Biomedical subjects

N B Chilton

Publications and source records attributed to N B Chilton.

At least 19 recordsLinked to original sources

PubMed2006

Isolation and characterization of class II myosin genes from Haemonchus contortus.

In this study, cDNAs encoding myosin from the parasitic nematode Haemonchus contortus were isolated and characterized. Several exhibited a considerable degree of sequence variation at the nucleotide and limited divergence at the amino acid levels within the various functional domains. The results suggest that the cDNAs isolated represented a single myosin heavy chain, which, by comparison with a number of other myosins, is inferred to represent a homologue of a muscle myosin (CeMHCA) of the free-living nematode Caenorhabditis elegans. The findings could have implications for investigating cytoskeletal dynamics and/or signalling pathways.

Amino Acid Sequence↗

PubMed2006

Class II myosins in nematodes-- genetic relationships, fundamental and applied implications.

Myosins are represented by a wide range of different classes of molecule, of which the most extensively studied are the class II myosins which drive muscle contraction and cell organization; the functional unit of class II myosins comprises two myosin heavy chains (MHCs). This minireview gives an update on class II MHCs of nematodes and describes a comparative analysis of MHC genes from nematodes and other organismal groups. Genetic analyses of sequence data for the four functional domains of MHCs (i.e., the SH3-like N-terminal, head, neck and tail domains) reveal a delineation between both the nematode and non-nematode myosins and between muscle and non-muscle myosins. The distinctiveness of the MHCs of nematodes suggests functional and tissue specialization. The elucidation of the functional roles of myosins and other molecules in specific signaling pathways in nematodes has the potential to lead to new intervention strategies for parasites via the specific disruption or interruption of key developmental processes, having biotechnological implications in the longer term.

Amino Acid Sequence↗

PubMed2005

Genetic variation in the mitochondrial cytochrome c oxidase subunit 1 within three species of Progamotaenia (Cestoda: Anoplocephalidae) from macropodid marsupials.

Sequence variation within 3 morphologically defined species of the anoplocephalid cestode genus Progamotaenia (P. ewersi, P. macropodis and P. zschokkei) was investigated using the cytochrome c oxidase subunit 1 gene. The magnitude of genetic variation detected within each morphospecies suggests that, in each instance, several cryptic species are present. Within P. ewersi, 5 genetically distict groups of cestodes were detected, 1 shared by Macropus robustus and M. parryi in Queensland, 1 in M. agilis from Queensland, 1 in Petrogale assimilis from Queensland, 1 in Macropus fuliginosus from South Australia and 1 in Wallabia bicolor from Victoria. In P. macropodis, cestodes from M. robustus from Queensland, Western Australia and the Northern Territory, M. parryi from Queensland and M. eugenii from South Australia were genetically distinct from those in Wallabia bicolor from Queensland and Victoria and from M. fuliginosus from South Australia. P. zschokkei consisted of a number of genetically distinct groups of cestodes, 1 in Lagorchestes conspicillatus and L. hirsutus from Queensland and the Northern Territory respectively, 1 in Petrogale herberti, P. assimilis and M. dorsalis from Queensland, 1 in Onychogalea fraenata from Queensland, 1 in M. agilis from Queensland and 1 in Thylogale stigmatica and T. thetis from Queensland. In general, genetic groups within each morphospecies were host specific and occurred predominantly in a particular macropodid host clade. Comparison of genetic relationships of cestodes with the phylogeny of their hosts revealed examples of colonization (P. zschokkei in M. agilis) and of host switching (P. zschokkei in M. dorsalis).

Amino Acid Sequence↗

PubMed2005

Molecular and morphological evidence indicates that Pseudorhabdosynochus lantauensis (Monogenea: Diplectanidae) represents two species.

Sequences of the first internal transcribed spacer (ITS-1) and the D1-D3 domains of the large subunit (LSU) of the ribosomal DNA (rDNA) were determined for multiple specimens of 4 operational taxonomic units (OTUs) of the monogenean, Pseudorhabdosynochus lantauensis. OTUs were defined based on their collecting localities, host and/or morphological characteristics. All P. lantauensis specimens of one group (OTUs 1 and 3) differed in their sequences of the ITS-1 and partial LSU rDNA when compared with specimens of a second group (OTUs 2 and 4) by 12% and 2%, respectively. Results of the phylogenetic analyses of the LSU rDNA sequence data showed total (100%) bootstrap support for the separation of P. lantauensis into 2 distinct clades. At least 11 of the 18 nucleotide differences in the LSU sequence between the two P. lantauensis clades were derived (i.e. autapomorphic) characters when the morphologically distinct species, P. epinepheli and P. coioidesis, were used as outgroups. Furthermore, there were several autapomorphic character states for each P. lantauensis clade. This provides sufficient evidence to reject the null hypothesis that P. lantauensis represents a single species. Morphological and morphometric differences between these two clades provided additional strong support for the separation of P. lantauensis into two species. These two parasite species were found to co-exist on one of the two species of serranid fish (i.e. Epinephelus coioides) examined in the South China Sea (Guangdong Province, China).

