A review of the Acanthocephala parasitising freshwater fishes in Australia.

The acanthocephalan fauna of Australian freshwater fishes was documented from field surveys, a literature survey and examination of specimens registered in Australian museums. From the 4030 fishes, representing 78 of the 354 Australian freshwater fish species (22%), examined for infection seven species of acanthocephalan were recovered. These species comprised five endemic species, three in endemic genera, two species in cosmopolitan genera, one species not fully identified and 1 putative exotic species recovered from eight species of fish. Of these Edmonsacanthus blairi from Melanotaenia splendida, was the only acanthocephalan found at a relatively high prevalence of 38·6%. These findings are indicative of a highly endemic and possibly depauperate acanthocephalan fauna. Species richness was higher in the tropical regions than the temperate regions of the country. Exotic acanthocephalan species have either not been introduced with their exotic hosts or have been unable to establish their life cycles in Australian conditions. Consequently, acanthocephalans have not yet invaded endemic Australian fish hosts.

First detection of Edwardsiella ictaluri (Proteobacteria: Enterobacteriaceae) in wild Australian catfish.

The bacterium Edwardsiella ictaluri is considered to be one of the most significant pathogens of farmed catfish in the United States of America and has also caused mortalities in farmed and wild fishes in many other parts of the world. E. ictaluri is not believed to be present in wild fish populations in Australia, although it has previously been detected in imported ornamental fishes held in quarantine facilities. In an attempt to confirm freedom from the bacterium in Australian native fishes, we undertook a risk-based survey of wild catfishes from 15 sites across northern Australia. E. ictaluri was detected by selective culturing, followed by DNA testing, in Wet Tropics tandan (Tandanus tropicanus) from the Tully River, at a prevalence of 0.40 (95% CI 0.21-0.61). The bacterium was not found in fishes sampled from any of the other 14 sites. This is the first report of E. ictaluri in wild fishes in Australia.

Cryptosporidium species in sheep and goats from Papua New Guinea.

Species of Cryptosporidium are extensively recognised as pathogens of domesticated livestock and poultry, companion animals, wildlife, and are a threat to public health. Little is known of the prevalence of Cryptosporidium spp. in humans, domesticated animals or wildlife in Papua New Guinea (PNG). The aim of the present study was to screen sheep and goats for Cryptosporidium using molecular tools. A total of 504 faecal samples were collected from sheep (n=276) and goats (n=228) in village, government and institutional farms in PNG. Samples were screened by nested PCR and genotyped at the 18S rRNA and at the 60kDa glycoprotein (gp60) loci. The overall prevalences were 2.2% for sheep (6/278) and 4.4% (10/228) for goats. The species/genotypes identified were Cryptosporidium hominis (subtype IdA15G1) in goats (n=6), Cryptosporidium parvum (subtypes IIaA15G2R1and IIaA19G4R1) in sheep (n=4) and in goats (n=2), Cryptosporidium andersoni (n=1) and Cryptosporidium scrofarum (n=1) in sheep, Cryptosporidium xiao (n=1) and Cryptosporidium rat genotype II (n=1) in goats. This is the first report of Cryptosporidium spp. identified in sheep and goats in PNG. Identification of Cryptosporidium in livestock warrants better care of farm animals to avoid contamination and illness in vulnerable population. The detection of zoonotic Cryptosporidium in livestock suggests these animals may serve as reservoirs for human infection.

Infection levels of gastrointestinal parasites in sheep and goats in Papua New Guinea.

