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Background: Staphylococcus aureus is a common cause of hospital-acquired infections. Medical undergraduates are frequently exposed to hospital-adapted bacterial strains especially S. aureus and therefore are always at risk of colonisation. Materials and Methods: A prospective study was conducted on 150 undergraduate students, of different semesters, to detect S. aureus nasal colonisation. A detailed history regarding attending the hospital ward teaching, hospitalisation, frequency of nose touching and immunocompromised condition was unequivocally obtained from all the participants. Anterior nares were swabbed and inoculated on mannitol salt and blood agar plates. S. aureus isolates were subsequently verified by conventional microbiological techniques. The antibiotic susceptibility testing was done by the modified Kirby-Bauer disc diffusion method. In addition, PCR-based detection of mecA gene and pvl gene was performed. Results: Out of 150 students, we found 18 students (3 of 1st, 10 of 5th and 5 of 9th semester, respectively) to be colonised by S. aureus. Of these 18 isolates, 8 were found to be methicillin-resistant. We observed a significant association (P < 0.05) of hospital exposure, ward teaching/hospitalisation with S. aureus colonisation. These S. aureus isolates exhibited varying susceptibility towards different antibiotics with multiple antibiotic resistance indexes ranging from 0.08 to 0.58. Interestingly, all eight methicillin-resistant S. aureus isolates displayed mecA positivity while all were found to be pvl negative. Conclusion: The results indicate the importance of imparting knowledge about infection control practices during the initial years of undergraduate training in medical schools, which may increase awareness amongst students before getting outpatient or ward postings. [ABSTRACT FROM AUTHOR]

The study is aimed at establishing the optimal parameters for RNA purification of pooled specimens, in SARS-CoV-2 assay. This research work evaluates the difference of extracted RNA purity of pooled samples with and without treatment with isopropyl alcohol and its effect on real-time RT-PCR. As per the protocol of the Indian Council of Medical Research (ICMR), 5 sample pools were analysed using qRT-PCR. A total of 100 pooled samples were selected for the study by mixing 50 μL of one COVID-19 positive nasopharyngeal/oropharyngeal (NP/OP) specimen and 50 μL each of 4 known negative specimens. Pool RNA was extracted using the column-based method, and 1 set of pooled extracted RNA was tested as such, while RNA of the second set was treated additionally with chilled isopropyl alcohol (modified protocol). Further, the purity of extracted RNA in both the groups was checked using Microvolume Spectrophotometers (Nanodrop) followed by RT-PCR targeting E-gene and RNaseP target. The results showed that the purity index of extracted RNA of untreated pooled specimens was inferior to isopropyl alcohol-treated templates, which was observed to be 85% sensitivity and 100% specificity. The average Cq (E gene) in the unpurified and purified pool RNA group was 34.66 and 31.48, respectively. The nanodrop data suggested that purified RNA concentration was significantly increased with an average value of 24.73 ± 1.49 ng / uL , which might be the reason for high sensitivity and specificity. Thus, this group testing of SARS-CoV-2 cases using pools of 5 individual samples would be the best alternative for saving molecular reagents, personnel time, and can increase the overall testing capacity. However, purity of RNA is one of the important determinants to procure unfailing results, thus, this additional purification step must be included in the protocol after RNA has been extracted using commercially available kit before performing qRT-PCR. [ABSTRACT FROM AUTHOR]

Introduction: There are 15 high risk genotypes of Human Papilloma Viruses (HPV) which are associated with Carcinoma Cervix (CaCx), of which genotype 16 and 18 are the commonest. Aim: To evaluate nested multiplex Polymerase Chain Reaction (PCR) protocol for simultaneous detection of HPV, and typing of high risk genotypes mainly genotypes 16 and 18 in cervical scrape/biopsy samples. Materials and Methods: This prospective cohort study was done on 63 females in the age group 21-70 years attending outpatient Department of Gynaecology, Sir Sunderlal Hospital, BHU, Uttar Pradesh, India from March 2015 to August 2016. Cervical biopsy (n=14) and cervical scrape (n=49) samples from 63 females were subjected to histopathology/cytology and nested multiplex PCR for detection and typing of HPV. Further, in HPV positive tissue samples, in which virus was not typed as genotype 16 and 18, 142 bp consensus sequence of L1 capsid gene was amplified by nested PCR employing MY/GP+ primers and subjected for sequencing to ascertain the HPV genotypes. Results: On histological examination, 13 (92.8%) out of 14 biopsies were diagnosed as cases of Squamous Cell Carcinoma (SCC). Of the 13 cases of SCC of uterine cervix, all were HPV positive and yielded either 134 bp L1 capsid amplicons (n=12) or HPV 16 specific amplicon (n=1). On subjecting cervical scrape samples for the PCR, 24.3%, 100%, 57.1%, and 66.7% cases with Negative for Intraepithelial Lesion or Malignancy (NILM), Atypical Squamous Cells of Undetermined Significance (ASCUS), Low Grade Intraepithelial Lesion (LSIL), and High Grade Intraepithelial Lesion (HSIL) respectively were positive for HPV. Conclusion: Detection of High Risk HPVs (HR HPV) in all the tissue samples of SCC of uterine cervix indicates the usefulness of the nested multiplex PCR protocol in detection and typing of HPVs in tissue samples of CaCx. Further, this protocol may be used to detect HPVs in scrape samples along with cervical cytology as a tool for screening CaCx. [ABSTRACT FROM AUTHOR]

