Steven A. Nadler

Phylogenetic relationships of Steinernema Travassos, 1927 (Nematoda: Cephalobina: Steinernematidae) based on nuclear, mitochondrial and morphological data

Entomopathogenic nematodes of the genus Steinernema are lethal parasites of insects that are used as biological control agents of several lepidopteran, dipteran and coleopteran pests. Phylogenetic relationships among 25 Steinernema species were estimated using nucleotide sequences from three genes and 22 morphological characters. Parsimony analysis of 28S (LSU) sequences yielded a well-resolved phylogenetic hypothesis with reliable bootstrap support for 13 clades. Parsimony analysis of mitochondrial DNA sequences (12S rDNA and cox 1 genes) yielded phylogenetic trees with a lower consistency index than for LSU sequences, and with fewer reliably supported clades. Combined phylogenetic analysis of the 3-gene dataset by parsimony and Bayesian methods yielded well-resolved and highly similar trees. Bayesian posterior probabilities were high for most clades; bootstrap (parsimony) support was reliable for approximately half of the internal nodes. Parsimony analysis of the morphological dataset yielded a poorly resolved tree, whereas total evidence analysis (molecular plus morphological data) yielded a phylogenetic hypothesis consistent with, but less resolved than trees inferred from combined molecular data. Parsimony mapping of morphological characters on the 3-gene trees showed that most structural features of steinernematids are highly homoplastic. The distribution of nematode foraging strategies on these trees predicts that S. hermaphroditum, S. diaprepesi and S. longicaudum (US isolate) have cruise forager behaviours.

Molecular characterization and phylogeny of anisakid nematodes from cetaceans from southeastern Atlantic coasts of USA, Gulf of Mexico, and Caribbean Sea

In the present study, 407 anisakid nematodes, collected from 11 different species of cetaceans of the families Delphinidae, Kogiidae, Physeteridae, and Ziphiidae, from the southeastern Atlantic coasts of USA, the Gulf of Mexico, and the Caribbean Sea, were examined morphologically and genetically characterized by PCR restriction fragment length polymorphism to identify them to species level, assess their relative frequencies in definitive hosts, and determine any host preference. Sequence data from nuclear ribosomal internal transcribed spacer and mitochondrial cox2 genes were analysed by maximum parsimony and Bayesian inference methods, as separate and combined datasets, to evaluate phylogenetic relationships among taxa. The results revealed a highly diverse ascaridoid community. Seven Anisakis species and Pseudoterranova species were recovered as adult parasites. Larval forms of Contracaecum multipapillatum were also found in a coastal population of bottlenose dolphins. The phylogenetic trees obtained from the combined dataset (and most individual datasets) revealed the existence of distinct clades, the first including species of the Anisakis simplex complex (A. simplex s.s., Anisakis pegreffii, A. simplex C), (Anisakis nascettii, Anisakis ziphidarum) and the second including Pseudoterranova ceticola ((Anisakis paggiae, (Anisakis physeteris, Anisakis brevispiculata)). This finding, excluding the relationship of P. ceticola, is consistent with the morphology of adult and larval specimens. Considering the presence versus absence of an intestinal cecum, the relationship of P. ceticola with the members of the second clade of Anisakis appears inconsistent with morphological evidences but consistent with host preference. The position of Anisakis typica as the sister group to the two main anisakid clades indicates that it represents a third distinct lineage.

Molecular characterization and phylogeny of whipworm nematodes inferred from DNA sequences of cox1 mtDNA and 18S rDNA.

A molecular phylogenetic hypothesis is presented for the genus Trichuris based on sequence data from the mitochondrial cytochrome c oxidase 1 (cox1) and ribosomal 18S genes. The taxa consisted of different described species and several host-associated isolates (undescribed taxa) of Trichuris collected from hosts from Spain. Sequence data from mitochondrial cox1 (partial gene) and nuclear 18S near-complete gene were analyzed by maximum likelihood and Bayesian inference methods, as separate and combined datasets, to evaluate phylogenetic relationships among taxa. Phylogenetic results based on 18S ribosomal DNA (rDNA) were robust for relationships among species; cox1 sequences delimited species and revealed phylogeographic variation, but most relationships among Trichuris species were poorly resolved by mitochondrial sequences. The phylogenetic hypotheses for both genes strongly supported monophyly of Trichuris, and distinct genetic lineages corresponding to described species or nematodes associated with certain hosts were recognized based on cox1 sequences. Phylogenetic reconstructions based on concatenated sequences of the two loci, cox1 (mitochondrial DNA (mtDNA)) and 18S rDNA, were congruent with the overall topology inferred from 18S and previously published results based on internal transcribed spacer sequences. Our results demonstrate that the 18S rDNA and cox1 mtDNA genes provide resolution at different levels, but together resolve relationships among geographic populations and species in the genus Trichuris.

