Preeti Mehta

Transmitted HIV Drug Resistance Among HIV-Infected Voluntary Counseling and Testing Centers (VCTC) Clients in Mumbai, India

A survey for transmitted HIV drug resistance (HIVDR) was conducted according to WHO guidelines among clients newly diagnosed with HIV-1 infection at two voluntary counseling and testing centers (VCTC) in Mumbai. HIVDR testing was performed using the ViroSeq RT-PCR method (Abbott). Out of 50 successfully amplified and sequenced specimens, analysis of the first 34 consecutively collected specimens revealed no nucleoside reverse transcriptase inhibitor, nonnucleoside reverse transcriptase inhibitor, or protease inhibitor mutations from the 2007 WHO list of mutations for surveillance of transmitted HIVDR, indicating that the prevalence of transmitted HIVDR to all three drug classes was <5% among recently infected VCTC clients in Mumbai. The phylogenetic analysis revealed that all samples belonged to HIV-1 subtype C. Continued ART program monitoring and further evaluation of transmitted HIV drug resistance in coming years are essential in Mumbai as well as in other regions of the country in which ART is being scaled up rapidly.

Prevalence of multidrug-resistant enterococci in a tertiary care hospital in Mumbai, India.

Introduction Enterococci are one of the major causes of nosocomial and community-acquired infections. In recent years, the evolution of antimicrobial resistance in enterococci has posed enormous challenges for clinicians. The antimicrobial therapy of enterococcal infections is complicated because of the inherent resistance shown by enterococci to several commonly used antibiotics such as cephalosporins, low-level aminoglycosides, and low-level clindamycin and perhaps more importantly, because of their acquired resistance to all currently available antibiotics, that leaves limited medicative options and results in the selection and spreading of multidrug-resistant (MDR) strains in hospitals [1,2]. Empirical use of antibiotics, absence of national guidelines for screening patients for MDR bacteria and lack of sufficient information and programs to control rapid spread of enterococci has led to increased mortality caused by enterococcal infections [3-5]. Knowledge of the antibiogram is essential to formulate therapeutic strategies for treating enterococcal infections [1]. This prospective study aimed to investigate species prevalence and extent of antimicrobial resistance among clinical isolates of enterococci in a tertiary care hospital in India.

Early dengue diagnosis: Role of rapid NS1 antigen, NS1 early ELISA, and PCR assay

Context: Early and accurate dengue diagnosis is important for disease surveillance, and also to start effective control measures in endemic countries. Aims: In this study, we evaluated the performance of three techniques available for early dengue diagnosis, viz., rapid nonstructural 1 (NS1) antigen test, NS1 early enzyme-linked immunosorbent assay (ELISA) test, and reverse transcription polymerase chain reaction (RT-PCR) test. Settings and Design: Retrospective analytical study. Materials and Methods: Performance of three different techniques, viz., rapid NS1 antigen, NS1 early ELISA, and RT-PCR on serum samples of suspected dengue cases. Statistical Analysis Used: Sensitivity, specificity, negative predictive value, and positive predictive value calculated for rapid test and ELISA test, keeping PCR as the gold standard Results: Amongst 155 samples received from clinically-suspected dengue cases within 1-9 days of fever, 56 samples were received during 1-3 days, 52 samples were received during 4-6 days, and 47 samples were received during 7-9 days. Of these 155 samples, 38 (24.5%) were positive by rapid NS1 antigen, 47 (30.3%) by NS1 ELISA, and 54 (34.8%) by reverse transcriptase polymerase chain reaction (RT-PCR). Rapid NS1 antigen and NS1 ELISA showed the highest positivity on days 1-3, while highest positivity for PCR was on days 4-6. The sensitivity and specificity of NS1 ELISA were 66.6% and 89.1%, while sensitivity and specificity of rapid NS1 antigen were 55.5% and 92%, respectively. Conclusions: With high mortality and morbidity associated with dengue, it is imperative to diagnose the disease during the early phase. All three assay formats are helpful in early diagnosis and to provide information of dengue cases in time.

