Ultrastructural and Molecular Characterisation of an Heterosporis-Like Microsporidian in Australian Sea Snakes (Hydrophiinae).

Four sea snakes (two Hydrophis major, one Hydrophis platurus, one Hydrophis elegans) were found washed ashore on different beaches in the Sunshine Coast region and Fraser Island in Queensland, Australia between 2007-2013. Each snake had multiple granulomas and locally extensive regions of pallor evident in the hypaxial and intercostal musculature along the body. Lesions in two individuals were also associated with vertebral and rib fractures. Histological examination revealed granulomas scattered throughout skeletal muscle, subcutaneous adipose tissue and fractured bone. These were composed of dense aggregates of microsporidian spores surrounded by a mantle of macrophages. Sequences (ssrRNA) were obtained from lesions in three sea snakes and all revealed 99% similarity with Heterosporis anguillarum from the Japanese eel (Anguillarum japonica). However, ultrastructural characteristics of the organism were not consistent with those of previous descriptions. Electron microscopic examination of skeletal muscle revealed large cysts (not xenomas) bound by walls of fibrillar material (Heterosporis-like sporophorocyst walls were not detected). The cysts contained numerous mature microsporidian spores arranged in small clusters, sometimes apparently within sporophorous vesicles. The microspores were monomorphic, oval and measured 2.5-3.0 µm by 1.6-1.8 µm. They contained isofilar polar filaments with 11 (infrequently 9-12) coils arranged in two ranks. This is the first published report of a microsporidian infection in hydrophiid sea snakes. This discovery shows microsporidia with molecular affinities to Heterosporis anguillarum but ultrastructural characters most consistent with the genus Pleistophora (but no hitherto described species). Further studies are required to determine whether the microsporidian presented here belongs to the genus Heterosporis, or to a polymorphic species group as suggested by the recognition of a robust Pleistophora/Heterosporis clade by molecular studies. The gross and histological pathology associated with these infections are described.

Phylogeny and biogeography of the "Australian" trichostomes (Ciliophora: Litostomata).

The phylogenetic relationships of members of the ciliate class Litostomatea were determined by a molecular phylogeny using the small subunit of the ribosomal RNA (ssu-rRNA) gene and a morphological phylogeny based on ultrastructural analyses of the group. Molecular analyses consistently supported the monophyly of Trichostomatia, Entodiniomorphida and the "Australian" trichostomes but provided limited support for a monophyletic Vestibuliferida and Haptoria. The results of the morphological analyses depended on the way in which the dataset was treated: "unordered" and "ordered" recovered a monophyletic Trichostomatia, Haptoria and the "Australian" trichostomes but challenged the monophyly of Entodinimorphida and Vestibuliferida; "dollo" recovered a monophyletic Trichostomatia and Entodiniomorphida but at the cost of a greatly longer tree than either "unordered" or "ordered" datasets. The monophyly of each "Australian" trichostome family was supported in all analyses and by both approaches. These results suggest that the trichostome ciliates may have become associated with mammals in Gondwana with the "Australian" trichostome ciliates entering Australia with primitive herbivorous marsupials. Subsequent diversification of the "Australian" families was probably a result of dietary specialization and oral and cortical synapomorphies define each family. We decline at this time to erect a formal taxon name for the "Australian" trichostomes due to the instability of other superfamilial taxa within the Litosomatea and concerns about the stability of tree topology until a better taxon sample of litostome ciliates is available.

Drug resistance in the sexually transmitted protozoan Trichomonas vaginalis.

Trichomoniasis is the most common, sexually transmitted infection. It is caused by the flagellated protozoan parasite Trichomonas vaginalis. Symptoms include vaginitis and infections have been associated with preterm delivery, low birth weight and increased infant mortality, as well as predisposing to HIV/AIDS and cervical cancer. Trichomoniasis has the highest prevalence and incidence of any sexually transmitted infection. The 5-nitroimidazole drugs, of which metronidazole is the most prescribed, are the only approved, effective drugs to treat trichomoniasis. Resistance against metronidazole is frequently reported and cross-resistance among the family of 5-nitroimidazole drugs is common, leaving no alternative for treatment, with some cases remaining unresolved. The mechanism of metronidazole resistance in T. vaginalis from treatment failures is not well understood, unlike resistance which is developed in the laboratory under increasing metronidazole pressure. In the latter situation, hydrogenosomal function which is involved in activation of the prodrug, metronidazole, is down-regulated. Reversion to sensitivity is incomplete after removal of drug pressure in the highly resistant parasites while clinically resistant strains, so far analysed, maintain their resistance levels in the absence of drug pressure. Although anaerobic resistance has been regarded as a laboratory induced phenomenon, it clearly has been demonstrated in clinical isolates. Pursuit of both approaches will allow dissection of the underlying mechanisms. Many alternative drugs and treatments have been tested in vivo in cases of refractory trichomoniasis, as well as in vitro with some successes including the broad spectrum anti-parasitic drug nitazoxanide. Drug resistance incidence in T. vaginalis appears to be on the increase and improved surveillance of treatment failures is urged.