Utility of multiplex real-time PCR in the diagnosis of extrapulmonary tuberculosis

AbstractObjectiveThe diagnosis of extrapulmonary tuberculosis is still a challenge because of its pauci-bacillary nature. The aim of the study was to evaluate the role of a multiplex PCR assay in the diagnosis of extrapulmonary tuberculosis and to compare the efficiency of two targets, IS6110 and MPB64 to detect Mycobacterium tuberculosis.Methods150 extrapulmonary samples (61 pus/aspirate, 46 tissue, 32 body fluids, and 11 urine) from clinically suspected cases of tuberculosis were included in the study. All the samples were subjected to direct fluorescent microscopy, TB culture (BacT/ALERT 3D, biomerieux, Durham, North Carolina, USA) and a Multiplexed Tandem PCR targeting two mycobacterial DNA sequences, IS6110 and MPB64. Master-Mix reagents and primers were prepared by AusDiagnostics Pvt. Ltd (Alexandria, New South Wales, Australia). The performance of the assay was assessed using a composite gold standard, which included clinical characteristics, microbiology smear as well as culture, histopathology, cytology, radiology, and response to antitubercular therapy.Results20.3%, 23.6%, and 45.3% of specimens were positive by smear, culture, and PCR, respectively. The sensitivity and specificity of the multiplex PCR was 91.9% and 88.4%, respectively, using the composite gold standard. Positive and negative predictive values of the PCR were estimated as 85.1% and 93.8%, respectively. Higher positivity was observed with target IS6110 (44.6%) as compared to target MPB64 (18.9%). The sensitivities of IS6110 and MPB64 individual targets were 90.3% and 64.5%, respectively, and specificities were 88.4% and 97.7%, respectively.ConclusionPCR can play an important role in rapid and accurate diagnosis of extrapulmonary tuberculosis. IS6110 alone is an effective target in our part of the country

SARS-CoV-2 B.1.617.2 Delta variant replication and immune evasion

Abstract: The B.1.617.2 (Delta) variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in the state of Maharashtra in late 2020 and spread throughout India, outcompeting pre-existing lineages including B.1.617.1 (Kappa) and B.1.1.7 (Alpha)1. In vitro, B.1.617.2 is sixfold less sensitive to serum neutralizing antibodies from recovered individuals, and eightfold less sensitive to vaccine-elicited antibodies, compared with wild-type Wuhan-1 bearing D614G. Serum neutralizing titres against B.1.617.2 were lower in ChAdOx1 vaccinees than in BNT162b2 vaccinees. B.1.617.2 spike pseudotyped viruses exhibited compromised sensitivity to monoclonal antibodies to the receptor-binding domain and the amino-terminal domain. B.1.617.2 demonstrated higher replication efficiency than B.1.1.7 in both airway organoid and human airway epithelial systems, associated with B.1.617.2 spike being in a predominantly cleaved state compared with B.1.1.7 spike. The B.1.617.2 spike protein was able to mediate highly efficient syncytium formation that was less sensitive to inhibition by neutralizing antibody, compared with that of wild-type spike. We also observed that B.1.617.2 had higher replication and spike-mediated entry than B.1.617.1, potentially explaining the B.1.617.2 dominance. In an analysis of more than 130 SARS-CoV-2-infected health care workers across three centres in India during a period of mixed lineage circulation, we observed reduced ChAdOx1 vaccine effectiveness against B.1.617.2 relative to non-B.1.617.2, with the caveat of possible residual confounding. Compromised vaccine efficacy against the highly fit and immune-evasive B.1.617.2 Delta variant warrants continued infection control measures in the post-vaccination era

Multidrug-resistant pulmonary & extrapulmonary tuberculosis: A 13 years retrospective hospital-based analysis

Background & objectives: Multidrug-resistant tuberculosis (MDR-TB) is a public health problem of great significance in India. In the present study an attempt was made to analyse the progression of MDR-TB pattern during a course of 13 years (2000-2012) among the patient population at a tertiary care centre in New Delhi, India. Methods: Mycobacterial isolates obtained on Lowenstein-Jensen (L-J) medium/BacT/ALERT 3D were identified using AccuProbe molecular identification system, routine biochemical tests or GenoType Mycobacteria CM. Antimycobacterial susceptibility testing was performed using resistance ratio method on L-J medium (2000-2004) and one per cent proportion method on BacT/ALERT 3D system (2005-2012). Results:Of the total 14,849 samples subjected to mycobacterial culture, 6569 pulmonary and 8280 extrapulmonary, 2364 were detected positive for mycobacteria. The average percentage positivity rate was 15.9 per cent (18.9 and 13.6% in case of pulmonary and extrapulmonary samples, respectively). Our study revealed a significant increase (P<0.001) in multidrug resistance by 12 per cent (4.7% in 2000 to 19.8% in 2012). MDR-TB was more in case of pulmonary (28.2%) than extrapulmonary (11.6%) TB (P<0.001). Only 6.5 per cent (154) of mycobacterial isolates were non-tuberculous mycobacteria and rapid growers represented by Mycobacterium fortuitum and M. abscessus were the most commonly isolated species. Interpretation & conclusions: Increase in prevalence of MDR-TB by 12 per cent in the past 13 years is alarming. Policy modifications may have to be done to strengthen the existing TB control programmes to encourage contact tracing and culture and drug susceptibility testing for all smear positive pulmonary cases to ensure early and appropriate therapy