Gastrointestinal parasites of livestock cause diseases of important socio-economic concern worldwide. The present study investigated the prevalence of gastrointestinal parasites in sheep and goats in lowland and highland regions of Papua New Guinea (PNG). Faecal samples were collected from a total of 165 small ruminants (110 sheep and 55 goats) from February to April 2011. Analysis by a modified McMaster technique revealed that 128 animals (72% of sheep and 89% of goats) were infected with one or more species of gastrointestinal parasites. The gastrointestinal parasites found and their prevalences in sheep (S) and in goats (G) were as follows: strongyle 67.3% (S), 85.5% (G); Eimeria 17.3% (S), 16.4% (G); Strongyloides, 8.2% (S), 23.6% (G); Fasciola, 5.5% (S), 18.2% (G); Trichuris, 1.8% (S), 3.6% (G); and Nematodirus, 1.8% (S), 3.6% (G). Two additional genera were found in goats: Moniezia (9.1%) and Dictocaulus (3.6%). This is the first study to quantitatively examine the prevalence of gastrointestinal parasites in goats in PNG. The high rates of parasitism observed in the present study are likely to be associated with poor farming management practices, including lack of pasture recovery time, lack of parasite control measures and poor-quality feed.

Parasites of native and exotic freshwater fishes in south-western Australia.

In this study, 1429 fishes of 18 different species (12 native and six exotic) were sampled from 29 localities to compare the levels of parasitism between native and exotic fish species and to examine the relationship between environmental degradation and parasite diversity. Forty-four putative species of parasites were found and most of these appear to be native parasites, which have not previously been described. Two parasite species, Lernaea cyprinacea and Ligula intestinalis, are probably introduced. Both were found on or in a range of native fish species, where they may cause severe disease. Levels of parasitism and parasite diversity were significantly greater in native fishes than in exotic species, and this may contribute to an enhanced demographic performance and competitive ability in invading exotics. Levels of parasitism and parasite diversity in native fishes were negatively related to habitat disturbance, in particular to a suite of factors that indicate increased human use of the river and surrounding environment. This was due principally to the absence in more disturbed habitats of a number of species of endoparasites with complex life cycles, involving transmission between different host species.

The diversity, distribution and host-parasite associations of trypanosomes in Western Australian wildlife.

Little is known regarding the diversity, distribution or host-parasite associations of Trypanosoma spp. in Australian wildlife. Here we report on an investigation based on divergence of the 18S rRNA gene of trypanosomes isolated from a range of hosts and varied geographical locations. A total of 371 individuals representing 19 species of native animals from 14 different locations were screened. In total, 32 individuals from 9 different species tested positive for the parasite. Phylogenetic analysis revealed considerable parasite diversity with no clear geographical distribution and no evidence of host specificity. In general, it appears that Australian Trypanosoma spp. are widespread, with several genotypes appearing in multiple host species and in varied locations including both mainland areas and offshore islands. Some host species were found to be susceptible to multiple genotypes, but no individuals were infected with more than a single isolate.

Trypanosomes in a declining species of threatened Australian marsupial, the brush-tailed bettong Bettongia penicillata (Marsupialia: Potoroidae).

The brush-tailed bettong (Bettongia penicillata), or woylie, is a medium-sized macropod marsupial that has undergone a rapid and substantial decline throughout its home range in the Upper Warren region of Western Australia over a period of approximately 5 years. As part of an investigation into possible causes of the decline a morphologically distinct Trypanosoma sp. was discovered by light microscopy in the declining population but was absent in a stable population within the Karakamia Wildlife Sanctuary. Further investigations employing molecular methods targeting variations in the 18s rRNA gene determined that the trypanosome was novel and was also present within the Karakamia population albeit at a much lower overall prevalence and individual parasitaemia levels. Phylogenetic analysis suggests the novel Trypanosoma sp. to be closely related to other trypanosomes isolated from native Australian wildlife species. Although it appears unlikely that the parasite is solely responsible for the decline in woylie population size, it may (singularly or in conjunction with other infectious agents) predispose woylies to increased mortality.

Phylogenetic Pattern, Evolutionary Processes and Species Delimitation in the Genus Echinococcus.

