Characterisation of OXA-48-like carbapenemases in Escherichia coli and Klebsiella pneumoniae from North India.

The oxacillinase-48 (OXA-48)-like carbapenemases are class D ß-lactamases and increasingly reported in Enterobacterial species. The detection of these carbapenemases is challenging and little information is available on the epidemiology and plasmid characteristics of OXA-48-like carbapenemase producers. We detected the presence of OXA-48-like carbapenemases in 500 clinical isolates of Escherichia coli and Klebsiella pneumoniae, followed by detection of other carbapenemases, extended spectrum ß-lactamases (ESBLs) and 16S rRNA methyltransferases in OXA-48 producers. Clonal relatedness was studied using pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST). Finally, plasmid characterisation was performed through conjugation experiment, S1-PFGE and Southern hybridisation. Around 40% of E. coli and K. pneumoniae isolates harboured OXA-48-like ß-lactamases. Two OXA-48 allele variants, OXA-232 and OXA-181 were detected in our study. OXA-48 producers co-harbored diverse drug-resistant genes belonging to other classes of carbapenemases, ESBLs and 16S rRNA methyltransferases. OXA-48-like carbapenemase producers exhibited high clonal diversity. Bla OXA-48 carrying plasmids were conjugative, untypable and their size was ~ 45 kb and ~ 104.5 kb in E. coli and K. pneumoniae respectively. In conclusion, OXA-48-like carbapenemases have emerged as major cause of carbapenem resistance in Enterobacteriaceae and probably still being under reported. Strict surveillance and adequate detection methods are needed to prevent the dissemination of OXA-48-like carbapenemases.

Localized outbreaks of Pseudomonas aeruginosa belonging to international high-risk clones in a south Indian hospital.

Introduction. Pseudomonas aeruginosa is now considered as a major bacterial pathogen associated with hospital infections. Frequently, multidrug-resistant (MDR) and extensively drug-resistant (XDR) P. aeruginosa are being encountered. Unusual increase in the P. aeruginosa infections led to the suspicion of outbreaks in the urology ward and cardiothoracic and vascular surgery intensive care unit (CTVS-ICU).Hypothesis. We hypothesize that the localized outbreaks may have originated from environmental sources within the hospital premises. An alternative possibility is the transmission from a previously infected patient or hospital attendant. Understanding the drug-resistance profile and genome characteristics of these clinical samples would determine the likely source of infection and spread.Aim. To perform epidemiological and molecular investigations on the suspected outbreaks of P. aeruginosa in the study centre and identify potential sources of infection.Methodology. Fourteen drug-resistant P. aeruginosa isolated from patients of the urology ward, CTVS-ICU and tap waters collected during the suspected outbreaks were subjected to microbiological and genomic analysis. Comparative genome (CG) analysis of these 14 study genomes with 284 complete P. aeruginosa genomes was performed.Results. Multilocus sequence typing analysis revealed that the isolates belonged to five different sequence types (ST235, ST357, ST639, ST654 and ST1203) and clustered into three distinct groups while two CTVS-ICU isolates remained as singletons. Genome analysis distinguished that the outbreaks in the urology ward and CTVS-ICU are independent, epidemiologically unrelated to each other and with the tap-water isolates.Conclusion. This study highlights the presence of distinct, clonally unrelated, drug-resistant P. aeruginosa within a hospital setting. The genome analysis of the two localized outbreaks revealed their distinct genetic background and phylogenetically unrelated origin. Vigilant screening and effective implementation of infection control measures led to the successful containment of potential environmental reservoirs of P. aeruginosa within the premises.

My experience on taeniasis and neurocysticercosis.