Animals↗

PubMed2004

Lack of evidence for co-speciation in a parasitic nematode of grey kangaroos.

Multilocus enzyme electrophoresis was used to compare specimens of the parasitic nematode Cloacina obtusa from the stomach of the eastern grey kangaroo, Macropus giganteus and the western grey kangaroo, M. fuliginosus. Allelic variation among nematodes was detected at 17 (85%) of 20 loci, but there was only a single fixed genetic difference (at the locus for isocitrate dehydrogenase, IDH) between C. obtusa from M. fuliginosus and those from M. giganteus in areas where each host occurred in allopatry. However, this fixed difference was not apparent within the zone of host sympatry. Although electrophoretic data indicate genetic divergence among allopatric populations of C. obtusa in the two host species, the magnitude of the electrophoretic difference (5%) between these populations does not refute the hypothesis that C. obtusa represents a single species. The 'usual' situation for parasitic helminths of grey kangaroos is that pairs of parasite species occur in the two host species. This situation differs for C. obtusa, where there has been a lack of speciation following a speciation event in its macropodid marsupial hosts. This finding suggests that a speciation event in the host does not necessarily lead to a speciation event for all its parasites and further highlights our lack of understanding of which processes drive speciation in parasites.

Animals↗

PubMed2003

Structure and organization of the mitochondrial genome of the canine heartworm, Dirofilaria immitis.

This study determined the complete mitochondrial (mt) genome sequence of the canine heartworm, Dirofilaria immitis, and compared its structure, organization and other characteristics with Onchocerca volvulus and other secernentean nematodes. The D. immitis mt genome is 13814 bp in size and contains 36 of the 37 genes typical of metazoan organisms, and lacks the ATP synthetase subunit 8 gene. All of the genes are transcribed in the same direction. For the entire genome, the nucleotide contents are approximately 55% (T), approximately 19% (each for A and G) and approximately 7% (C), which is very similar to those of the protein-coding genes. In the latter genes, most (approximately 69%) third codon positions have a T, but rarely (approximately 1-9%) have an A or a C. The C content (8-12%) is higher at the first and second codon positions compared with the third position (approximately 1%). These nucleotide biases have a significant effect on the codon usage patterns and, thus, on the amino acid composition of the proteins. The mt genome organization of D. immitis is essentially the same as that of O. volvulus, but is distinctly different from other secernentean nematodes sequenced thus far. Irrespective of transpositions of transfer RNA (trn) genes and the non-coding, AT-rich region, there are 4 gene- or gene block-translocations between the mt genome of D. immitis and those of Caenorhabditis elegans, Ascaris suum and the 2 human hookworms, Ancylostoma duodenale and Necator americanus. For D. immitis, the 22 trn genes have secondary structures typical of other secernentean nematodes, and possess a TV-replacement loop instead of a TpsiC arm and loop. Like O. volvulus, the mt trnK and trnP of D. immitis use the anticodons CUU and AGG, whereas in other nematodes, UUU and UGG are employed, respectively. Also, the secondary structures of the 2 ribosomal RNA (rrn) genes are similar to the models for other nematodes. Overall, the availability of the complete D. immitis mt genome sequence provides a resource for future studies of the comparative mt genomics and of the population genetics and/or phylogeny of parasitic nematodes.

Amino Acid Sequence↗

PubMed2002

Long PCR-based amplification of the entire mitochondrial genome from single parasitic nematodes.