An accurate and stable alpha taxonomy requires a clear conception of what constitutes a species and agreed criteria for delimiting different species. An evolutionary or general lineage concept defines a species as a single lineage of organisms with a common evolutionary trajectory, distinguishable from other such lineages. Delimiting evolutionary species is a two-step process. In the first step, phylogenetic reconstruction identifies putative species as groups of organisms that are monophyletic (share a common ancestor) and exclusive (more closely related to each other than to organisms outside the group). The second step is to assess whether members of the group possess genetic exchangeability (where cohesion is maintained by gene flow among populations) or ecological exchangeability (where cohesion is maintained because populations occupy the same ecological niche). Recent taxonomic reviews have recognized nine species within the genus Echinococcus. Phylogenetic reconstructions of the relationships between these putative species using mtDNA and nuclear gene sequences show that for the most part these nine species are monophyletic, although there are important incongruences that need to be resolved. Applying the criteria of genetic and ecological exchangeability suggests that seven of the currently recognized species represent evolutionarily distinct lineages. The species status of Echinococcus canadensis and Echinococcus ortleppi could not be confirmed. Coalescent-based analyses represent a promising approach to species delimitation in these closely related taxa. It seems likely, from a comparison of sister species groups, that speciation in the genus has been driven by geographic isolation, but biogeographic scenarios are largely speculative and require further testing.

Finding genetic markers for complex phenotypic traits in parasites.

The identification, mapping and eventual cloning of genes which determine or influence important epidemiological traits in parasites can have great benefits for the control of parasitic disease. In this review, strategies are outlined for identifying genetic markers for complex, quantitative traits. A genetic marker is a variable DNA sequence which co-occurs with a variable quantitative trait. Candidate markers are chosen because they are thought to directly influence the trait whereas random markers are expected to be linked to another DNA sequence which influences the trait. Association studies compare the value of a quantitative trait between different marker genotype classes in a population, without regard to family structure. Linkage studies compare the value of a quantitative trait between marker genotype classes within families or within a population (usually derived from a cross between inbred lines) which is segregating for both marker and quantitative trait loci. The most commonly used analytical methods for determining the significance of association or linkage between marker and quantitative trait loci, and for estimating parameters such as recombination rate and quantitative gene action, are least-squares and maximum likelihood. Both methods may be used to test either single markers or the interval between flanking markers, and both suffer from the need to minimize type I and type II error rates with multiple tests.

Genetic differences between cysts of Echinococcus granulosus from the same host.

Isozyme differences were found between protoscoleces derived from different cysts in three sheep and three macropod marsupials. Isozymes were interpreted as the products of different alleles at corresponding enzyme loci, indicating that the same host may contain cysts derived from genetically different embryos. Genetic variation on this scale may cause confusion in epidemiological studies, if protoscoleces from several cysts are pooled prior to strain-typing.

Echinococcus: biology and strain variation.

Biology and strain variation in the causative agent of hydatid disease is reviewed with emphasis on developmental and genetic aspects. In vitro cultivation experiments have made a significant contribution to current knowledge of the developmental plasticity of Echinococcus. However, the mechanisms which regulate and determine developmental strategies in the parasite, as well as the characteristics, source and cytodifferentiation of germinal cells, are not understood. The nature, significance and origin of strain variation in Echinococcus are examined. Before we can fully appreciate the phenotypic consequences of genetic differentiation between populations, we need to know something about the genetic and environmental components of variation in traits such as development rate, host preference, host specificity, virulence and drug resistance. There is an urgent need for research on the developmental pathways by which genetic differences within and between strains of E. granulosus are translated to phenotypic differences in these traits.

Comparative studies on the growth dynamics of two genetically distinct isolates of Giardia duodenalis in vitro.