Taeniasis and neurocysticercosis (NCC) are major public health problems in developing countries. NCC is the leading cause of community-acquired active epilepsy. NCC may present as a medical emergency, especially when there is cysticercotic encephalitis or raised intracranial hypertension. Systematic community-based studies on taeniasis and NCC are lacking. We studied taeniasis and NCC-related active epilepsy disease burden in the pig farming community of Lucknow district, Uttar Pradesh, India. Based on the 30 cluster sampling approach as recommended by the World Health Organization, we estimated the prevalence of taeniasis, NCC-related active epilepsy, and silent NCC in the community. We also estimated the prevalence of swine cysticercosis. Taeniasis was detected in 18.6% of populations. Expulsions of tapeworm segments in stool, consumption of undercooked pork, age above 15 years, and handwash with clay or plain water after defecation were associated with taeniasis. On molecular analyses of positive stool samples, T. solium was identified in 40% and Taenia asiatica in 60% of cases. Active epilepsy was identified in 5.8% of subjects; 48% of them had NCC. On neuroimaging, NCC was detected in 15% of asymptomatic individuals. We observed that host genetic factors such as toll-like receptor-4, matrix metalloproteinase-9, intercellular adhesion molecule-1, and glutathione-S transferase gene polymorphisms were associated with seizure in NCC. When peripheral blood mononuclear cells (PBMCs) from NCC subjects were exposed to cysticerci fluid antigens in-vitro, PBMCs from symptomatic and asymptomatic subjects showed significantly higher Th 1 and Th 2 cytokines response respectively, symptomatic patients had significant Th-1 cytokines response, while asymptomatic individuals showed Th-2 response. Porcine cysticercosis was detected in 26% of swine; 38% of them had cysticerci in the brain. Swine with brain involvement showed clinical signs such as excessive salivation, excessive blinking and tearing, and subconjunctival nodule. On molecular analysis, 15% of cysticerci in swine were identified as T. asiatica. Infected swine when treated with albendazole plus/minus steroid, the response rate of cysticerci (either dead or resolved lesion) was 100% in albendazole-treated group and 71% in albendazole plus steroid-treated group. The above studies suggest that taeniasis and NCC are alarmingly high in the pig farming community of North India. Taeniasis in human and cysticercosis in swine due to T. asiatica call for further studies on this parasite.

Complete genome sequence of an extensively drug-resistant Pseudomonas aeruginosa ST773 clinical isolate from North India.

OBJECTIVES: Pseudomonas aeruginosa is an opportunistic Gram-negative pathogen causing a wide range of community- and hospital-acquired infections. Here we report the complete genome sequence of an extensively drug-resistant (XDR) P. aeruginosa strain (PA790) in order to understand the antibiotic resistance genes (ARGs) harboured by such a strain. METHODS: Whole-genome sequencing (WGS) was performed using Illumina HiSeq and Nanopore MinION platforms. Genome assembly was performed using Unicycler v.0.4.8. The genome was annotated using the NCBI Prokaryotic Genome Annotation Pipeline (PGAP). In silico predictions were fulfilled using curated bioinformatics tools. RESULTS: Pseudomonas aeruginosa PA790 was classified as XDR and belongs to sequence type 773 (ST773). The complete genome size is 6 932 250 bp with a G+C content of 66.02% and a BUSCO (Benchmarking Universal Single-Copy Orthologs) score of 100. Strain PA790 harboured 12 different ARGs conferring resistance to eight different classes of antibiotics. It was identified as the nineteenth ST773 strain among 5785 whole-genome sequences of P. aeruginosa available in the NCBI database. CONCLUSION: Pseudomonas aeruginosa PA790 belongs to ST773 and was identified as the nineteenth such isolate to be submitted to NCBI and the first complete ST773 genome from India. The WGS data with multiple ARGs of P. aeruginosa PA790 (ST773) will aid in understanding the evolution and phylogeny of such high-risk clones and provide a solid basis for further research on XDR strains.

Genetic Characterisation of Colistin Resistant Klebsiella pneumoniae Clinical Isolates From North India.