Mitochondrial genome sequences provide useful markers for investigating population genetic structures because of their maternal inheritance and high evolutionary rates. There is, however, a paucity of information on mitochondrial genomes for many parasitic organisms, including nematodes, which appears to relate mainly to technical limitations and (for modestly funded laboratories) the cost associated with full mitochondrial genome sequencing. In this article, we describe a simple, relatively inexpensive long-PCR approach for the amplification (using two sets of primers) of the entire mitochondrial genome from individual parasitic nematodes for subsequent sequencing, which overcomes these limitations. We employed two species of human hookworm (Ancylostoma duodenale and Necator americanus; order Strongylida) to establish the long-PCR conditions, and then extended its use to a number of other species of parasitic nematode of the class Secernentea (orders Strongylida, Ascaridida and Rhabditida). The long-PCR method for the amplification of the entire mitochondrial genome from single nematodes, coupled with direct sequencing of amplicons, provides a useful tool for the comparative analysis of genome organisation and evolution of a range of nematode groups. It also creates a platform for molecular ecological and population genetic studies.

Ancylostoma↗

PubMed2002

Review of Papillostrongylus Johnston & Mawson, 1939 (Nematoda: Strongyloidea) from wallabies and kangaroos (Marsupialia: Macropodidae) using morphological and molecular techniques, with the description of P. barbatus n. sp.

The strongyloid nematode genus Papillostrongylus Johnston & Mawson, 1939, from kangaroos and wallabies, is reviewed using morphological and molecular methods. P. labiatus Johnston & Mawson, 1939 is re-described from material from the type-host, the black-striped wallaby Macropus dorsalis, from eastern Queensland, Australia, in which it is a relatively common parasite. Additional records from M. parryi and Thylogale thetis are confirmed and considered to represent examples of host-switching. A geographically disjunct population of the nematode species occurs in M. bernardus and Petrogale brachyotis in Arnhem Land, Northern Territory, but assessment of its status requires additional material. Nematodes from M. rufus, M. giganteus, M. fuliginosus and M. robustus from inland regions of Australia, formerly attributed to P. labiatus, are here assigned to a new species, P. barbatus, distinguished by the presence of an external leaf-crown, larger size, by greater spicule length in the male and by a sinuous vagina in the female. Additional hosts of P. barbatus n. sp. are Petrogale assimilis and Pet. lateralis purpureicollis. Sequence analyses of the second internal transcribed spacer of ribosomal DNA (ITS-2) also showed that P. barbatus n. sp. differed at 40 (16.7%) of the 240 alignment positions when compared with P. labiatus. Most of these interspecific sequence differences occurred in loops or bulges of the predicted precursor rRNA secondary structure, or represented partial or total compenstory base pair changes in stems.

Animals↗

PubMed2001

Applications of single-strand conformation polymorphism (SSCP) to taxonomy, diagnosis, population genetics and molecular evolution of parasitic nematodes.

The analysis of genetic variation in parasitic nematodes has important implications for studying aspects of taxonomy, diagnosis, population genetics, drug resistance and molecular evolution. This article highlights some applications of PCR-based single-strand conformation polymorphism (SSCP) for the analysis of sequence variation in individual parasites (and their populations) to address some of these areas. It also describes the principles and advantages of SSCP, and provides some examples for future applications in parasitology.

Animals↗

PubMed2001

Single-strand conformation polymorphism analysis of genetic variation in Labiostrongylus longispicularis from kangaroos.

Single-strand conformation polymorphism (SSCP) analysis was employed to screen for sequence heterogeneity in the second internal transcribed spacer (ITS-2) of ribosomal (r) DNA of Labiostrongylus longispicularis, a parasitic strongylid nematode occuring in some species of kangaroo in different geographical regions of Australia. The results showed that most of the nematodes screened had different SSCP profiles, which were subsequently shown to correspond to polymorphisms and/or an indel in the ITS-2 sequence. These variable sites related mainly to unpaired regions of the predicted secondary structure of the precursor rRNA molecule. SSCP profiles could be used to distinguish L. longispicularis in Macropus robustus robustus (New South Wales) from L. longispicularis in Macropus robustus erubescens and Macropus rufus (South Australia). This difference corresponded to a transversional change in the ITS-2 sequence at alignment position 82. The study demonstrated clearly the effectiveness of SSCP analysis for future large-scale population genetic studies of L. longispicularis in order to test the hypothesis that L. longispicularis from different geographical regions represents multiple sibling species.

Animals↗

PubMed2001

Evolutionary relationships of trichostrongyloid nematodes (Strongylida) inferred from ribosomal DNA sequence data.