Review of research on hookworms (Uncinaria lucasi Stiles, 1901) in northern fur seals (Callorhinus ursinus Linnaeus, 1758)

The objective of this article is to review knowledge on the hookworm Uncinaria lucasi Stiles, 1901 in northern fur seals, Callorhinus ursinus Linnaeus, 1758. Emphasis is placed on research on this host–parasite system in the Pribilof Islands, AK, USA where the bulk of the studies has been performed.

First report and characterization of adult Uncinaria spp. in New Zealand Sea Lion ( Phocarctos hookeri ) pups from the Auckland Islands, New Zealand

Two species of hookworms (Uncinaria lucasi and Uncinaria hamiltoni) have been formally described from pinnipeds, but dissimilar types are noted from these hosts. This report is the first description of hookworms (Uncinaria spp.) from the New Zealand sea lion, Phocarctos hookeri. The nematodes were collected from dead pups on Enderby Island (Auckland Islands, 50°30, 166°17) during January and February, 2004. Standard measurements of male and female hookworms were obtained, providing a general morphometric characterization of the hookworm species in P. hookeri. Considerable variations in the body length of adult hookworms were noted within the same host. The arrangement of some of the bursal rays differs from that described for U. lucasi and U. hamiltoni.

Investigations of peritoneal and intestinal infections of adult hookworms (Uncinaria spp.) in northern fur seal (Callorhinus ursinus) and California sea lion (Zalophus californianus) pups on San Miguel Island, California (2003)

The peritoneal cavity (PNC) and intestine of northern fur seal (Callorhinus ursinus) pups and California sea lion (Zalophus californianus) pups that died in late July and early August, 2003, on San Miguel Island, California, were examined for hookworms. Prevalence and morphometric studies were done with the hookworms in addition to molecular characterization. Based on this and previous molecular studies, hookworms from fur seals are designated as Uncinaria lucasi and the species from sea lions as Uncinaria species A. Adult hookworms were found in the PNC of 35 of 57 (61.4%) fur seal pups and of 13 of 104 (12.5%) sea lion pups. The number of hookworms located in the PNC ranged from 1 to 33 (medianu2009=u20093) for the infected fur seal pups and 1 to 16 (medianu2009=u20092) for the infected sea lion pups. In addition to the PNC, intestines of 43 fur seal and 32 sea lion pups were examined. All of these pups were positive for adult hookworms. The worms were counted from all but one of the sea lion pups. Numbers of these parasites in the intestine varied from 3 to 2,344 (medianu2009=u2009931) for the fur seal pups and 39 to 2,766 (medianu2009=u2009643) for the sea lion pups. Sea lion pups with peritoneal infections had higher intensity infections in the intestines than did pups without peritoneal infections, lending some support for the hypothesis that peritoneal infections result from high-intensity infections of adult worms. There was no difference in intestinal infection intensities between fur seal pups with and without peritoneal infections. Female adult hookworms in the intestines of both host species were significantly larger than males, and sea lion hookworms were larger than those in fur seals. Worms in the intestine also were larger than worms found in the PNC. Gene sequencing and (RFLP) analysis of (PCR) amplified (ITS) ribosomal DNA were used to diagnose the species of 172 hookworms recovered from the PNC and intestine of 18 C. ursinus and seven Z. californianus hosts. These molecular data revealed that U. lucasi (hookworm of C. ursinus) and Uncinaria species A (of Z. californianus) infrequently mature in the intestine of the opposite host species in California rookeries. However, there is no support from molecular data for the hypothesis that cross-infection with “the wrong” Uncinaria species is a contributing factor in these cases of host peritonitis. The major significance of this research is the unusual finding of adult hookworms in the PNC of so many dead pups. No obvious explanation for this occurrence could be determined. Further research, like in the present study, should help understand and monitor the apparent ever changing role of hookworm disease in the health of northern fur seal and California sea lion pups on SMI.