Background: Increasing use of colistin has led to the world-wide emergence of mobile colistin resistant gene (mcr). The present study aimed to identify and characterise mcr and other drug-resistant genes in colistin resistant Klebsiella pneumoniae clinical isolates. Methods: Twenty-two colistin resistant K. pneumoniae were analysed for mcr and other drug-resistant genes, efflux pumps, and virulence genes, and for their biofilm forming ability. Pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) were performed for all mcr-1 positive isolates. S1-PFGE and Southern hybridisation were performed for localisation of mcr-1 and blaNDM. Results: Nineteen colistin resistant K. pneumoniae harboured mcr-1 and 3 had mgrB disruption. All isolates harboured blaOXA-48-type and ESBL genes; eight strains (five with mcr-1 and three with mgrB disruption) co-harboured blaNDM. Efflux pumps genes AcrAB and mdtK were detected in all 22 and tol-C in 21 isolates. Virulence-related genes entB and irp-1 were detected in all 22, mrkD in 20, and fimH-1 in 18 isolates; 11 isolates were strong biofilm producers. PFGE clustered mcr-1 positive isolates into eight groups based on ≥90% similarity; MLST revealed diverse sequence types, predominant being ST-15 (n = 4) and ST-16 (n = 4). Both mcr-1 and blaNDM were localised on plasmid and chromosome; mcr-1 was present on IncFII type and blaNDM on IncFIB and IncA/C type plasmids. Conclusions: Colistin resistance in K. pneumoniae was predominantly mediated by mcr-1. Co-existence of colistin, carbapenem, and other drug-resistant genes along with efflux pumps indicates towards enormous genomic plasticity in K. pneumoniae with ability to emerge as super-spreader of drug-resistance.

First report of peritonitis caused by the vancomycin-resistant coccus Pediococcus pentosaceus in a patient on continuous ambulatory peritoneal dialysis.

INTRODUCTION: Worldwide, about one-tenth of end-stage renal disease (ESRD) patients are on peritoneal dialysis (PD). Peritonitis is a major cause of PD failure and change of therapy to haemodialysis. An update on peritoneal dialysis-related infections has recommended the use of a first generation cephalosporin or vancomycin as an empirical therapy for Gram-positive organisms. Pediococcus spp. is a Gram-positive environmental cocci that have been increasingly reported from various nosocomial infections but very rarely from peritoneal dialysis infections. It is intrinsically resistant to Vancomycin but sensitive to ampicillin. So, diagnosis of this bacteria is important if isolated from PD infections. CASE PRESENTATION: An elderly female patient of ESRD on continuous ambulatory peritoneal dialysis (CAPD) was admitted with complaints of high fever and cloudy PD effluent for 2 days. She was started with vancomycin and imipenem empirically but did not improve even after 4 days. Pus cells were seen when PD fluid was examined microscopically. BACTEC culture of PD fluid isolated growth of Gram-positive cocci, which was confirmed as Pediococcus pentosaceus . It was resistant to vancomycin. The antibiotic of the patient was changed to ciprofloxacin IV. The patient responded in 2 days and was discharged after 7 days. CONCLUSION: This is the first case report of Pediococcus pentosaceus peritonitis in an ESRD patient on CAPD. Accurate diagnosis and antibiotic sensitivity test of the bacteria is important especially if isolated in critical patients as it is intrinsically resistant to vancomycin.

Expression levels of A disintegrin and metalloproteases (ADAMs), and Th17-related cytokines and their association with Helicobacter pylori infection in patients with gastroduodenal diseases.

Expression levels of A disintegrin and metalloproteases (ADAMs) (10 and 17) and Th17-related cytokines [interleukin (IL) 17A, IL-17F, IL-33, IL-23, IL-23R] were investigated by quantitative real time polymerase chain reaction in gastric biopsies of patients with different gastroduodenal pathologies in the presence and absence of Helicobacter pylori infection. Patients with gastric cancer (GC) (n = 70, intestinal-type 38 and diffuse type 32), peptic ulcer disease [n = 50, duodenal ulcer (DU) 16 and gastric ulcer (GU) 34] and functional dyspepsia (n = 120) were included in the study. Further, the expression levels of ADAMs and Th17 cytokines were correlated with H. pylori cytotoxin-associated genes pathogenicity island (cagPAI) status. Expression levels of ADAMs (10 and 17) and Th17-related cytokines (IL-17A, IL-23, IL-23R) were significantly higher in H. pylori-positive than in H. pylori-negative gastric biopsies. Significant increase in ADAM17 and Th17 cytokines (IL-17A and IL-23) expressions was observed in patients with GU and intestinal-type GC in the presence of H. pylori infection and in strains harbouring intact cagPAI. Expression levels of IL-17A, IL-23 and ADAM17 were strongly correlated with GU and intestinal-type GC and weakly with DU and diffuse-type GC in the presence of H. pylori infection. Higher expression levels of ADAM17 and Th17 cytokines (IL-17A and IL-23), and their strong correlation with GU and intestinal-type GC patients in the presence of H. pylori and its intact cagPAI status, suggest a possible role of strain specificity in the pathogenesis of these diseases.

