Naveen Grover

First Report of blaNDM-1 in Raoultella ornithinolytica

The genus Raoultella is composed of Gram-negative, aerobic, oxidase-negative, catalase-positive, nonmotile, capsulated rods belonging to the Enterobacteriaceae family ([1][1]). Raoultella ornithinolytica is found in aquatic and hospital environments ([2][2]). Occurrence of Raoultella in human

Reverse transcription loop-mediated isothermal amplification (RT-LAMP) for diagnosis of dengue.

BACKGROUND Dengue is an emerging public health problem causing serious morbidity and mortality in tropical developing countries. Early, sensitive and specific diagnosis is paramount for clinical decision making. Currently available diagnostic tests are limited in scope and utility. This study highlights applicability of RT-LAMP in dengue diagnosis. METHODS 100 dengue confirmed cases, 100 dengue negative cases and 79 healthy negative controls from dengue epidemic between Sep 2009 to Jul 2011 were included. Dengue cases were profiled using WHO guidelines 2006, haematological and biochemical parameters evaluated and diagnosed using NS1 antigen, IgM and IgG enzyme immunoassay, RT-PCR and RT-LAMP. Positive cases were serotyped, genotyped and various tests were compared. RESULTS Mean haematocrit, PT, PTT, platelet count, activated lymphocytes, serum fibrinogen, transaminases, bilirubin, lactate dehydrogenase, protein and sodium were significantly elevated in DHF/DSS as compared to DF. NS1 antigen, RT-PCR and RT-LAMP were sensitive during 1-3 days while μ-capture IgM EIA was specific after 5-7 days of initial infection. DEN-1 genotype III was predominant. CONCLUSION Deranged haematocrit and liver function tests are indicators of the severity of the disease. RT-LAMP is rapid, cost effective, highly sensitive and specific qualitative and quantitative technique which can detect dengue infection in both early and intermediary stages when NS1 antigen titres are not in the detectable range and the IgM antibody titres have just started to rise. Its superiority over existing techniques, amenability for automation and promising utility in low resource healthcare setups and field conditions raise it as the new gold standard for dengue diagnosis.

Emergence of Escherichia coli, Co-Producing NDM-1 and OXA-48 Carbapenemases, in Urinary Isolates, at a Tertiary Care Centre at Central India.

OBJECTIVE To detect genes encoding carbapenem resistance in urinary isolates of Escherichia coli recovered from hospitalized patients in tertiary care centre in Pune, India. METHODS From Jan 2012 to Dec 2012, a total of 300 consecutive non-duplicate (one isolate per patient) clinical isolates of Escherichia coli were recovered from urine cultures of hospitalized patients including hospital acquired infection cases admitted to the medical and surgical intensive care units. Polymerase chain reaction (PCR) assays and sequencing was used to determine the presence of beta-lactamase encoding genes. Conjugation experiments were performed to determine the transferability of beta-lactamase. RESULTS All the isolates were completely resistant to the second and third generation cephalosporins tested as well as carbapenems. All the isolates showed 100% susceptibility to tigecycline and colistin in vitro. Conjugation experiments demonstrated that blaNDM-1 was transferable via plasmid. All the isolates showed presence of blaNDM-1 and co-association of blaOXA-48 was 25/45(55%) of the isolates. Repetitive element based PCR (REP PCR), Enterobacterial Repetitive Intergenic Consensus (ERIC PCR) and Randomly Amplified Polymorphic DNA (RAPD) revealed a diversity of six clonal types among E.coli isolates. CONCLUSION Co-production of NDM-1with OXA-48 in urinary isolates of E. coli was detected for the first time in India. Transmission of plasmid carrying these resistant genes to other members of Enterobacteriaceae will increase incidence of multidrug resistance. Early detection of these genes will help in prevention and adequate infection control by limiting the spread of these organisms.