LED fluorescence microscopy: Novel method for malaria diagnosis compared with routine methods

Rapid and accurate diagnosis of malaria is the need of hour for effective management and controlling drug resistance. The conventional and gold-standard method, Light microscopy (LM), is time-consuming, requires trained staff and well-maintained equipments. The newly developed, rapid diagnostic tests (RDT) are fast and reliable, but give only qualitative results, are expensive and have short shelf life. Light Emission Diode fluorescence microscopy (LED FM) may provide a reliable alternative which can be used for routine diagnosis. In order to assess the effectiveness of LED fluorescence microscopy in malaria diagnosis, a cross-sectional study was conducted at a tertiary care teaching hospital in Mumbai. 2-3ml of blood of 300 patients, who were clinically suspected of having malaria but were not on anti-malarial treatment, was collected in EDTA vials. These specimens were processed to diagnose malaria by three methods, namely-Peripheral smear examination with LM, Peripheral smear examination with LED FM and RDT. The results of all the 3 tests were compared, taking Light Microscopy as the gold standard method. Of the 300 specimens, LM, LED FM and RDT reported 111 (37%), 86 (28.67%) and 107 (35.67%), respectively, as positive. The sensitivity and specificity were respectively 71.2% and 96.3% for LED FM and 91% and 96.8% for RDT. Of the LM positive cases, 53 (47.75%) had parasitic index (PI) <1% and 58 (52.25%) had PI ≥1%. LED FM was found to be only moderately sensitive but highly specific in comparison to Light microscopy. In order to improve the performance of this technique, more precise training in fluorescence staining and reading of the slides, will be required.

Diversity in KIR gene repertoire in HIV-1 exposed infected and uninfected infants: A study from India.

Natural killer (NK) cells have antiviral activity mediated through killer immunoglobulin receptors (KIRs). Studies have shown the importance of KIR receptors in HIV infection. However reports on association of KIR genes in HIV infection from Indian population are limited, not a single study is reported in HIV exposed uninfected (EU) and infected infants. This study compared the KIR gene repertoire of HIV‐1 positive (n = 29) with EU (n = 76) infants to elucidate its association with transmission. KIR genotyping was analysed using the PCR‐SSP method. Viral load of mothers, CD4 count of both mothers and infected infants were done using commercial kits. The data was analysed using SPSS software. Results revealed presence of significantly high frequencies of activating gene KIR 2DS5 (P = 0.040) and inhibitory gene KIR 2DL3 (P = 0.013) in EU infants as compared to HIV‐1 positive infants, confirmed with multivariable linear regression modelling. Fifty‐nine KIR genotypes were identified in these 105 infants. Nine genotypes were unique, reported for the first time. Twenty six genotypes were shared with the World populations. Twenty four genotypes were reported for the first time from India. Specific KIR genotype combinations (GIDs) were exclusively present either in HIV‐1 positive (n = 19) or in EU infants (n = 30). The Linkage disequilibrium (LD) analysis shows a strong linkage between four pairs of genes in HIV‐1 positive and three pairs of genes in EU infants. In conclusion, this study revealed that, besides maternal confounding factors such as ART and viral load, specific KIR genes are associated independently with perinatal HIV infection. J. Med. Virol. 88:417–425, 2016.

Diagnosis of pediatric pulmonary tuberculosis with special reference to polymerase chain reaction based nucleic acid amplification test