Nuclear and mitochondrial genes for inferring Trichuris phylogeny

Nucleotide sequences of the triose phosphate isomerase (TPI) gene (624xa0bp) and mitochondrial cytochrome b (cob) gene (520xa0bp) were obtained by PCR and evaluated for utility in inferring the phylogenetic relationships among Trichuris species. Published sequences of one other nuclear gene (18S or SSU rRNA, 1816–1846xa0bp) and one additional mitochondrial (mtDNA) gene (cytochrome oxidase 1, cox1, 342xa0bp) were also analyzed. Maximum likelihood and Bayesian inference methods were used to infer phylogenies for each gene separately but also for the combined mitochondrial data (two genes), the combined nuclear data (two genes), and the total evidence (four gene) dataset. Few Trichuris clades were uniformly resolved across separate analyses of individual genes. For the mtDNA, the cob gene trees had greater phylogenetic resolution and tended to have higher support values than the cox1 analyses. For nuclear genes, the SSU gene trees had slightly greater resolution and support values than the TPI analyses, but TPI was the only gene with reliable support for the deepest nodes in the tree. Combined analyses of genes yielded strongly supported clades in most cases, with the exception of the relationship among Trichuris clades 1, 2, and 3, which showed conflicting results between nuclear and mitochondrial genes. Both the TPI and cob genes proved valuable for inferring Trichuris relationships, with greatest resolution and support values achieved through combined analysis of multiple genes. Based on the phylogeny of the combined analysis of nuclear and mitochondrial genes, parsimony mapping of definitive host utilization depicts artiodactyls as the ancestral hosts for these Trichuris, with host-shifts into primates, rodents, and Carnivora.

Panagrobelus stammeri Rühm, 1956 and Plectonchus hunti n. sp.: Implications of new morphological observations for characterisation of these genera (Nematoda: Panagrolaimoidea)

A detailed morphological description (light and scanning electron microscope observations) is given of two isolates currently used in molecular and developmental studies: PDL0024, Panagrobelus stammeri Ruhm, 1956 and PDL0025, a new species herein described as Plectonchus hunti n. sp. We redescribe the lip region, interpret the orientation of the lips in P.stammeri and provide additional morphological and morphometric diagnostic features. Plectonchus hunti n. sp. is characterised by the bifurcating spicule tip and by the placement of the male genital papillae in a 3/3 + p + 2 arrangement. Additionally, females of this new species can be differentiated from other species of Plectonchus by the more anterior position of the vulva, the more anterior location of the excretory pore and the tail morphology. An emended diagnosis for this genus is provided. In this study we demonstrate that both studied taxa share morphological similarities, such as the presence of a cephaloboid-type spermatheca. Our observations also indicate slight but consistent differences in cell composition of the female reproductive system between these two taxa.

The complete mitochondrial genome of Koerneria sudhausi (Diplogasteromorpha: Nematoda) supports monophyly of Diplogasteromorpha within Rhabditomorpha

Testing hypotheses of monophyly for different nematode groups in the context of broad representation of nematode diversity is central to understanding the patterns and processes of nematode evolution. Herein sequence information from mitochondrial genomes is used to test the monophyly of diplogasterids, which includes an important nematode model organism. The complete mitochondrial genome sequence of Koerneria sudhausi, a representative of Diplogasteromorpha, was determined and used for phylogenetic analyses along with 60 other nematode species. The mtDNA of K. sudhausi is comprised of 16,005xa0bp that includes 36 genes (12 protein-coding genes, 2 ribosomal RNA genes and 22 transfer RNA genes) encoded in the same direction. Phylogenetic trees inferred from amino acid and nucleotide sequence data for the 12 protein-coding genes strongly supported the sister relationship of K. sudhausi with Pristionchus pacificus, supporting Diplogasteromorpha. The gene order of K. sudhausi is identical to that most commonly found in members of the Rhabditomorphaxa0+xa0Ascaridomorphaxa0+xa0Diplogasteromorpha clade, with an exception of some tRNA translocations. Both the gene order pattern and sequence-based phylogenetic analyses support a close relationship between the diplogasterid species and Rhabditomorpha. The nesting of the two diplogasteromorph species within Rhabditomorpha is consistent with most molecular phylogenies for the group, but inconsistent with certain morphology-based hypotheses that asserted phylogenetic affinity between diplogasteromorphs and tylenchomorphs. Phylogenetic analysis of mitochondrial genome sequences strongly supports monophyly of the diplogasteromorpha.

Review paper: The evolutionary relationships of rotifers and acanthocephalans

Advances in morphological and molecular studies of metazoan evolution have led to a better understanding of the relationships among Rotifera (Monogononta, Bdelloidea, Seisonidea) and Acanthocephala, and their relationships to other bilateral animals. The most accepted morphological analysis places Acanthocephala as a sister group to Rotifera, although other studies have placed Acanthocephala as a sister taxon to Bdellodea or Seisonidea. Molecular analyses using nuclear 18S rRNA and mitochondrial 16S rRNA genes support Acanthocephala as a sister taxon to Bdelloidea, although no molecular data is available for Seisonidea. Combining molecular and morphological analyses of Bilateria leads to a tree with Platyhelminthes, Rotifera, Acanthocephala and Gnathostomulida (and probably Gastrotricha) as a sister group to the annelid-mollusc lineage of the Spiralia (Lophotrochozoa).