Ecology of blood stream infection and antibiotic resistance in intensive care unit at a tertiary care hospital in North India

OBJECTIVE: To analyse the prevalent microorganisms and their antimicrobial resistance among intensive care unit patients in a tertiary care centre in New Delhi.METHODS: A retrospective study of all consecutive blood cultures from various intensive care unit patients in the hospital during four years (January 2008 to December 2011). Antibiotic consumption data in the intensive care units were also analysed during the same period.RESULTS: Out of the total 22,491 blood cultures processed, 2846 samples were positive and 3771 microorganisms were isolated. The blood culture positivity was estimated as 12.7% of which 67.5% were monomicrobial and 32.5% polymicrobial infections. Gram negative bacilli, Gram positive cocci, and fungi were isolated in 49%, 33%, and 18% cases, respectively. Coagulase negative staphylococcus was the commonest single isolate followed by Candida spp. A drastic shift in the distribution of Candida spp. towards nonalbicans along with high resistance to azole group of antifungals suggest echinocandins for the empiric therapy of candidemia. High penicillin resistance in Gram positive isolates suggest vancomycin, linezolid and tigecycline as the options for empiric therapy, whereas tigecycline and colistin are the only options remaining for highly resistant Gram negative isolates. Aminoglycosides were observed to have better sensitivity and reduced usage when compared with cephalosporins and ß-lactam + ß-lactam inhibitor combinations.CONCLUSIONS: High frequencies of multidrug resistant organisms were observed in intensive care units which is a warning as to use the only few effective antimicrobials wisely to reduce selective pressure on sensitive strains

Multi-omics approaches for remediation of bisphenol A: Toxicity, risk analysis, road blocks and research perspectives

In this &quot;plastic era&quot; with the increased use of plastic in day today&apos;s life the accumulation of its degraded products like microplastics or plastic additives such as Bisphenol A(BPA) is also increasing. BPA is an endocrine-disrupting chemical used as a plasticizing agent in clear plastic, building materials, coatings, and epoxy resin. Several enzymes including laccases and lipases have been studied for the reduction of BPA toxicity. Over the decades of encountering these toxicants, microorganisms have evolved to degrade different classes of plastic additives. Since the degradation of BPA is a long process thus meta-omics approaches have been employed to identify the active microbiota and microbial dynamics involved in the mitigation of BPA. It is also necessary to investigate the impact of processing activities on transit of BPA in food items and to limit its entrance in food world. This review summarizes a comprehensive overview on BPA sources, toxicity, bio-based mitigation approaches along with a deeper understanding of multi-omics approaches for its reduction and risk analysis. Knowledge gaps and op-portunities have been comprehensively compiled that would aid the state-of-the-art information in the available literature for the researchers to further address this issue

Transmission of B.1.617.2 Delta variant between vaccinated healthcare workers

AbstractBreakthrough infections with SARS-CoV-2 Delta variant have been reported in doubly-vaccinated recipients and as re-infections. Studies of viral spread within hospital settings have highlighted the potential for transmission between doubly-vaccinated patients and health care workers and have highlighted the benefits of high-grade respiratory protection for health care workers. However the extent to which vaccination is preventative of viral spread in health care settings is less well studied. Here, we analysed data from 118 vaccinated health care workers (HCW) across two hospitals in India, constructing two probable transmission networks involving six HCWs in Hospital A and eight HCWs in Hospital B from epidemiological and virus genome sequence data, using a suite of computational approaches. A maximum likelihood reconstruction of transmission involving known cases of infection suggests a high probability that doubly vaccinated HCWs transmitted SARS-CoV-2 between each other and highlights potential cases of virus transmission between individuals who had received two doses of vaccine. Our findings show firstly that vaccination may reduce rates of transmission, supporting the need for ongoing infection control measures even in highly vaccinated populations, and secondly we have described a novel approach to identifying transmissions that is scalable and rapid, without the need for an infection control infrastructure.</jats:p