Examination of Sarcocystis spp. of giant snakes from Australia and Southeast Asia confirms presence of a known pathogen - Sarcocystis nesbitti

Abstract

We examined Sarcocystis spp. in giant snakes from the Indo-Australian Archipelago and Australia using a combination of morphological (size of sporocyst) and molecular analyses. We amplified by PCR nuclear 18S rDNA from single sporocysts in order to detect mixed infections and unequivocally assign the retrieved sequences to the corresponding parasite stage. Sarcocystis infection was generally high across the study area, with 78 (68%) of 115 examined pythons being infected by one or more Sarcocystis spp. Among 18 randomly chosen, sporocyst-positive samples (11 from Southeast Asia, 7 from Northern Australia) the only Sarcocystis species detected in Southeast Asian snakes was S. singaporensis (in reticulated pythons), which was absent from all Australian samples. We distinguished three different Sarcocystis spp. in the Australian sample set; two were excreted by scrub pythons and one by the spotted python. The sequence of the latter is an undescribed species phylogenetically related to S. lacertae. Of the two Sarcocystis species found in scrub pythons, one showed an 18S rRNA gene sequence similar to S. zamani, which is described from Australia for the first time. The second sequence was identical/similar to that of S. nesbitti, a known human pathogen that was held responsible for outbreaks of disease among tourists in Malaysia. The potential presence of S. nesbitti in Australia challenges the current hypothesis of a snake-primate life cycle, and would have implications for human health in the region. Further molecular and biological characterizations are required to confirm species identity and determine whether or not the Australian isolate has the same zoonotic potential as its Malaysian counterpart. Finally, the absence of S. nesbitti in samples from reticulated pythons (which were reported to be definitive hosts), coupled with our phylogenetic analyses, suggest that alternative snake hosts may be responsible for transmitting this parasite in Malaysia.

Fig 1. Phylogram of selected taxa of the Sarcocystidae including the new Sarcocystis isolates and species examined in this study (black symbols).

Selected taxa of the Eimeriidae and the fish-host species Goussia janae served as root. Branches A and B indicate the subfamilies Toxoplasmatinae and Sarcocystinae, respectively. In addition to snake-host taxa, branch D includes Sarcocystis spp. of ruminating mammals as intermediate hosts. The tree was reconstructed by Minimum Evolution (ME) algorithm with 1557 sites of the 18S rRNA gene under analysis. Bootstrap values ≥ 50% of 1000 iterations are indicated. See further explanations in the text. Key to generic name abbreviations as follows: B = Besnoitia, C = Cystoisospora, E = Eimeria, G = Goussia, H = Hyaloklossia, N = Neospora, S = Sarcocystis, T = Toxoplasma.

Fig 2

a) Sporocysts of Sarcocystis cf. nesbitti (arrow) and Sarcocystis sp.1 (arrowhead, tentatively S. zamani) from the Australian scrub python. Asterisk indicates residual body, sp = sporozoite; b) Oocyst of S. cf. nesbitti. Arrowhead indicates the thin oocyst wall.