OBJECTIVE To determine the utility of polymerase chain reaction (PCR) for diagnosing pediatric pulmonary tuberculosis (PPTB). METHOD A prospective cross-sectional study was carried out on 100 children less than 14 years of age, with strong clinical suspicion and radiological evidence suggestive of pulmonary tuberculosis (TB). Sputum samples/gastric lavage were collected. Direct smears and culture on Lowenstein Jensen (LJ) media were performed. DNA extraction and amplification was performed using Genei™ Amplification Reagent set for Mycobacterium tuberculosis (MTB) (by Genei, Bangalore, India). This test is based on the principle of single-tube nested PCR which amplifies the repetitive insertion sequence IS6110. RESULTS When compared with culture, sensitivity and specificity of PCR was 93.55% and 92.75%, respectively. The PPV was 85.29% and the NPV was 96.97%. When intention to treat (ITT) was used as the standard, sensitivity, specificity, PPV and NPV of PCR was 47.88%, 93.1%, 94.4%, and 42.19%, respectively, and that of culture was 40.85%, 100%, 100% and 40.85%, respectively. Against response to treatment (RTT), PCR demonstrated sensitivity, specificity, PPV and NPV of 50.9%, 93.1%, 93.33% and 50%, respectively, and for culture it was 43.64%, 100%, 100% and 48.33%, respectively. CONCLUSION/RECOMMENDATION The present study reinforces better case detection rate with PCR-based nucleic acid amplification test as compared with microscopy and culture in pediatric pulmonary TB. PCR showed a higher correlation with clinical diagnosis as compared with microscopy and solid culture. Hence, a molecular platform should be the test of choice for detecting PPTB.

Xpert(®) MTB/RIF for improved case detection of extra-pulmonary TB in a tertiary care setting in urban India.

SETTING Department of microbiology at a tertiary care hospital, Mumbai, India. OBJECTIVE To determine 1) the sensitivity and specificity of the Xpert(®) MTB/RIF assay in comparison with microscopy and culture in extra-pulmonary tuberculosis (EPTB), and 2) the number of additional cases of EPTB and rifampicin (RMP) resistance detected using this assay. DESIGN The study was conducted from July 2013 to April 2015. All consecutive patients with clinically suspected EPTB referred for microscopic examination to the Department of Microbiology that were sufficient in specimen volume were included in the study. RESULTS Of the 728 specimens included in the study, respectively 5.5%, 23.5% and 20.9% were positive on smear, culture and Xpert. Compared to culture, Xpert had a sensitivity of 84.2% (95%CI 81.4-86.6) and specificity of 98.2% (95%CI 90-104). All specimens with high and medium load on assay were positive on culture; 28 (18.4%) specimens were RMP-resistant and 124 (81.6%) were Xpert-susceptible. No additional RMP-resistant cases were detected using Xpert as compared to phenotypic drug susceptibility testing. CONCLUSION The ability of the Xpert assay to rapidly detect a significantly greater number of bacteriologically confirmed EPTB cases, including RMP-resistant cases, makes it an important diagnostic tool in a TB-endemic country.

Seropositivity for Antibodies to DRS-G, a Virulence Factor from Streptococcus dysgalactiae subsp. equisimilis, Is an Independent Risk Factor for Poststreptococcus Glomerulonephritis and Chronic Kidney Disease in Mumbai, India

ABSTRACT The disease spectrum caused by Streptococcus dysgalactiae subsp. equisimilis resembles that of S. pyogenes (group A streptococcus [GAS]). These two bacterial species are closely related and possess many common virulence characteristics. While some GAS strains express virulence factors called streptococcal inhibitor of complement (SIC) and distantly related to SIC (DRS), some S. dysgalactiae subsp. equisimilis isolates express an orthologue of DRS, which is referred to as DRS-G. We reported previously that seropositivity for either anti-SIC or anti-DRS antibodies (Abs) is associated with poststreptococcal glomerulonephritis (PSGN). However, only seropositivity for anti-SIC Abs is associated with chronic kidney disease (CKD). We now extend the study to test whether seropositivity for anti-DRS-G Abs is also associated with these renal diseases. Stored serum samples collected for our previous study were tested by an enzyme-linked immunosorbent assay (ELISA) for Abs to DRS-G. The samples represented sera from 100 CKD adult patients, 70 adult end-stage renal disease (ESRD) patients, 25 PSGN pediatric patients, and corresponding age-matched control subjects. The proportion of PSGN, CKD, and ESRD patients who showed seroreaction to anti-DRS-G Abs was significantly higher than that of the corresponding age-matched controls, who in general exhibited seropositivity rates commensurate with the isolation rate of drsG-positive S. dysgalactiae subsp. equisimilis in the community during this study period. Since higher rates of seropositivity for anti-DRS-G Abs in the renal disease categories are resultant of previous infections with DRS-G-positive S. dysgalactiae subsp. equisimilis strains, we conclude the seropositivity is an additional risk factor for these renal diseases. In this regard, anti-DRS-G Abs have attributes similar to those of the anti-SIC Abs.