The in vitro growth behaviour of the intestinal protozoan Giardia duodenalis was studied in detail and comparisons were made between two genetically and biologically distinct cloned isolates. Replicates of each clone were grown at six different initial cell concentrations and in culture media at four different pH values. Significant differences in in vitro growth were found between the two isolates, BAH12 and P1. BAH12 had a specific narrow pH requirement, with satisfactory growth only obtained at pH 6. The mean generation time of BAH12 at pH 6 between days 1 and 3 was 10.8 h, compared to an average of 6 h for the same period for P1, both at pH 6 and pH 7. Comparative health of cultures was assessed during both the pH and growth experiments using a suite of six variables. Consistent changes in the health of cultures over time were found to reflect growth behaviour over time. These results provide the first detailed evidence that genetically different isolates of Giardia may differ in such fundamental biological parameters as growth rate and pH requirements. These differences may have important epidemiological and taxonomic implications.

The role of companion animals in the emergence of parasitic zoonoses.

Pets offer individuals and the community significant benefits, however cognisance must be taken of the potential for transmission of infectious agents from these animals to humans. The prevalence of many parasites, such as Giardia and Cryptosporidium, has increased over the past few decades while others, such as Toxocara and Ancylostoma, have decreased. These changes could be real, associated with the ready availability of efficacious anthelmintic products or could be artificial due to the type of surveys conducted, the animals surveyed and the diagnostic tests used. Immunocompromised people, in particular, must be aware of the potential risk of acquiring parasitic infections from their pets. However, with the adoption of good hygiene and a thorough knowledge of the transmission of these parasites, immunocompromised people should be able to continue to enjoy the significant benefits of pet ownership. As many owners are not aware of the zoonotic parasites that could be carried by their pets or their mode of transmission, it is concluded that veterinarians need to play a greater role in the education of their clients.

The origin of a new focus of infection with Echinococcus granulosus in Tasmania.

Hydatid cysts were discovered in cattle on King Island, north of Tasmania, where Echinococcus granulosus was thought to have been eradicated. Using enzyme electrophoresis, isolates from King Island were compared genetically with isolates from Tasmania and the mainland of Australia. The genetic distinctness of the King Island isolates make it unlikely that they originated from a recent introduction from either Tasmania or mainland Australia. Alternative possibilities, that the infection resulted from a recent introduction from another source or from previously undetected persistence of E. granulosus on King Island, could not be distinguished from available data.

Isoenzyme electrophoresis of 30 isolates of Giardia from humans and felines.

Thirty isolates of Giardia duodenalis from humans and felines were compared by isoenzyme electrophoresis. Using 10 enzyme systems, 13 different zymodemes were distinguished. The majority of zymodemes could be divided into two groups: one group comprising human and feline isolates with worldwide geographic distribution; the other group containing human isolates restricted to Western Australia. A number of isolates showed multiple-banded patterns and the genetic significance of these findings is discussed. The marked heterogeneity of G. duodenalis demonstrated in this study is considered in relation to the epidemiology of giardiasis. The findings are consistent with felines serving as a reservoir of infection to humans.

Characterization of Giardia isolates using a non-radiolabeled DNA probe, and correlation with the results of isoenzyme analysis.

Forty-seven isolates, identified morphologically as Giardia duodenalis, were compared by restriction endonuclease analysis of DNA with hybridization to a non-radiolabeled probe. Seven schizodemes were distinguished, compared to 15 zymodemes identified by isoenzyme electrophoresis. Despite the greater sensitivity of isoenzyme electrophoresis, DNA analysis did detect previously unsuspected variations between isolates in 1 zymodeme. Eighteen different genetic groups were detected among the 49 isolates by isoenzyme and DNA analyses. Genetic differences between groups, calculated from DNA restriction fragment variation, were significantly correlated with differences calculated from allozyme variation. This correlation between the 2 techniques suggests that G. duodenalis consists of a complex of genetically diverse clones. Such a genetic structure has important implications for the taxonomy of Giardia and the epidemiology of giardiasis.

Differences in antigen expression within and between 10 isolates of Giardia duodenalis.