The Era of Device Colonizers: Chryseobacterium indologenes Infections from a Tertiary Care Center in North India.

BACKGROUND: Chryseobacterium indologenes is a hospital environment contaminant and can cause healthcare-associated infections. METHODS: Patients with C. indologenes infections in a tertiary care center in North India for 6 months were evaluated for susceptibility patterns, comorbidities, mechanical devices, risk factors, and treatment outcomes. The organism was provisionally identified phenotypically, and identification was confirmed by the BD Phoenix automated microbiology system. Minimum inhibitory concentration values of antibiotic susceptibility were determined. RESULTS: A total of 12 isolates of C. indologenes were recovered from 11 patients. Five patients had C. indologenes bloodstream infection (BSI), one had ventilator-associated pneumonia (VAP), and one had both BSI and VAP. In four others, the organism was isolated from the catheterized urinary tract. All VAP and BSI patients were admitted to the Intensive Care Units and mechanically ventilated; all had central lines and history of colistin therapy during the past 15 days. The common underlying risk factors were diabetes, hypertension, and coronary artery disease. CONCLUSIONS: C. indologenes infections are increasing because of higher use of carbapenems and colistin, to which it is intrinsically resistant.

Clinical and laboratory standards institute versus European committee for antimicrobial susceptibility testing guidelines for interpretation of carbapenem antimicrobial susceptibility results for Escherichia coli in urinary tract infection (UTI).

BACKGROUND: Carbapenems show excellent activity against resistant uropathogens, and they are the antibiotics of choice for urinary tract infections (UTIs). The choice of carbapenem prescription is strongly influenced by antimicrobial susceptibility testing (AST) report. With the publication of recent AST guidelines by the European Committee on AST (EUCAST), we were curious to evaluate the difference in results between Clinical and Laboratory Standards Institute (CLSI) and the EUCAST guidelines for the interpretation of carbapenems. METHODS: During a period of 1 year, midstream urine specimens received in the laboratory were cultured by conventional techniques and 2932 of them grew significant colony counts of Escherichia coli. Out of them, 501 E. coli isolates which were resistant to at least six first-line antibiotics were further subjected to second-line antimicrobials imipenem and meropenem, reported by E-tests (bioMerieux, France). The E-test results were interpreted by both CLSI 2016 and EUCAST 6.0 (2016) guidelines. Weighted kappa was used to determine absolute agreement, and McNemar's Chi-square test was used to test the difference in proportions of susceptibility between two methods, respectively. RESULTS: Taking CLSI guidelines as a gold standard, there was 100% sensitivity in a susceptible category by the EUCAST guidelines for both the carbapenems. Weighted kappa showed good and moderate agreement between them for imipenem and meropenem, respectively. However, McNemar Chi-square test in the nonsusceptible category between the two tests was 9.38% and 33.03% for imipenem and meropenem, respectively, and they were highly significant (P < 0.001). CONCLUSIONS: A laboratory can follow EUCAST guidelines as well and the guidelines are more useful in urinary concentrated antibiotics such as carbapenems. Further other antibiotics need to be evaluated by both these guidelines.

Novel variant NDM-11 and other NDM-1 variants in multidrug-resistant Escherichia coli from South India.