Background: The Ganga River is a life-line, a symbol of purity and virtue for countless people of India. Nearly all the sewage, industrial effluent, runoff from chemical fertilizers and pesticides used in agriculture within the basin, and large quantities of solid waste, including thousands of animals carcasses and hundreds of human corpses are dumped in the river everyday which posed a considerable public health threat to the religious bathers and a large number of people living along the river who uses Ganges water for drinking and other household purposes. Objective: To assess the physiochemical and biological quality of river Ganges water and to study the seasonality pattern of pollution of River Ganges. Materials and Methods: Two points namely Samneghat and Rajghat (entry and exit point of Ganga into the city) were selected as the first and the last sampling stations, beside 8 other intermediate sampling stations to assess the physiochemical and biological quality of River Ganga water. The study was conducted from April 2013 to March 2014 and this period was crudely divided into three seasons viz Winter, Summer and Rainy season for the purpose of studying seasonality pattern of pollution. Results: As the river progress through city of Varanasi, Total dissolved solids (TDS), Chloride, Conductivity, Biochemical Oxygen Demand (BOD), Nitrate, Nitrite and Most Probable Number (MPN) increases, DO decreases progressively and Temperature and pH remain same. Conclusion: As far as quality of water with regard to recreational and industrial purposes are concerned, Ganges water may be considered useful but most of the parameters observed across different seasons indicates that it may not be used as potable water. [ABSTRACT FROM AUTHOR]

Background: Control of the global burden of tuberculosis is obstructed due to lack of simple, rapid and cost effective diagnostic techniques that can be used in resource poor-settings. To facilitate the early diagnosis of TB directly from clinical specimens, we have standardized and validated the use of nested multiplex PCR, targeting gene fragments IS6110, MTP40 and 32kD α-antigen encoding genes specific for Mycobacterium tuberculosis complex and non-tubercular mycobacteria (NTM), in comparison to smear microscopy, solid culture and single step multiplex PCR. The results were evaluated in comparison to a composite reference standard (CRS) comprising of microbiological results (smear and culture), clinical, radiological and cytopathological findings, clinical treatment and response to anti-tubercular therapy.Methods: The nested multiplex PCR (nMPCR) assay was evaluated to test its utility in 600 (535 pulmonary and 65 extra-pulmonary specimens) clinically suspected TB cases. All specimens were processed for smear, culture, single step multiplex PCR and nested multiplex PCR testing.Results: Out of 535 screened pulmonary and 65 extra-pulmonary specimens, 329 (61.5%) and 19 (29.2%) cases were culture positive for M. tuberculosis. Based on CRS, 450 patients had "clinical TB" (definitive-TB, probable-TB and possible-TB). Remaining 150 were confirmed "non-TB" cases. For culture, the sensitivity was low, 79.3% for pulmonary and 54.3% for extra-pulmonary cases. The sensitivity and specificity results for nMPCR test were evaluated taken composite reference standard as a gold standard. The sensitivity of the nMPCR assay was 97.1% for pulmonary and 91.4% for extra-pulmonary TB cases with specificity of 100% and 93.3% respectively.Conclusion: Nested multiplex PCR using three gene primers is a rapid, reliable and highly sensitive and specific diagnostic technique for the detection and differentiation of M. tuberculosis complex from NTM genome and will be useful in diagnosing paucibacillary samples. Nested multiplex PCR assay was found to be better than single step multiplex PCR for assessing the diagnosis of TB. [ABSTRACT FROM AUTHOR]

Abstract: Eighty-six strains of Acinetobacter baumannii from India and Nepal were investigated for the presence of integrons in relation to multiple drug resistance by integrase gene polymerase chain reaction (PCR). Integrons were found to be present at a rate of 43.02% (37/86). Integrons were significantly correlated with multidrug resistance to several antibiotics. Class 1 integrons were detected in 81.1% of integron-positive strains, whilst 18.9% were found to be positive for class 2 integrons. The majority of class 2 integrons (71%) were encountered in strains isolated from post-operative wards of both countries. The highest integron carriage in isolates of A. baumannii (63.6%) was observed in 2005. Hence, it is likely that integrons play an important role in antibiotic resistance and possibly indicate epidemic behaviour of A. baumannii. Integrase gene PCR may be used as routine screening and identification for the surveillance of clinical isolates of A. baumannii with epidemic potential. [Copyright &y& Elsevier]

Abstract: In the present study, genetic diversity analysis of Mycobacterium tuberculosis isolated from patients attending a tertiary care hospital, North India, has been attempted. Eighty three isolates of M. tuberculosis were subjected to DNA fingerprinting using spoligotyping and IS6110-RFLP techniques. Spoligotype patterns showed that central Asian (32.5%), ill defined T (13.2%) and Beijing (10.8%) families were predominant in ongoing transmission of the bacterium. Two STs; ST26 (CAS_Delhi) and ST1 (Beijing) represented 36.1% of the total M. tuberculosis population in eastern Uttar Pradesh, North India. IS6110 RFLP analysis showed that isolates having low and zero copy number of the IS element were 15.6% and 19.2%, respectively. Out of the 47 isolates clustered by spoligotyping, 40 could be further differentiated as unique strains by IS6110-RFLP. Therefore, this study recommends that both the techniques be used simultaneously for DNA fingerprinting of M. tuberculosis in India. [Copyright &y& Elsevier]