SOLiD™ Sequencing of Genomes of Clinical Isolates of Leishmania donovani from India Confirm Leptomonas Co-Infection and Raise Some Key Questions

Abstract

BackgroundKnown as ‘neglected disease’ because relatively little effort has been applied to finding cures, leishmaniasis kills more than 150,000 people every year and debilitates millions more. Visceral leishmaniasis (VL), also called Kala Azar (KA) or black fever in India, claims around 20,000 lives every year. Whole genome analysis presents an excellent means to identify new targets for drugs, vaccine and diagnostics development, and also provide an avenue into the biological basis of parasite virulence in the L. donovani complex prevalent in India.Methodology/Principal FindingsIn our presently described study, the next generation SOLiD™ platform was successfully utilized for the first time to carry out whole genome sequencing of L. donovani clinical isolates from India. We report the exceptional occurrence of insect trypanosomatids in clinical cases of visceral leishmaniasis (Kala Azar) patients in India. We confirm with whole genome sequencing analysis data that isolates which were sequenced from Kala Azar (visceral leishmaniasis) cases were genetically related to Leptomonas. The co-infection in splenic aspirate of these patients with a species of Leptomonas and how likely is it that the infection might be pathogenic, are key questions which need to be investigated. We discuss our results in the context of some important probable hypothesis in this article.Conclusions/SignificanceOur intriguing results of unusual cases of Kala Azar found to be most similar to Leptomonas species put forth important clinical implications for the treatment of Kala Azar in India. Leptomonas have been shown to be highly susceptible to several standard leishmaniacides in vitro. There is very little divergence among these two species viz. Leishmania sp. and L. seymouri, in terms of genomic sequence and organization. A more extensive perception of the phenomenon of co-infection needs to be addressed from molecular pathogenesis and eco-epidemiological standpoint.

Isolation of Crithidia spp. from lesions of immunocompetent patients with suspected cutaneous leishmaniasis in Iran

“…Sing et al . reported that after whole genome sequencing, only L. symori was isolated from the spleen of some immune‐competent patients with visceral leishmaniasis.…”

“…the occurrence of Leptomonas.symori as a lower trypanosomatids in visceral leishmaniasis patients [29,30]. Sing et al [31] reported that after whole genome sequencing, only L. symori was isolated from the spleen of some immune-competent patients with visceral leishmaniasis.…”

The evolution of trypanosomatid taxonomy

“…BHU [91]. Several similar cases have been reported since, with later studies incriminating Leptomonas seymouri as the opportunistic monoxenous agent [80, 85, 90]. In such cases, Leptomonas may be detected in bone marrow and splenic aspirates, reminiscent of VL [90].…”

“…Leptomonas seymouri ) exploring the dixenous niche [80, 84–86] (discussed below). While these cases are seemingly rare exceptions rather than common events, they highlight that isolation of promastigotes from a vertebrate does not confirm a Leishmania infection which does confound classic taxonomic definitions.…”

“…It was proposed that the monoxenous flagellates were able to infect these patients due to immune-suppression associated with visceral leishmaniasis.2012Ghosh et al [85]In a study of Indian Leishmania donovani infected patients, 4/29 (13.8%) patients with visceral leishmaniasis and post-kala-azar dermal leishmaniasis (PKDL) were co-infected with the previously identified Leptomonas sp. BHU, which was later confirmed as Leptomonas seymouri .2013Singh et al [80]Through whole genome sequencing of the L. donovani clinical isolates from India, the presence of monoxenous trypanosomatids in cases of visceral leishmaniasis was reported as Leptomonas seymouri . It is important to note that a recent study by Kraeva et al found that some of the DNA sequences of Leptomonas seymouri were misidentified as Leishmania donovani in GenBank [53].…”