Although the suprageneric-level relationships have been elucidated with the use of molecular markers, the lack of representation of some groups obscure the phylogenetic relationships among families, rendering the classification unstable. Here, we tested the phylogenetic position of the family Proterodiplostomidae Dubois, 1936 based on partial 28S rDNA and complete 18S rDNA sequences for Crocodilicola pseudostoma (Willemoes-Suhm, 1870), a crocodile parasite that has been found as a progenetic metacercaria parasitising the pale catfish Rhamdia guatemalensis (Günther) in Mexico and in other siluruforms in the Neotropics. We augmented the representation of the species, genera and families within the Diplostomida, including mostly representatives of the superfamily Diplostomoidea, and assembled a dataset that contains 49 species for the 28S rRNA gene, and 45 species for the 18S rRNA gene. Additionally, we explored the phylogenetic signal of the mitochondrial gene cox1 in reconstructing the phylogenetic relationships of selected members of the superfamily. Our analyses showed that the family Proterodiplostomidae is the sister taxon to the paraphyletic Diplostomidae Poirier, 1886 and Strigeidae Railliet, 1919, with Cyathocotylidae Mühling, 1898 + Brauninidae Wolf, 1903 as their sister group. Analysis of concatenated 18S + 28S sequences revealed the Liolopidae Odhner, 1912 as the basal group of the superfamily Diplostomoidea, although analyses of independent datasets showed that the position of this family remains uncertain. Analysis based on cox1 unequivocally resolved the Proterodiplostomidae as the sister taxon to the Diplostomidae and Strigeidae, although the Cyathocotylidae was nested in a different clade, along with brachylaimoids and schistosomatoids.