In this study indirect immunofluorescence was performed on both live and fixed trophozoites to determine the level of variability in surface antigen expression between 14 Giardia duodenalis isolates, using a monoclonal antibody raised against the Portland 1 isolate (ATCC No. 30888). Subsets of antigen positive cells were detected in 13 isolates ranging in number from < 1% to 50% of the total population. The differences in antigen expression between 10 uncloned isolates did not correlate with genetic differences determined using isoenzyme analysis. Examination of four clones of the Portland 1 isolate showed that all of the progeny expressed surface antigen at significantly different levels to the parent.

The porcine intestinal spirochaetes: identification of new genetic groups.

The weakly beta-haemolytic isolates were divided into 56 electrophoretic types (ETs), contained in three distinct genetic groups (A,B and C). Group A corresponded to the genus Serpulina, and could be divided into three divisions. It contained 17 weakly haemolytic isolates in divisions b and c, as well as all 98 isolates of S. hyodysenteriae, located in division a. All seven weakly beta-haemolytic isolates that produced indole and had alpha-glucosidase but not alpha-galactosidase activity fell into division b. These spirochaetes may represent a distinct species. The other ten weakly beta-haemolytic spirochaetes, in division c, fitted the description of S. innocens. Group B contained 17 of the weakly beta-haemolytic isolates (18.9%) in ten ETs. Isolates in this group differed from typical S. innocens in that they lacked alpha-galactosidase activity. Group B represented a distinct group of weekly beta-haemolytic spirochaetes, which may constitute a new genus. Group C contained 56 of the weakly beta-haemolytic isolates (62.2%) located in 29 ETs. The original isolate from "spirochaetal diarrhoea" (P43/6/78-Taylor et al., 1980) was located in this group, together with Australian isolates from a similar condition. Spirochaetes in group C were morphologically distinct from those in groups A and B in that they possessed only four, five or occasionally six, subterminal axial filaments, were more slender, and had more pointed ends to their cells. We consider that group C represents a new genus of spirochaetes, members of which may be associated with spirochaetal diarrhoea.

Genetic relationships between isolates of Serpulina (Treponema) hyodysenteriae, and comparison of methods for their subspecific differentiation.

Multilocus enzyme electrophoresis (MEE) was used to examine the extent of genetic diversity amongst 98 isolates of Serpulina (Treponema) hyodysenteriae. The species contained four major genetic divisions (A, B, C and D) and 29 electrophoretic types (ETs). Division D was relatively distinct, being separated from the other three divisions by fixed allelic differences at an average of 6.6 of 15 enzyme loci. Electrophoretic differences were compared with results of DNA restriction endonuclease analysis (REA) and serological typing of the isolates. Most isolates with the same or similar REA banding patterns shared the same or similar ETs. This demonstrated that both techniques could be used as sensitive and specific methods of identifying closely related isolates. However, using MEE analysis, some isolates that had quite different REA patterns were found to be genetically closely related. Therefore ET designations had an advantage over REA patterns in that they were readily quantifiable as a means of estimating genetic relatedness between isolates. Most isolates that were genetically similar to each other were of the same serological group, but some antigenic types were widely distributed across the genetic divisions.

Multilocus enzyme electrophoresis for identification and typing of Treponema hyodysenteriae and related spirochaetes.

An assessment was made of multilocus enzyme electrophoresis as a means of identifying and typing spirochaetes isolated from pigs. Using five enzyme systems, 36 isolates from Australia, the U.K. and the U.S.A. were divided into 12 electrophoretic types or multilocus genotypes, comprising four major, genetically distinct groups. All 26 isolates of Treponema hyodysenteriae fell into one group, members of which showed relatively little genetic diversity. Ten isolates of non-pathogenic spirochaetes fell into three genetically different groups. Although the technique was capable of typing organisms within the groups, it was not always as discriminatory as DNA-restriction endonuclease analysis. Examination of additional enzyme loci should increase the sensitivity of the method for typing and for overall assessment of genetic relationships between spirochaetes.