OBJECTIVES: Carbapenem resistance mediated by New Delhi metallo-ß-lactamase 1 (NDM-1) and its variants has caused a major public-health concern worldwide. Here we report for the first time an Escherichia coli isolate positive for a novel variant (NDM-11). METHODS: blaNDM genes were investigated in E. coli by PCR and sequencing, and blaNDM variants were further characterised. The susceptibility pattern of novel blaNDM-11 towards different antimicrobials was compared with blaNDM-1 by cloning and expression in E. coli TOP10. RESULTS: A total of 33 carbapenem-resistant E. coli isolates were screened by PCR for the presence of blaNDM, of which 15 (45.5%) were positive. Sequencing of the PCR products revealed 10 isolates with NDM-1 and 5 isolates with NDM variants (one each of NDM-4, NDM-8 and NDM-11 and two NDM-5). Other resistance genes, including blaTEM-1, blaCTX-M-15, blaVIM, plasmid-encoded AmpC blaCMY-2 and 16S methyltransferases (rmtB and rmtC), were also associated with NDM variants in different combinations. The blaNDM variants were located on a transferable IncF-type plasmid of >100kb. Pulsed-field gel electrophoresis (PFGE) showed that all five E. coli isolates were unrelated, and multilocus sequence typing (MLST) revealed that they all belonged to ST131. Expression of the blaNDM-1 and blaNDM-11 genes in E. coli TOP10 showed no significant difference in MICs to various ß-lactams, including carbapenems. CONCLUSIONS: This study underlines the spread of NDM variants with other antimicrobial resistance genes in E. coli in South India. It also describes a novel NDM variant (blaNDM-11) having an antimicrobial resistance pattern similar to blaNDM-1.

Isolation of bacterial DNA followed by sequencing and differing cytokine response in peritoneal dialysis effluent help in identifying bacteria in culture negative peritonitis.

AIM: The treatment of peritoneal dialysis related culture negative peritonitis is empirical which increases the cost of therapy and moreover antibiotic resistance. We aimed the study to isolate bacterial DNA from PD effluent and indentify bacteria causing peritonitis in culture negative situations. We have also studied the cytokine response with different bacteria causing peritonitis. METHODS: We have isolated bacterial DNA from PD effluent of culture negative and culture positive peritonitis patients. Bacterial DNA was subjected to polymerase chain reaction using universal bacteria specific primers and subsequently to Gram type specific primers for the differentiation of the etiologic agents into Gram-positive and Gram-negative. The amplified products were sequenced and subjected to blast search to identify agent at genus/ species level. RESULTS: Of the 30 molecular method positive samples, 16 (53.33%) samples were positive for Gram-negative bacteria and 4 (13.33%) for Gram-positive, while the remaining10 (33.33%) were positive for both Gram-positive and Gram-negative bacteria. We have found organisms that usually do not grow on normal culture methods. TNF-α was significantly associated with Gram-positive peritonitis and regulatory cytokine IL-10 with Gram-negative peritonitis. CONCLUSIONS: The molecular techniques are helpful in detecting and identifying organisms from culture negative PD effluent.

Prevalence and Molecular Characterization of New Delhi Metallo-Beta-Lactamases in Multidrug-Resistant Pseudomonas aeruginosa and Acinetobacter baumannii from India.

New Delhi metallo-beta-lactamase (NDM)-mediated carbapenem resistance in Pseudomonas aeruginosa and Acinetobacter baumannii is a major concern. We investigated the presence of NDM and its variants in P. aeruginosa and A. baumannii at a tertiary hospital in North India. A total of 236 isolates (130 P. aeruginosa and 106 A. baumannii) were included; 38 (29.23%) P. aeruginosa and 20 A. baumannii isolates (18.8%) were resistant to carbapenems and all of them were blaNDM positive. All 38 carbapenem-resistant P. aeruginosa harbored blaNDM-1, while 12 (60%) of 20 A. baumannii harbored blaNDM-2. Pulsed-field gel electrophoresis showed that all 58 isolates were clonally unrelated. By Southern blot analysis, blaNDM-2 was located on chromosome. The blaNDM-2-positive isolates were more frequently recovered from tracheal aspirate (67% vs.16%; p = 0.02) and intensive care unit (67% vs. 20%; p = 0.001) than blaNDM-1. Among other carbapenemases, VIM was significantly associated with blaNDM-1 than blaNDM-2 (61% vs. 17%; p = 0.006). Mortality between blaNDM-1- and blaNDM-2-infected patients was comparable. When expressed in Escherichia coli, blaNDM-2 transformant conferred one doubling dilution higher MIC value for cefotaxime, piperacillin/tazobactam than blaNDM-1. The study shows the emergence of blaNDM-mediated resistance among P. aeruginosa and A. baumannii and rapid evolution of blaNDM-2 in A. baumannii with its chromosomal localization.

Simultaneous isolation of Chryseobacterium gleum from bloodstream and respiratory tract: first case report from India.

Introduction. Species of the genus Chryseobacterium are emerging healthcare-associated pathogens, often colonizing the hospital environment. There are no clear guidelines available for antimicrobial susceptibility of this organism. In this report we present the first case, to our knowledge, of simultaneous central-line-associated bloodstream infection (CLABSI) and ventilator-associated pneumonia (VAP) due to Chryseobacterium gleum from India. Case presentation. A 62 years old man with a history of a road traffic accident 1 month previously was referred to our center for further management. He developed features of sepsis and aspiration pneumonia on day 3 of admission. Four blood cultures (two each from central and peripheral lines) and two tracheal aspirate cultures grew pure yellow colonies of bacteria. Both matrix assisted laser desorption ionization time of flight mass spectrometry, (MALDI-TOF MS; bioMérieux, Marcy-L'Etoile, France,) and BD Phoenix (BD Biosciences, Maryland, USA) identified the organism as C. gleum. However, BD Phoenix failed to provide MIC breakpoints. The isolates of C. gleum both from blood and tracheal aspirate showed identical susceptibility patterns: resistant to cephalosporins and carbapenems and susceptible to ciprofloxacin, levofloxacin, amikacin, trimethoprim+sulfamethoxazole, piperacillin-tazobactam, cefoperazone-sulbactam, doxycycline, minocycline and vancomycin. Following levofloxacin therapy, the fever responded within 48 h and procalcitonin levels decreased without removal of the central line or endotracheal tube. However, the patient developed sudden cardiac arrest on day 10 of treatment and could not be resuscitated. Conclusion. Rapid and accurate identification of C. gleum in the laboratory, preferably based on MALDI-TOF, is essential for guiding therapy. C. gleum responds well to fluoroquinolones without the need to remove indwelling catheters.

Assessment of Doripenem, Meropenem, and Imipenem against Respiratory Isolates of Pseudomonas aeroginosa in a Tertiary Care Hospital of North India.

OBJECTIVE: Pseudomonas aeruginosa is one of the leading pathogen causing healthcare-associated infections, particularly in immunocompromised and critically ill patients. The development of carbapenem resistance in P. aeruginosa infections is worrisome. Data specifically comparing the susceptibility of the three available carbapenems are lacking in the Indian subcontinent. MATERIALS AND METHODS: We evaluated the minimum inhibitory concentrations (MICs) of the three commonly used carbapenems- imipenem, meropenem, and doripenem against, 435 P. aeruginosa isolates obtained from respiratory samples and compared their susceptibility patterns to determine the best possible carbapenem among those available that may be used in combination regimes. RESULTS: Overall, 222 (51.0%) of isolates were susceptible to doripenem followed by imipenem 206 (47.3%) and meropenem 195 (44.8%), respectively. Two hundred and sixty-two (60.23%) strains were intermediate or resistant to at least one carbapenem. The MIC90 of all three carbapenems was >32 µg/ml while the MIC50 of meropenem was 16 µg/ml which was higher than MIC50 of both imipenem (4 µg/ml) and doripenem (2 µg/ml). CONCLUSION: Our study revealed that doripenem exerted better in vitro activity against the tested bacteria compared to imipenem and meropenem, but the difference was not statistically significant.

Survivability and fitness cost of heterogeneous vancomycin-intermediate Staphylococcus aureus.

The aim of this study was to observe the survivability and fitness cost of heterogeneous vancomycin-intermediate Staphylococcus aureus(hVISA) isolates. Survivability study was performed on dry cotton swab, and fitness cost was evaluated by estimating growth kinetics and generation time constant in BACTEC automated system. Total mean maximum time of recovery on primary culture was 4.1 and 7.1 weeks (P = 0.0001) for hVISA and vancomycin-sensitive S. aureus (VSSA), respectively, in dry starved condition. No significant difference between the mean value of lag phase duration (P = 0.89) was noted between hVISA and VSSA isolate in growth kinetics. However, we observed lesser generation time of hVISA isolates compared to S. aureus ATCC 29213 (P = 0.0076). This study concluded that a significant difference in generation time between VSSA and hVISA and suggests that hVISA have fitness cost compared to VSSA. However, further studies with more cases are required.

Matrix metalloproteinases-2 and -9 in Campylobacter jejuni-induced paralytic neuropathy resembling Guillain-Barré syndrome in chickens.

Inflammation in Guillain-Barré syndrome (GBS) is manifested by changes in matrix metalloproteinase (MMP) and pro-inflammatory cytokine expression. We investigated the expression of MMP-2, -9 and TNF-α and correlated it with pathological changes in sciatic nerve tissue from Campylobacter jejuni-induced chicken model for GBS. Campylobacter jejuni and placebo were fed to chickens and assessed for disease symptoms. Sciatic nerves were examined by histopathology and immunohistochemistry. Expressions of MMPs and TNF-α, were determined by real-time PCR, and activities of MMPs by zymography. Diarrhea developed in 73.3% chickens after infection and 60.0% of them developed GBS like neuropathy. Pathology in sciatic nerves showed perinodal and/or patchy demyelination, perivascular focal lymphocytic infiltration and myelin swelling on 10th- 20th post infection day (PID). MMP-2, -9 and TNF-α were up-regulated in progressive phase of the disease. Enhanced MMP-2, -9 and TNF-α production in progressive phase correlated with sciatic nerve pathology in C. jejuni-induced GBS chicken model.

High frequency of SCCmec type V and agr type I among heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) in north India.

OBJECTIVES: The objective of this study was to compare the genetic features of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) and vancomycin-sensitive methicillin-resistant S. aureus (VS-MRSA) isolates. METHODS: The presence of staphylococcal cassette chromosome mec (SCCmec) types, Panton-Valentine leukocidin (PVL), accessory gene regulator (agr) types, and vanA and vanB genes in hVISA and VS-MRSA isolates was evaluated by PCR. Genetic relatedness was studied by pulsed-field gel electrophoresis (PFGE). RESULTS: The distribution of SCCmec types in hVISA was as follows: 13/29 (44.8%) each of types II and V, 1/29 (3.4%) type III and 2/29 (6.9%) type IVa. Among VS-MRSA isolates, 20/50 (40.0%) were SCCmec type II, 17/50 (34.0%) were type III, 3/50 (6.0%) were type IVa and 10/50 (20.0%) were type V. SCCmec type V was significantly associated with hVISA, whereas SCCmec type III showed an association with VS-MRSA (P=0.020 and P=0.001, respectively). The PVL gene was detected in 9/29 hVISA (31.0%) and 13/50 VS-MRSA (26.0%). By PFGE analyses, both hVISA and VS-MRSA strains were found to be clonally unrelated. In hVISA isolates, 24/29 (82.8%) were agr type I, 3/29 (10.3%) were type III and 2/29 (6.9%) were non-typeable. However, in VS-MRSA isolates, 25/50 (50.0%) were type II, 15/50 (30.0%) were type I, 7/50 (14.0%) were type III and 3/50 (6.0%) were non-typeable. CONCLUSIONS: The study shows that healthcare-associated MRSA strains may harbour community-acquired MRSA genetic markers. The changing molecular epidemiology and role of agr I in reduced vancomycin susceptibility in hVISA requires further investigation.

Microbiology, clinical spectrum and outcome of peritonitis in patients undergoing peritoneal dialysis in India: Results from a multicentric, observational study.

