Shelf life enhancement of guava (Psidium guajava cv. Baruipur) stored under pilot scale modified atmosphere storage system.

PRACTICAL APPLICATION: The developed modified atmosphere storage (MAS) system has increased the shelf life of Guava up to 20 and 9 days at 10 °C and 25 °C, respectively. This extended period will be very much effective for providing the buffer period to the fresh Guava and facilitate extra time to the farmers for its marketing. The uniquely developed MAS system is helpful for the farmers for on-farm storage of fresh Guava at a large scale and will provide smooth handling and transportation for retailing and marketing.

Multilocus Sequence Typing of Clinical Isolates of Cryptococcus from India.

Cryptococcosis is a life-threatening infection caused by Cryptococcus neoformans and C. gattii species complex. In the present study, to understand the molecular epidemiology of 208 clinical isolates of Cryptococcus from different parts of India, multilocus sequence typing (MLST) using ISHAM MLST consensus scheme for C. neoformans/C. gattii species complex was used. MLST analysis yielded a total of 10 Sequence Types (STs)-7 STs for C. neoformans and 3 for C. gattii species complex. The majority of isolates identified as C. neoformans belonged to molecular type VNI with predominant STs 31 and 93. Only 3 isolates of C. gattii species complex were obtained, belonging to ST58 and ST215 of VGI and ST69 of VGIV. Phylogenetic analysis revealed less diversity among the clinical Indian isolates compared to the global MLST database. No association between prevalent STs and HIV status, geographical origin or minimum inhibitory concentration (MIC) could be established.

Diagnostic accuracy of nucleic acid amplification based assays for tuberculous meningitis: A meta-analysis.

BACKGROUND: Numerous in-house and commercial nucleic acid amplification tests (NAAT) have been evaluated using variable reference standards for diagnosis of TBM but their diagnostic potential is still not very clear. METHODS: We conducted a meta-analysis to assess the diagnostic accuracy of different NAAT based assays for diagnosing TBM against 43 data sets of confirmed TBM (n = 1066) and 61 data sets of suspected TBM (n = 3721) as two reference standards. The summary estimate of the sensitivity and the specificity were obtained using the bivariate model. QUADAS-2 tool was used to perform the Quality assessment for bias and applicability. Publication bias was assessed with Deeks' funnel plot. RESULTS: Studies with confirmed TBM had better summary estimates as compared to studies with clinically suspected TBM irrespective of NAAT and index tests used. Among in-house assays, MPB as the gene target had best summary estimates in both confirmed [sensitivity:90%(83-95), specificity:97-%(87-99), DOR:247 (50-1221), AUC:99%(97-100), PLR:38.8-(6.6-133), NLR:0.11(0.05-0.18), I2 = 15%] and clinically suspected [sensitivity:69%(47-85), specificity:96%(90-98), DOR:62(16.8-232), AUC:94%(92-97), PLR:16.9(6.5-36.8), NLR:0.33(0.16-0.56), I2:15.3%] groups. GeneXpert revealed good diagnostic accuracy only in confirmed TBM group [sensitivity = 57%(38-74), specificity = 98%(89-100), DOR = 62(7-589), AUC = 87%(79-96), PLR = 33.2(3.8-128), NLR = 0.45(0.26-0.68), I2 = 0%]. CONCLUSIONS: This meta-analysis identified potential role of MPB gene among in-house assays and GeneXpert as commercial assay for diagnosing TBM.

Isoniazid and rifampicin heteroresistant Mycobacterium tuberculosis isolated from tuberculous meningitis patients in India.

BACKGROUND: Heteroresistant Mycobacterium tuberculosis (mixture of susceptible and resistant subpopulations) is thought to be a preliminary stage to full resistance and timely detection, initiation of correct treatment is vital for successful anti tubercular therapy. The aim of this study was to detect multi drug resistant (MDR) and heteroresistant M. tuberculosis with the associated gene mutations from patients of tuberculous meningitis. METHODS: A total of 197 M. tuberculosis isolates from 478 patients of TBM were isolated from July 2012 to July 2015 and subjected to drug susceptibility testing (DST) by BACTEC MGIT and Genotype MTBDR line probe assay (LPA). Heteroresistance was defined as presence of both WT and mutant genes in LPA. RESULTS: Of 197 M. tuberculosis isolates, 11 (5.6%) were MDR, 23 (11.6%), 1 (0.5%) were mono resistant to isoniazid (INH) and rifampicin (RMP) respectively. Heteroresistance was detected in 8 (4%), 2 (1%) isolates to INH and RMP respectively. INH heteroresistant strains had WT bands with mutation band S315T1 whereas RMP heteroresistant strains had WT bands with mutation band S531L. CONCLUSION: The prevalence of MDR M. tuberculosis was 5.6% in TBM patients with the most common mutation being ΔWT band with S315T1 for INH and ΔWT band with S531T for RMP. MGIT DST was found to be more sensitive for detecting overall resistance in M. tuberculosis but inclusion of LPA not only reduced time for early initiation of appropriate treatment but also enabled detection of heteroresistance in 8 (4%), 2 (1%) isolates for INH and RMP respectively.

Predictors of adverse outcome in patients of tuberculous meningitis in a multi-centric study from India.

INTRODUCTION: This study aimed to investigate the factors which may predict mortality and neurological disability at one year follow up in patients of tuberculous meningitis (TBM) in India. METHODOLOGY: Patients with TBM were prospectively enrolled from July 2012 to September 2014 from four tertiary care hospitals of Delhi. The demographic characteristics, clinical features and laboratory findings were collected and patients were followed up till 1 year. These were analyzed by univariate and multivariate multinomial logistic regression analysis to identify predictors of adverse patient outcome at 1 year follow up. RESULTS: Out of 478 patients enrolled, 391 patients could be followed up to 1 year. Sixty-four patients (16.3%) died and 150 patients (39%) survived with one or more neurological disability. Altered sensorium, motor deficit, cranial nerve palsy, seizures, isolation of M. tuberculosis and presence of multi-drug resistance were independently associated with any adverse outcome (death or disability) but by multivariate analysis only motor deficit, altered sensorium and isolation of M. tuberculosis on culture produced a statistically significant model for prediction of patient outcome. CONCLUSION: The three-predictor model with motor deficit, altered sensorium and isolation of M. tuberculosis produced a statistically significant model with correct prediction rate of 60.4%. These three variables predicted death with odds ratio of 39.2, 6.7 and 2.1 respectively in comparison to recovery whereas only motor deficit and isolation of M. tuberculosis predicted neurological disability at 1 year with odds ratio of 3.9, 2.4 respectively.

Evaluation of Geno Type MTBDRplus Line Probe Assay for Early Detection of Drug Resistance in Tuberculous Meningitis Patients in India.

BACKGROUND: Molecular methods which allow for rapid and reliable detection of drug resistance have yet not been sufficiently evaluated for timely management of patients with tuberculous meningitis. AIMS: We aimed to evaluate Geno Type MTBDRplus line probe assay for early detection of drug resistance in Mycobacterium tuberculosis isolates and CSF samples of confirmed tuberculous meningitis patients. SETTINGS AND DESIGN: This was a multicentric prospective study carried out from July 2011 to December 2013 in tertiary care hospitals of Delhi. MATERIALS AND METHODS: The assay was performed on 89 M. tuberculosis isolates and 31 direct CSF samples from microbiologically confirmed tuberculous meningitis patients. The sensitivity and specificity of this assay was calculated in comparison to drug susceptibility testing by BACTEC MGIT 960 system. RESULTS: The sensitivity, specificity for detection of resistance to Isoniazid was 93%, 97% and to Rifampicin was 80%, 98.8%, respectively by this assay in comparison with the phenotypic drug susceptibility testing. The line probe assay could detect M. tuberculosis in 55% of CSF samples from patients with microbiologically confirmed tuberculous meningitis. Only 5/89 isolates (5.6%) were resistant to both Isoniazid and Rifampicin while 9/89 (10%) isolates were additionally resistant to Isoniazid. Resistance to any of the drugs, namely Isoniazid, Rifampicin, Streptomycin or Ethambutol, was seen in 24.7% of strains. CONCLUSION: The line probe assay has a good sensitivity and specificity for detection of drug resistance to Isoniazid and Rifampicin in M. tuberculosis culture isolates. However, this assay has limited role in detection of M. tuberculosis and drug resistance from direct samples with confirmed diagnosis of tuberculous meningitis.

Human soft tissue infection by the emerging pathogen Shewanella algae.

Shewanella soft tissue infections usually occur in immunocompromised patients with a preexisting cutaneous ulcer, mostly after exposure to a marine environment or contaminated water. A 35-year-old male presented with a non-healing ulcer over the distal end of his right leg but had no predisposing factors.  Cultures of exudates from the wound grew Shewanella on repeated occasions. Recovery was uneventful following surgical debridement and antimicrobial therapy. Early suspicion, diagnosis, and treatment with potent antibiotics are needed to prevent any further complications resulting from infection by this emerging pathogen.

Comparison of DNA Extraction Protocols for Mycobacterium Tuberculosis in Diagnosis of Tuberculous Meningitis by Real-time Polymerase Chain Reaction.

BACKGROUND: Several nucleic acid amplification techniques are available for detection of Mycobacterium tuberculosis (MTB) in pulmonary and extrapulmonary samples, but insufficient data are available on the diagnostic utility of these techniques in tubercular meningitis where bacilli load is less. The success of final amplification and detection of nucleic acid depends on successful extraction of DNA from the organism. AIMS: We performed this study to compare four methods of extraction of MTB DNA from cerebrospinal fluid (CSF) samples so as to select one method of DNA extraction for amplification of nucleic acid from clinical samples. MATERIALS AND METHODS: Four methods of extracting MTB DNA from CSF samples for testing by real-time polymerase chain reaction (PCR) were compared: QIAGEN(R) protocol for DNA purification using QIAamp spin procedure (manual), AMPLICOR(R) respiratory specimen preparation kit, MagNA Pure(R) kit extraction, combined manual DNA extraction with automated extraction by MagNA Pure(R). Real-time PCR was performed on COBAS TaqMan 48 Analyzer(R) with known positive and negative controls. RESULTS: The detection limit for the combined manual and MagNA Pure(R) extraction protocol was found to be 100 copies of MTB DNA per reaction as against 1,000 copies of MTB DNA per reaction by the QIAGEN(R), AMPLICOR(R), and the MagNA Pure(R) extraction protocol. CONCLUSION: The real-time PCR assay employing the combination of manual extraction steps with MagNA Pure(R) extraction protocol for extraction of MTB DNA proved to be better than other extraction methods in analytical sensitivity, but could not detect less than 10(2) bacilli /ml.

Laboratory diagnosis of tuberculous meningitis - is there a scope for further improvement?

AIMS: Tuberculous meningitis (TBM) still remains a diagnostic challenge because of inconsistent clinical presentation and lack of rapid, sensitive and specific tests. This study was carried out to diagnose TBM by a combination of direct microscopy on Ziehl-Neelsen (ZN) staining, culture by conventional Lowenstein Jensen (LJ) media and Bactec MGIT 960 system in clinically suspected cases, supported by laboratory parameters. MATERIALS AND METHODS: A total of 164 cerebrospinal fluid (CSF) samples from suspected cases of TBM were processed for direct acid fast bacilli (AFB) smear examination, and culture on Bactec MGIT 960 and LJ media. RESULTS: AFB were detected on direct smears in 13 of 164 (7.9%) specimens and Mycobacterium tuberculosis was isolated by at least one of the culture methods from 49 (29.8%) CSF samples of which 45 (27.4%) were detected positive for M. tuberculosis by MGIT 960 culture and 18 (10.9%) by culture on LJ medium. The mean time of detection in MGIT and LJ media for M. tuberculosis were 18 and 38 days, respectively. CONCLUSIONS: A combination of laboratory parameters like smear microscopy, conventional culture and automated method like Bactec MGIT increases the sensitivity of diagnosing TBM as compared to any single method.

Prevalence of HIV-associated cryptococcal meningitis and utility of microbiological determinants for its diagnosis in a tertiary care center.

CONTEXT: Human immunodeficiency virus (HIV) infection continues to be the most important risk factor for the development of central nervous system (CNS) cryptococcosis, which in turn is an important contributor to morbidity and mortality in HIV-infected patients. Early diagnosis of such patients is the key to their therapeutic success. AIMS: This study was undertaken to find out the prevalence of CNS cryptococcosis and to assess the role of microbiological parameters for its specific diagnosis in HIV-reactive hospitalized patients admitted with meningeal signs in a tertiary care setting. MATERIALS AND METHODS: A total of 104 patients suspected to be suffering from meningitis/meningoencephalitis were subjected to cerebrospinal fluid (CSF) analysis (including India ink preparation, culture by conventional methods and Bactec MGIT 960 system, antigen detection) and tests for HIV antibodies by standard laboratory operating procedures. RESULTS: The prevalence of HIV infection in our study group was 12.5% (13/104), while the prevalence of cryptococcal CNS infection in HIV-reactive cohort was 46% (6/13). Additionally, 15.3% (2/13) of the patients from this cohort were positive for Mycobacterium tuberculosis. CONCLUSIONS: High prevalence of cryptococcal CNS infections in HIV-infected patients underscores the importance of precise and early microbiological diagnosis for better management of such patients.

