![]() Cytokine array to check the profile of different cytokines ![]() During screening, already known virus-positive and negative samples were used as respective controls for RNA extraction and RT-PCR. Serum samples from LD-negative and otherwise healthy individuals (n = 11) belonging to the endemic regions were found negative within the limits of threshold of the PCR used (data not shown). All LD-LS positive samples were also virus-positive. LS-specific PCR was also carried out and LS-DNA was detectable in only five of the twenty-eight serum samples from the aforesaid LD diagnosed patients (5/28 or 18%). The above-mentioned LD diagnosed samples (by LD-specific diagnostic tests) were reconfirmed of LD positivity by LD-specific PCR 8. The serum levels of IL-18 for virus-positive and negative LD infected patients’ serum samples as well as endemic controls were assessed. IL-18 has been previously associated with LD infection and considered as important cytokine to elicit anti-LD adaptive immunity 13, 14. We have also analyzed the cytokine profile of serum samples of VL patients. In the current study, we have screened for Lepsey NLV1 in archived and recently-collected human serum samples, from LD-diagnosed patients as well as endemic controls, rather than the promastigote stage of the parasites cultured in artificial growth media. However, it is to be noted that this coinfection has also been demonstrated in lesional skin biopsy specimens from PKDL cases and in direct biopsy materials, namely peripheral blood from VL patients, at least on one occasion 10. LD-LS coinfection of patients had been inferred from the analysis of cultured isolates in majority of the studies that have been published, including ours 8, 9, 11, 12. Ours was the first report that Indian VL/kala-azar victims are exposed to the LD-LS-Lepsey NLV1 triple pathogen complex 8. In our previous study, 20 out of the 22 (91%) LD isolates were LS-positive by PCR-based detection and 15 (75%) of the 20 LD-LS co-infected samples were Lepsey NLV1-positive. Historically, LD clinical isolates from India often showed high incidence of LS co-infection 9, 10, 11, 12. The latter (LS) is another protozoon transmitted by the sand fly vector of LD. Lepsey NLV1 was not of LD but of Leptomonas seymouri (LS) origin. Surprisingly, a different protozoan virus called Leptomonas seymouri narna-like virus 1 (Lepsey NLV1) was found in a great majority of these multi-passaged samples 8. However, LRVs have never been reported from VL samples and despite rigorous attempts we too could not detect LRVs in the 22 well-characterized Indian VL cultured isolates 8. LRV2 reported from old world leishmaniasis cases, showed a similar trend 5, 6, 7. In South American countries, an aggravated form of cutaneous leishmaniasis (CL) has been attributed to be influenced by a protozoan dsRNA virus called Leishmania RNA virus1 (LRV1) 3, 4. Some patients develop post-kala-azar dermal leishmaniasis (PKDL) whose pathogenesis is still considered an unresolved mystery 1 and it is indicated that PKDL patients might act as silent parasite pool for transmission of leishmaniasis in kala-azar endemic areas 2. India is endemic for VL and sand flies ( Phlebotomus spp) act as the insect vector. VL is commonly known as kala-azar in India. Visceral leishmaniasis (VL) is one of the most important protozoan diseases caused by Leishmania donovani (LD). The study emphasizes the need to revisit LD pathogenesis in the light of the association and persistence of a protozoan virus in kala-azar and PKDL patients. The Lepsey NLV1 interaction with the immune system results in reduced IL-18 which favors LD survival and increased parasitic burden. IL-18 is crucial for Th1 and macrophage activation which eventually clears the parasite. Cytokine profiling showed significantly elevated IL-18 in only LD infected patients compared to the virus-positive LD and control samples. The presence of the virus was confirmed in thirteen of eighteen (72%) recently collected VL and PKDL samples. RNA extracted from same samples was subjected to RT-PCR, qRT-PCR and sequencing using virus-specific primers to detect/identify and quantify the virus. DNA from the serum of twenty-eight LD diagnosed patients was subjected to LD-specific and LS-specific PCR to reconfirm the presence of LD parasites and to detect LD-LS co-infections. A pertinent question was whether this virus of LS is detectable in direct clinical samples. Leptomonas seymouri narna-like virus 1 (Lepsey NLV1) has been reported in multi-passaged laboratory isolates of VL samples which showed LD-LS co-infection. Kala-azar/Visceral Leishmaniasis (VL) caused by Leishmania donovani (LD) is often associated with Leptomonas seymouri (LS) co-infection in India.
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |