Laboratory evaluation of the rapid diagnostic tests for the detection of Vibrio cholerae O1 using diarrheal samples

BackgroundCholera, an acute diarrheal disease is a major public health problem in many developing countries. Several rapid diagnostic tests (RDT) are available for the detection of cholera, but their efficacies are not compared in an endemic setting. In this study, we have compared the specificity and sensitivity of three RDT kits for the detection of Vibrio cholerae O1 and compared their efficiency with culture and polymerase chain reaction (PCR) methods.MethodsFive hundred six diarrheal stool samples collected from patients from two different hospitals in Kolkata, India were tested using SD Bioline Cholera, SMART-II Cholera O1 and Crystal-VC RDT kits. All the stool samples were screened for the presence of V. cholerae by direct and enrichment culture methods. Stool DNA-based PCR assay was made to target the cholera toxin (ctxAB) and O1 somatic antigen (rfb) encoding genes. Statistical evaluation of the RDTs has been made using STATA software with stool culture and PCR results as the gold standards. The Bayesian latent class model (LCM) was used to evaluate the diagnostic tests in the absence of the gold standard.ResultsInvolving culture technique as gold standard, the sensitivity and specificity of the cholera RDT kits in the direct testing of stools was highest with SAMRT-II (86.1%) and SD-Cholera (94.4%), respectively. The DNA based PCR assays gave very high sensitivity (98.4%) but the specificity was comparatively low (75.3%). After enrichment, the high sensitivity and specificity was detected with SAMRT-II (78.8%) and SD-Cholera (99.1%), respectively. Considering PCR as the gold standard, the sensitivity and specificity of the RDTs remained between 52.3–58.2% and 92.3–96.8%, respectively. In the LCM, the sensitivity of direct and enrichment testing was high in SAMRT-II (88% and 92%, respectively), but the specificity was high in SD cholera for both the methods (97% and 100%, respectively). The sensitivity/specificity of RDTs and direct culture have also been analyzed considering the age, gender and diarrheal disease severity of the patients.ConclusionOverall, the performance of the RDT kits remained almost similar in terms of specificity and sensitivity. Performance of PCR was superior to the antibody-based RDTs. The RTDs are very useful in identifying cholera cases during outbreak/epidemic situations and for making them as a point-of-care (POC) testing tool needs more improvement.     

The MARCH family E3 ubiquitin ligase K5 alters monocyte metabolism and proliferation through receptor tyrosine kinase modulation

Kaposi's sarcoma (KS) lesions are complex mixtures of KS-associated herpesvirus (KSHV)-infected spindle and inflammatory cells. In order to survive the host immune responses, KSHV encodes a number of immunomodulatory proteins, including the E3 ubiquitin ligase K5. In exploring the role of this viral protein in monocytes, we made the surprising discovery that in addition to a potential role in down regulation of immune responses, K5 also contributes to increased proliferation and alters cellular metabolism. This ubiquitin ligase increases aerobic glycolysis and lactate production through modulation of cellular growth factor-binding receptor tyrosine kinase endocytosis, increasing the sensitivity of cells to autocrine and paracrine factors. This leads to an altered pattern of cellular phosphorylation, increases in Akt activation and a longer duration of Erk1/2 phosphorylation. Overall, we believe this to be the first report of a virally-encoded ubiquitin ligase potentially contributing to oncogenesis through alterations in growth factor signaling cascades and opens a new avenue of research in K5 biology.     

Psychosocial Health Problems Associated with Increased HIV Risk Behavior among Men Who Have Sex with Men in Nepal: A Cross-Sectional Survey

Background

Men who have sex with men (MSM) are marginalized, hidden, underserved and at high risk for HIV in Nepal. We examined the association between MSM sub-populations, psychosocial health problems and support, access to prevention and non-use of condoms.

Methods

Between September-November of 2010, a cross-sectional survey on HIV-related risk behavior was performed across Nepal through snowball sampling facilitated by non-governmental organizations, recruiting 339 MSM, age 15 or older. The primary outcomes were: (a) non-use of condoms at least once in last three anal sex encounters with men and (b) non-use of condoms with women in the last encounter. The secondary outcome was participation in HIV prevention interventions in the past year.

Results

Among the 339 MSM interviewed, 78% did not use condoms at their last anal sex with another man, 35% did not use condoms in their last sex with a woman, 70% had experienced violence in the last 12 months, 61% were experiencing depression and 47% had thought of committing suicide. After adjustment for age, religion, marital status, and MSM subpopulations (bisexual, ta, meti, gay), non-use of condoms at last anal sex with a man was significantly associated with non-participation in HIV interventions, experience of physical and sexual violence, depression, repeated suicidal thoughts, small social support network and being dissatisfied with social support. Depression was marginally associated with non-use of condoms with women. The findings suggest that among MSM who reported non-use of condoms at last anal sex, the ta subgroup and those lacking family acceptance were the least likely to have participated in any preventive interventions.

Conclusions

MSM in Nepal have a prevalence of psychosocial health problems in turn associated with high risk behavior for HIV. Future HIV prevention efforts targeting MSM in Nepal should cover all MSM subpopulations and prioritize psychosocial health interventions.     

Ecological redundancy between coral reef sharks and predatory teleosts

Reef sharks may be ecologically redundant, such that other mesopredatory fishes compensate for their functions when they decline in number, preventing trophic cascades. Oral jaw gape, hereafter referred to as gape, determines maximum prey size in many piscivores and therefore affects the size structure of prey assemblages. Here, we examine whether gape and maximum prey size differ between five species of reef shark and 21 species of teleost (n?=?754) using data collected from 38 reefs in the Indo-Pacific. Sharks displayed relatively small gape dimensions compared to most teleost species and, at smaller sizes, the giant trevally Caranx ignobilis and other teleosts may be able to consume larger prey than similar-sized sharks. However, ecological redundancy between reef sharks and teleosts appears to decline at larger sizes, such that the grey reef shark Carcharhinus amblyrhynchos, for example, may be capable of consuming larger prey than any other reef predator at its largest sizes, regardless of prey body shape. Moreover, sharks may be able to consume proportionally larger prey as they grow, in contrast to reef teleosts, which may largely be limited by their gapes to ever-smaller prey as a proportion of their body size. Our results also suggest that reef sharks may be unable to swallow whole prey that are >?36% of their length, consistent with gut-content studies. Conservation of reef ecological function may therefore depend not only on the protection of sharks but also particular size classes and key components of the mesopredatory guild.

     

Efficiency of pretreatments for optimal enzymatic saccharification of soybean fiber

The effectiveness of several pretreatments [high-power ultrasound, sulfuric acid (H₂SO₄), sodium hydroxide (NaOH), and ammonium hydroxide (NH₃OH)] to enhance glucose production from insoluble fractions recovered from enzyme-assisted aqueous extraction processing of extruded full-fat soybean flakes (FFSF) was investigated. Sonication of the insoluble fraction at 144 μm_{pp (peak-to-peak)} for 30 and 60 s did not improve the saccharification yield. The solid fractions recovered after pretreatment with H₂SO₄ [1% (w/w), 90 °C, 1.5 h], NaOH [15% (w/w), 65 °C, 17 h], and NH₃OH [15% (w/w), 65 °C, 17 h] showed significant lignin degradation, i.e., 81.9%, 71.2%, and 75.4%, respectively, when compared to the control (7.4%). NH₃OH pretreatment resulted in the highest saccharification yield (63%) after 48 h of enzymatic saccharification. A treatment combining the extraction and saccharification steps and applied directly to the extruded FFSF, where oil extraction yield and saccharification yield reached 98% and 43%, respectively, was identified.     

Characterization of 2-octenoyl-CoA carboxylase/reductase utilizing pteB from Streptomyce avermitilis

The filipin biosynthetic gene cluster of Streptomyces avermitilis contains pteB, a homolog of crotonyl-CoA carboxylase/reductase. PteB was predicted to be 2-octenoyl-CoA carboxylase/reductase, supplying hexylmalonyl-CoA to filipin biosynthesis. Recombinant PteB displayed selective reductase activity toward 2-octenoyl-CoA while generating a broad range of alkylmalonyl-CoAs in the presence of bicarbonate.     

Caspase-2 cleaves DNA fragmentation factor (DFF45)/inhibitor of caspase-activated DNase (ICAD)

To investigate the signal transduction pathway of caspase-2, cell permeable Tat-reverse-caspase-2 was constructed, characterized and utilized for biochemical and cellular studies. It could induce the cell death as early as 2 h, and caspase-2-specific VDVADase activity but not other caspase activities including DEVDase and IETDase. Interestingly, nuclear DNA fragmentation occurred and consistently DNA fragmentation factor (DFF45)/Inhibitor of caspase-activated DNase (ICAD) was cleaved inside the cell as well as in vitro, suggesting a role of caspase-2 in nuclear DNA fragmentation.