Hepatitis B Virus (HBV) Infection in Liver Disease Patients in Mumbai, India with Special Reference to Hepatitis B Surface Antigen (HBsAg) Mutant Detection.

OBJECTIVES To determine the prevalence of Hepatitis B Surface antigen (HBsAg) in patients attending the Hepatology Out Patient Department (OPD) of a tertiary care hospital and to compare the routinely used HBsAg detection kit with the mutant detection kit to find out the presence of mutants in a given setting. MATERIALS AND METHODS A cross-sectional study was carried out in adult patients with liver disease attending the Hepatology OPD, of a tertiary care hospital in Mumbai, India. Age, gender and clinical history of the patient were recorded. Blood specimen was tested for HBsAg (Microscreen(TM) ELISA, Span diagnostics, India) and HBsAg mutants (Hepanostika HBsAg Ultra(TM) ELISA, Biomerieux, France). The samples with discordant results between these two ELISAs were confirmed by Hepatitis B Virus (HBV) Deoxyribonucleic Acid (DNA) Polymerase Chain Reaction (PCR) (Cobas Taqman(TM), Roche Molecular Systems, USA). RESULTS Seven hundred and eighteen patients were enrolled in the study. The mean age of patients in the study group was 41 years (range 17 to 69 years). Four hundred and ninety seven (69.22%) were males and remaining were females. The prevalence of HBsAg was found to be 17.4%. The positivity amongst the male population was 18.1% which was higher than the female population (15.8%). Of the 718 samples tested, 120 were positive for HBsAg by Microscreen(TM) ELISA and 132 were positive by Hepanostika HBsAg ultra(TM). Of the 12 discordant samples, HBV DNA was detected in five samples indicating 0.7% prevalence of mutants. CONCLUSION Hepatitis B is prevalent in liver disease patients. The mutant detecting assay is recommended in set-ups where missing HBsAg in patients would have tremendous impact on the outcome such as in blood donors, organ or tissue donors and antenatal screening of mothers. It is also helpful in chronic liver disease patients where the routine HBsAg detection test is negative and the other causes of chronic liver disease have been ruled out. However, it is not recommended for use in routine diagnostic set-ups where high false positivity would lead to over-diagnosis of the condition.

Seroprevalence of Streptococcal Inhibitor of Complement (SIC) suggests association of streptococcal infection with chronic kidney disease

BackgroundGroup A streptococcus (GAS) is an etiological agent for the immune mediated sequela post streptococcal glomerulonephritis (PSGN). In some populations PSGN is recognized as a risk factor for chronic kidney disease (CKD) and end-stage renal disease (ESRD). It was found that a significantly greater proportion of subjects with past history of PSGN than without the history exhibited seroreactions to streptococcal antigens called streptococcal inhibitor of complement (SIC) and to distantly related SIC (DRS). These antigens are expressed by major PSGN-associated GAS types. We therefore predicted that in populations such as India, which is endemic for streptococcal diseases and which has high prevalence of CKD and ESRD, greater proportions of CKD and ESRD patients exhibit seroreaction to SIC and DRS than healthy controls.MethodsTo test this we conducted a SIC and DRS seroprevalence study in subjects from Mumbai area. We recruited 100 CKD, 70 ESRD and 70 healthy individuals.ResultsNineteen and 35.7% of CKD and ESRD subjects respectively were SIC antibody-positive, whereas only 7% of healthy cohort was seropositive to SIC. Furthermore, significantly greater proportion of the ESRD patients than the CKD patients is seropositive to SIC (p=0.02; odds ratio 2.37). No association was found between the renal diseases and DRS-antibody-positivity.ConclusionsPast infection with SIC-positive GAS is a risk factor for CKD and ESRD in Mumbai population. Furthermore, SIC seropositivity is predictive of poor prognosis of CKD patients.