The evolutionary relationships of 21 species of trichostrongyloid nematodes were determined by use of sequence data of the second internal transcribed spacer of the ribosomal DNA aligned according to secondary structure information. Irrespective of the method of analysis used, the topologies of the phylogenetic trees derived from the molecular data differed with respect to all four hypotheses proposed previously for the evolutionary relationships of the different subfamilies within the Trichostrongylidae based on morphological data. Thus, the molecular data set did not resolve the conflict between the four previous proposals for the subfamilial relationships. Nonetheless, all trees derived from the molecular data showed strong support for the exclusion of the genera Filarinema and Amidostomum from the clade containing the species within the family Trichostrongylidae. This represents a major difference from the most recent proposal of the systematics of the Trichostrongyloidea in which these two genera were included within the Trichostrongylidae. Therefore, the molecular data support an earlier systematic framework in which Filarinema and Amidostomum were considered to be sister groups of the Trichostrongyloidea.

Animals↗

PubMed2001

Co-evolutionary relationships between the nematode subfamily Cloacininae and its macropodid marsupial hosts.

Morphologically based phylogenies of the cloacinine genera Cyclostrongylus, Macropostrongylus, Pharyngostrongylus, Popovastrongylus, Rugopharynx, Thallostonema, Wallabinema, and Zoniolaimus were constructed and compared with the phylogeny of their respective macropodid hosts. These comparisons show some evidence of co-speciation. However, there was little consistency among trees of different nematode genera, parasite species were scattered amongst hosts and basal parasite taxa were, in some instances, parasitic in hosts belonging to derived clades. A cladistic analysis, using as characters 208 cloacinine nematode species found in 23 species of host, produced a tree largely resembling that of the host tree but with significant differences explainable by host switching among macropodids occupying similar habitat. Nematodes were moderately host-specific, but some species occurred in three or more distantly related host species indicating a degree of host switching. The results are more consistent with the hypothesis of a colonisation of macropodid hosts by cloacinine nematodes rather than a prolonged period of co-speciation although alternative interpretations of the data are also considered.

Animals↗

PubMed2001

Mutation scanning analysis of microsatellite variability in the second internal transcribed spacer (precursor ribosomal RNA) for three species of Metastrongylus (Strongylida: Metastrongyloidea).

This study investigated sequence variability in the second internal transcribed spacer of ribosomal DNA for 3 species of Metastrongylus (porcine lungworms). The ITS-2 region was amplified by PCR from individuals of M. elongatus, M. pudendotectus and M. salmi, and then subjected directly to single-strand conformation polymorphism analysis (SSCP), which allowed the direct display of sequence variation within and among individuals representing each species. There were marked differences in SSCP profiles among species, making this approach useful for species identification. For individual species, representative bands were excised from electrophoretic gels, reamplified by PCR and subjected to direct sequencing. For all 3 taxa, variability in the ITS-2 was related chiefly to the presence of microsatellites. Eight different microsatellites were identified, namely (A)n, (TG)n, (TCG)n, (TA)n, (TATG)n, (G)n, (TACA)n and (T)n. Considerable variability in microsatellite repeat number (ranging from 1 to 23) was found among individual nematodes of a species and between species. The microsatellites were located to specific stem or loop regions in the predicted ITS-2 precursor rRNA secondary structure. The results may suggest that slipped-strand mispairing in microsatellite regions contributes to sequence variability in the ITS-2 of Metastrongylus species under structural constraint as a consequence of microsatellite location in the precursor rRNA. Similar studies of the ITS-2 for a wide range of parasitic nematodes may lead to a better understanding of concerted evolution in these organisms.

Animals↗

PubMed2001

Molecular approaches for studying ascaridoid nematodes with zoonotic potential, with an emphasis on Toxocara species.

Species-specific identification of ascaridoid nematodes at any developmental stage is a prerequisite for detailed investigation of the life cycles, systematics and epidemiology of this important group, and is also crucial for the diagnosis of associated infections. The morphological identification of some species and/or their larval stages can, however, present considerable difficulty. Recently, PCR-based methods, using genetic markers in the internal transcribed spacers (ITS) of ribosomal DNA, have been shown to provide reliable alternatives to more traditional methods for the specific identification of nematodes. This article provides an account of recent research on the development of PCR-based methods (utilizing ITS sequences) for the specific identification of ascaridoid nematodes of zoonotic potential, for the diagnosis of infections, and for the analysis of genetic variation within and among individual nematodes and their populations. Prospects for using these diagnostic and analytical tools to investigate epidemiological and population genetic questions relating to ascaridoid parasites are also discussed.

Animals↗

PubMed2000

Relationships among some ascaridoid nematodes based on ribosomal DNA sequence data.