BACKGROUND: Peritoneal dialysis (PD)-related peritonitis is a major risk factor for drop out of patients on continuous ambulatory PD (CAPD) and automated PD (APD). Factors affecting PD-related peritonitis and centre-specific microbiological data are lacking in India. A multicentric prospective observational study was designed to overcome the gaps in the existing data regarding causative organism and outcome. METHODOLOGY: The present study was a prospective, uncontrolled, open-label; observational study conducted in 21 centres representing all the four geographical regions (North, South, East and West) of India between April 2010 and December 2011. RESULTS: A total of 244 patients on chronic PD with peritonitis were enrolled in the study (CAPD and APD), who met the inclusion criteria, from 21 centres covering the different geographical areas of India. Amongst the 85 samples that were culture positive, 38 (44.7%) were in the monsoon season followed by 23 (27.1%) in the post-monsoon, 18 (21.2%) during winter and 11 (12.9%) during summer. Maximum culture positivity (72.7%) was observed with automated culture technique. Microorganisms could be isolated in only 85 cases (35.3%) while the remaining samples were culture negative (156/241, 64.7% of samples). Organisms isolated were Gram-negative in 47.8%, Gram-positive in 36.7%, fungal in 13.3% and Mycobacterium tuberculosis in 2.2%. CONCLUSION: This large multicentre study of peritonitis offers insights into the aetiology and outcomes of infectious complications of chronic PD in India that are germane to clinical decision-making.

Antimicrobial susceptibility to azithromycin among Salmonella enterica Typhi and Paratyphi A isolates from India.

Decreased ciprofloxacin susceptibility (DCS) and multidrug resistance in typhoidal Salmonella isolates in areas of endemicity are significant therapeutic problems. Guidelines for azithromycin disc diffusion and MIC interpretive criteria for Salmonella enterica serovar Typhi were published recently by the Clinical and Laboratory Standards Institute in 2015. We investigated the antimicrobial susceptibility pattern of azithromycin in 100 isolates of Salmonella Typhi (n=80), Paratyphi A (n=18) and B (n=2) recovered from bloodstream infections from January 2013 to December 2015. Zone sizes were extrapolated against MIC values, and a scatter plot was constructed. The azithromycin MICs by Etest ranged from 2 to 16 µg ml-1, while the disc diffusion diameters were from 13 to 22 mm. We observed that the margin of the zone of inhibition around the azithromycin disc may not be very clear and therefore difficult to interpret and that there was wide variation in the zone sizes for the same MIC value in both serovars. DCS was observed in 85 % of Salmonella Typhi recovered (68/80) and in 15/18 (83.3 %) Paratyphi A isolates. Judicious use of azithromycin is advocated as an alternative oral agent in endemic areas where DCS is common.

Antimicrobial susceptibility pattern and sequence analysis of DNA gyrase and DNA topoisomerase IV in Salmonella enterica serovars Typhi and Paratyphi A isolates with decreased susceptibility to ciprofloxacin.

BACKGROUND: We describe the antimicrobial susceptibility pattern of 100 typhoidal Salmonella isolates recovered from blood cultures and also investigate the association of decreased ciprofloxacin susceptibility with mutations in the genes coding for DNA gyrase and topoisomerase IV in 55 isolates. METHODS: The study was conducted between January 2013 and December 2015 at a tertiary care centre in north India. Antimicrobial susceptibility testing was performed by Kirby-Bauer disc diffusion and E-test. Genotypic characterization included the screening of mutations in the quinolone resistance-determining region of gyrA, gyrB, parC, and parE by PCR. DNA sequence analysis was done for 55 isolates. RESULTS: Out of 100 isolates recovered 80 were S. Typhi, 18 were Paratyphi A and two were Paratyphi B. Eighty two percent (66/80) of S. Typhi and 15/18 S. Paratyphi A showed decreased ciprofloxacin susceptibility. The most common mutation in gyrA led to a change at codon 83 of serine to phenylalanine (n=37) or tyrosine (n=12). Five S. Typhi isolates that were resistant to ciprofloxacin (MICs of 12, 16, 24 and 32 µg/ml) had a second mutation at codon 87 in the gyrA gene changing aspartate to asparagine. CONCLUSIONS: There is a need to urgently review the use of fluoroquinolones for the management of enteric fever in endemic areas.