A technically simple method for staining of acid-fast bacilli in cytology smears: an evaluation.

OBJECTIVE: To study the effects of modifications in the Ziehl-Neelsen staining procedure on predictive accuracy for acid fast bacilli in comparison to the conventional technique. Simplicity of procedure and reagent economy were the factors taken into consideration. DESIGN: Comparative evaluation between thick and thin air-dried smears stained conventionally and thick ethanol-fixed smears stained by the modified technique was done. RESULTS: Positive predictive accuracy of all the three smears, that is, thick air-dried, thin air-dried and thick ethanol-fixed, was 100%. Negative predictive accuracy for thick air-dried, thin air-dried and thick ethanol-fixed smears was 36.36%, 32.33% and 34.78%, respectively. Overall predictive accuracy was 66.67% for thick air-dried, 61.90% for thin air-dried and 64.29% for thick ethanol-fixed. These differences were found to be statistically insignificant. CONCLUSION: The modified method offers an accuracy comparable to the conventional technique, is simpler and with improved reagent economy. It is of special importance to diagnostic facilities in rural set-ups.

Mitochondrial encephalomyopathies: advances in understanding.

Mitochondrial encephalomyopathies encompass a group of disorders that have impaired oxidative metabolism in skeletal muscles and central nervous system. As the field of mitochondrial medicine takes shape and physicians in all specialties become increasingly aware of respiratory chain or oxidative phosphorylation (OXPHOS) related disorders, their prevalence remains largely unknown. The unique features of the mitochondrial genome and the dual control over this important cellular apparatus makes the clinical presentation variable and diagnosis difficult. There is a confounding variation in phenotype and genotype, and the natural history of the disorders in individual patients is not accurately predictable. Only recently have things begun to fall into place and some phenotypes defined. Diagnosis requires a complex battery of clinical studies coupled with diagnostic findings on muscle biopsy (abnormal structure, histochemistry, or enzyme studies) or DNA testing. However, a reasonably confident diagnosis can be made by viewing the clinical presentation in the light of family history and some basic, routinely available laboratory investigations. This review tries to give a brief account of mitochondrial structure, function and genetics, and clinical presentation, evaluation, and treatment in suspected cases of mitochondrial encephalomyopathies.

Neuropathology of schizophrenia--a review.

Despite clinical evidence of brain dysfunction in schizophrenia, little progress was made for most of the last century in determining its organic parameters. Neuropathology, over the past few decades, has made a substantial contribution to the understanding of cellular and molecular mechanisms of schizophrenia. During the last 10-15 years the concept of schizophrenia as a 'functional' psychosis has changed to the current paradigm of schizophrenia as a neurodevelopmental disorder. Much still has to be unravelled and learnt. This review gives a brief account of the relevant neuroanatomy, viral hypotheses of schizophrenia etiology, pathologic findings reported, concept of neurodevelopmental model and avenues for the future.

Human prion diseases.

Prion diseases is another name for a group of 'transmissible spongiform encephalopathies'. Creutzfeldt-Jakob disease, the first prion disease described in humans, occurs in sporadic, familial or iatrogenic form. Other transmissible spongiform encephalopathies in humans such as familial Creutzfeldt-]akob disease, Gerstmann-Sträussler-Scheinker disease and fatal familial Insomnia have been shown to be associated with specific prion protein gene mutations. In 1996, a new variant of Creutzfeldt-Jakob disease was reported in the United Kingdom among young patients with unusual clinical features and unique neuropathological findings. This new form could be due to transmission to humans of the agent causing bovine spongiform encephalopathy. While examination of brain tissue is the key to making a diagnosis, it is not always possible antemortem. Immunological tests such as ELISA or western blot assays along with tests for 1 4-3-3 protein in the cerebrospinal fluid remain the main tools of diagnosis. Conventional disinfection and sterilization practices are Ineffective for these agents. The unusual properties of prions pose a challenge for treatment, surveillance and control of these diseases.