The nuclear ribosomal DNA (rDNA) region spanning the first (ITS-1) and second (ITS-2) internal transcribed spacers was sequenced for 15 taxa of ascaridoid nematodes. The length of the ITS-1 and ITS-2 sequences in the 15 taxa ranged from 392-500 bp and 240 348 bp, respectively. While nucleotide variation of 0-2.9% in the ITS-1 and/or ITS-2 sequences was detected within taxa where multiple samples were sequenced, significantly higher level of nucleotide difference (9.4-66.6%) was detected between the taxa, except for Ascaris suum and A. lumbricoides whose taxonomic status remains uncertain. These interspecific differences were linked with the considerable size differences (0-108 bp) in the rDNA spacers. Phenograms based on the genetic differences among the 15 taxa showed some concordance with previous classification schemes derived from morphological data.

Animals↗

PubMed2000

Influence of temperature and relative humidity on the moulting success of Amblyomma limbatum and Aponomma hydrosauri (Acari: Ixodidae) larvae and nymphs.

This study compared the duration of the moulting periods of engorged larvae and nymphs of the ixodid ticks, Amblyomma limbatum and Aponomma hydrosauri, at different temperature/relative humidity regimes, and examined the relationships between the engorged weight of ticks and their weights after moulting. The results showed that for each species, there was a significant relationship between the weights of unfed nymphs and engorged larvae, and the weights of unfed adults and engorged nymphs. The weight of engorged nymphs was also a good indicator of their sex, with female ticks having heavier weights as engorged nymphs. Temperature and relative humidity had a marked effect on the moulting success of engorged ticks of both species. Aponomma hydrosauri larvae and nymphs were able to moult at lower temperatures than Amb. limbatum but most ticks, except Ap. hydrosauri larvae, failed to moult at 13 degrees C. Additionally, there was a marked decrease in the pre-moult times of ticks at higher temperatures, with larvae taking less time to moult than nymphs. At temperatures greater than 21 degrees C, Amb. limbatum took less time to moult than Ap. hydrosauri but this interspecific difference was less marked for nymphs. The interspecific differences in the responses of engorged larvae and nymphs to different temperatures and relative humidities correlated with interspecific differences in off-host behaviour and with the different climates the two species experience throughout most of their distributional range.

Animals↗

PubMed2000

Genetic variation within the ticks Ixodes holocyclus and Ixodes cornuatus from south-eastern Australia.

Ticks from mainland Australia (Victoria, New South Wales and Queensland) and Tasmania, identified morphologically as either Ixodes holocyclus or Ixodes cornuatus, were compared genetically using 24 enzyme loci. The results showed that ticks from three localities in Victoria were genetically similar to I. cornuatus in Tasmania, but both groups had fixed genetic differences at >45% of loci compared with other ticks on the mainland. In addition, there were fixed genetic differences at 0-60% of loci among I. holocyclus from different localities on the mainland. Ixodes holocyclus samples could be divided into four distinct clusters (with fixed genetic differences >15%), three of which were represented by one or two specimens. Nonetheless, these electrophoretic data suggest that I. holocyclus represents a species complex. The results also showed that the morphological criteria used to identify specimens were not always accurate because several specimens had been mis-identified morphologically. Despite limitations with the morphological identification, this study has demonstrated that I. cornuatus can be distinguished from the I. holocyclus species complex using six enzyme loci, providing the foundation for a re-examination of morphological characteristics. The present study has shown that I. cornuatus and the I. holocyclus complexes have a greater distribution than previously reported, with both occurring in sympatry at Cape Patterson, on the southern coastline of Victoria.

Alleles↗

PubMed2000

A molecular systematic framework for equine strongyles based on ribosomal DNA sequence data.

In this study, molecular data sets were used to address the controversies relating to the systematics of strongyloid nematodes of equids utilising morphological data sets. DNA sequences of the first and second internal transcribed spacers (ITS-1 and ITS-2) of ribosomal DNA were determined for 30 species of equine strongyles and the systematic relationships reconstructed using phenetic and phylogenetic tree-building methods. The molecular data provided support for the hypothesis that the genera with large subglobular buccal capsules are ancestral to those with small cylindrical buccal capsules, but did not provide support for the current division of the subfamilies Strongylinae and Cyathostominae or for some taxonomic groupings (i.e. generic designations of species) within the Cyathostominea based on morphological data. Although not entirely concordant, the current molecular data provide a systematic framework for future studies of equine strongyles, which could be exploited in combination with new, phylogenetically informative morphological data sets.

Animals↗