Genetic Traits of Vibrio cholerae O1 Haitian Isolates That Are Absent in Contemporary Strains from Kolkata,India

The world''s worst cholera epidemic in Haiti (2010) coerced to trace the origin and dissemination of the causative agent Vibrio cholerae O1 for proper management of cholera. Sequence analysis of the Haitian strain showed several variations in the genes encoding cholera toxin B subunit (ctxB); toxin-co-regulated pilus (tcpA), repeat in toxins (rtxA), quinolone resistance-determining region (QRDR) of gyrase A (gyrA), rstB of RS element along with the change in the number of repeat sequences at the promoter region of ctxAB. Our earlier studies showed that variant tcpA (tcpA CIRS) and ctxB (ctxB7) first appeared in Kolkata during 2003 and 2006, respectively. The present study revealed that a variant rtxA was first isolated in Kolkata during 2004 and probably formed the genetic background for the emergence of the ctxB7 allele as we were unable to detect a single strain with the combination of El Tor rtxA and ctxB7. The variant gyrA was first time detected in Kolkata during 1994. The Kolkata strains contained four heptad repeats (TTTTGAT) in their CT promoter regions whereas Haitian strains carried 5 heptad repeats. Haitian strains had 3 nucleotide deletions at the rstB gene, which is a unique feature of the classical biotype strains. But the Kolkata strains did not have such deletion mutations in the rstB. Our study demonstrated the existence of some Haitian genetic traits in Kolkata isolates along with the dissimilarities in genomic content with respect to rstB and ctxAB promoter region. Finally, we conclude that Haitian variant strain may be evolved due to sequential event in the Indian subcontinent strain with some cryptic modification in the genome.     

The AMA1-RON complex drives Plasmodium sporozoite invasion in the mosquito and mammalian hosts

Plasmodium sporozoites that are transmitted by blood-feeding female Anopheles mosquitoes invade hepatocytes for an initial round of intracellular replication, leading to the release of merozoites that invade and multiply within red blood cells. Sporozoites and merozoites share a number of proteins that are expressed by both stages, including the Apical Membrane Antigen 1 (AMA1) and the Rhoptry Neck Proteins (RONs). Although AMA1 and RONs are essential for merozoite invasion of erythrocytes during asexual blood stage replication of the parasite, their function in sporozoites was still unclear. Here we show that AMA1 interacts with RONs in mature sporozoites. By using DiCre-mediated conditional gene deletion in P. berghei, we demonstrate that loss of AMA1, RON2 or RON4 in sporozoites impairs colonization of the mosquito salivary glands and invasion of mammalian hepatocytes, without affecting transcellular parasite migration. Three-dimensional electron microscopy data showed that sporozoites enter salivary gland cells through a ring-like structure and by forming a transient vacuole. The absence of a functional AMA1-RON complex led to an altered morphology of the entry junction, associated with epithelial cell damage. Our data establish that AMA1 and RONs facilitate host cell invasion across Plasmodium invasive stages, and suggest that sporozoites use the AMA1-RON complex to efficiently and safely enter the mosquito salivary glands to ensure successful parasite transmission. These results open up the possibility of targeting the AMA1-RON complex for transmission-blocking antimalarial strategies.     

Plasmodium berghei leucine-rich repeat protein 1 downregulates protein phosphatase 1 activity and is required for efficient oocyst development

Protein phosphatase 1 (PP1) is a key enzyme for Plasmodium development. However, the detailed mechanisms underlying its regulation remain to be deciphered. Here, we report the functional characterization of the Plasmodium berghei leucine-rich repeat protein 1 (PbLRR1), an orthologue of SDS22, one of the most ancient and conserved PP1 interactors. Our study shows that PbLRR1 is expressed during intra-erythrocytic development of the parasite, and up to the zygote stage in mosquitoes. PbLRR1 can be found in complex with PbPP1 in both asexual and sexual stages and inhibits its phosphatase activity. Genetic analysis demonstrates that PbLRR1 depletion adversely affects the development of oocysts. PbLRR1 interactome analysis associated with phospho-proteomics studies identifies several novel putative PbLRR1/PbPP1 partners. Some of these partners have previously been characterized as essential for the parasite sexual development. Interestingly, and for the first time, Inhibitor 3 (I3), a well-known and direct interactant of Plasmodium PP1, was found to be drastically hypophosphorylated in PbLRR1-depleted parasites. These data, along with the detection of I3 with PP1 in the LRR1 interactome, strongly suggest that the phosphorylation status of PbI3 is under the control of the PP1–LRR1 complex and could contribute (in)directly to oocyst development. This study provides new insights into previously unrecognized PbPP1 fine regulation of Plasmodium oocyst development through its interaction with PbLRR1.     

Radio-attenuated leishmanial parasites as immunoprophylactic agent against experimental murine visceral leishmaniasis

The present study intends to evaluate the role of radio-attenuated leishmania parasites as immunoprophylactic agents for experimental murine visceral leishmaniasis. BALB/c mice were immunized with gamma (γ)-irradiated Leishmania donovani. A second immunization was given after 15 days of first immunization. After two immunizations, mice were infected with virulent L. donovani promastigotes. Protection against Kala-azar (KA) was estimated from spleen and liver parasitic burden along with the measurement of nitrite and superoxide anion generation by isolation of splenocytes and also by T-lymphocyte helper 1(Th1) and T-lymphocyte helper 2(Th2) cytokines release from the experimental groups. It was observed that BALB/c mice having prior immunization with radio-attenuated parasites showed protection against L. donovani infection through higher expression of Th1 cytokines and suppression of Th2 cytokines along with the generation of protective free radicals. The group of mice without prior priming with radio-attenuated parasites surrendered to the disease. Thus it can be concluded that radio-attenuated L. donovani may be used for.     

Sialic acid attenuates puromycin aminonucleoside-induced desialylation and oxidative stress in human podocytes

Sialoglycoproteins make a significant contribution to the negative charge of the glomerular anionic glycocalyx—crucial for efficient functioning of the glomerular permselective barrier. Defects in sialylation have serious consequences on podocyte function leading to the development of proteinuria. The aim of the current study was to investigate potential mechanisms underlying puromycin aminonucleosisde (PAN)-induced desialylation and to ascertain whether they could be corrected by administration of free sialic acid.     

Epidemio-Clinico-Microbiological Study of Mycotic Keratitis in North-West Region of Rajasthan

Aim and Objectives

Mycotic keratitis, with its diverse clinical presentation and difficulties in treatment, makes it a challenging task for clinicians and an important object of study. The aim of present study was to determine the frequency of occurrence and epidemiological association with identification of fungal isolates from mycotic keratitis cases.

Materials and Methods

This was a prospective and observational study conducted in Mycology Laboratory, Department of Microbiology, S.P. Medical College, Bikaner, on corneal scrapings and swabs of a total of 480 patients attending the Ophthalmology OPD, P.B.M. Hospital, Bikaner, during July 2005 to June 2012.

Results

Out of 480 suspected cases, 180 were found to be positive for fungus by smear/culture examination. Increased incidence was seen in the months of May to September with Aspergillus fumigatus being the most common isolate.

Conclusion

Mycotic keratitis, though an age-old disease, still presented with challenging aspects of diagnosis and treatment. The study showed fungal keratitis is prevalent in rural parts of north-west Rajasthan, mainly found in males (age group 21–40 years) with low socio-economic status (farm or factory workers). The most common cause of fungal keratitis was found to be species of Aspergillus.