Emergence of NDM – 1 in the Clinical Isolates of Pseudomonas aeruginosa in India

OBJECTIVE The present study was undertaken to detect the prevalence of the blaNDM-1 metallo beta lactamases (MBLs) in the isolates of Pseudomonas aeruginosa, which were recovered from various clinical samples from hospitalized patients in a tertiary care centre in Pune, India. METHODS A total of 200 isolates of P. aeruginosa which were obtained from various clinical samples were subjected to antibiotic susceptibility testing by the disc-diffusion method and their MICs were determined by the Vitek - 2 Automated Antimicrobial Identification and Susceptibility Testing System against imipenem, meropenem, ticarcillin, amikacin, gentamicin, tobramycin, ciprofloxacin, levofloxacin, moxifloxacin, tigecycline, trimethoprim/sulfamethoxazole, ampicillin/sulbactam, piperacillin/tazobactam, cefoperazone/sulbactam, cefepime, tetracycline, ceftazidime, ceftriaxone and colistin. Their MICs were also determined by the Etest method against imipenem, meropenem, piperacillin, tobramycin, ceftazidime, tigecycline and colistin. The presence of blaNDM-1 was detected by PCR and it was confirmed by sequencing the gene which was present in the isolates which exhibited carbapenem resistance. The experimental transferability of the plasmids which carried blaNDM-1 was determined by using E. coli J53 as the recipient. RESULT In the present study, four isolates of P. aeruginosa, which carried the blaNDM-1 gene, were resistant to imipenem and meropenem. These blaNDM-1 carrying isolates remained susceptible to colistin. The plasmid carrying blaNDM-1 was successfully transferred from the four isolates to E. coli J53 recipients. CONCLUSIONS We are reporting the emergence of the P. aeruginosa carrying NDM-1gene, which exhibited resistance to imipenem and meropenem, for the first time from India.

Therapeutic challenges of ESBLS and AmpC beta-lactamase producers in a tertiary care center.

BACKGROUND Resistance to broad-spectrum beta lactams mediated by extended spectrum beta lactamases (ESBLs) and AmpC beta lactamases (AmpC βLs) enzymes is an increasing problem worldwide. Determination of their prevalence is essential to formulate an effective antibiotic policy and hospital infection control measures. Present study was undertaken to determine the prevalence of ESBL and AmpC βL producers in ICU of a tertiary care center. METHODS A total of 262 clinical isolates comprising of Escherichia coli, Klebsiella pneumoniae and Proteus mirabilis that were recovered from various clinical specimens over a one year period, were studied. Antibiogram profile was determined to conventionally used antibiotics, along with recommended tests for detection of ESBL and AmpC βL production. RESULTS 40.07% (105/262) were found to be ESBL producers, 14.8% (39/262) were AmpC bL producers. The coexistence of ESBL and AmpC βL producers was detected in 9.9% (26/262) of the isolates. CONCLUSION Screening of multidrug resistant bacteria especially belonging to the Enterobacteriaceae poses considerable therapeutic challenges in critical care patients because of the production of ESBL and AmpC βL. Strategies to keep a check on the emergence of such drug resistant microbes by hospital environmental surveillance and laboratory monitoring should form an important aspect of Hospital Infection control policy guidelines.

Carbapenem Resistance among Enterobacter Species in a Tertiary Care Hospital in Central India.

Objective. To detect genes encoding carbapenem resistance among Enterobacter species in a tertiary care hospital in central India. Methods. Bacterial identification of Enterobacter spp. isolates from various clinical specimens in patients admitted to intensive care units was performed by routine conventional microbial culture and biochemical tests using standard recommended techniques. Antibiotic sensitivity test was performed by standard Kirby Bauer disc diffusion technique. PCR amplification and automated sequencing was carried out. Transfer of resistance genes was determined by conjugation. Results. A total of 70/130 (53.84%) isolates of Enterobacter spp. were found to exhibit reduced susceptibility to imipenem (diameter of zones of inhibition ≤13 mm) by disc diffusion method. Among 70 isolates tested, 48 (68.57%) isolates showed MIC values for imipenem and meropenem ranging from 32 to 64 mg/L as per CLSI breakpoints. All of these 70 isolates were found susceptible to colistin in vitro as per MIC breakpoints (<0.5 mg/L). PCR carried out on these 48 MBL (IP/IPI) E-test positive isolates (12 Enterobacter aerogenes, 31 Enterobacter cloacae, and 05 Enterobacter cloacae complex) was validated by sequencing for beta-lactam resistance genes and result was interpreted accordingly. Conclusion. The study showed MBL production as an important mechanism in carbapenem resistance in Enterobacter spp. and interspecies transfer of these genes through plasmids suggesting early detection by molecular methods.

Multidrug resistant NDM-1 metallo-beta-lactamase producing Klebsiella pneumoniae sepsis outbreak in a neonatal intensive care unit in a tertiary care center at central India

OBJECTIVE The objective of the following study is to detect genes encoding carbapenem resistance in Klebsiella pneumoniae sepsis outbreak in a neonatal intensive care unit (NICU). MATERIALS AND METHODS Antibiotic sensitivity test was performed by standard Kirby Bauer disc diffusion technique and minimum inhibitory concentrations of antibiotics was determined by VITEK-2. Polymerase chain reaction (PCR) assays and sequencing was used to determine the presence of beta-lactamase encoding genes. Conjugation experiments were performed to determine the transferability of beta-lactamase. Isolate relatedness were determined by repetitive-element PCR (REP), enterobacterial repetitive intergenic consensus (ERIC) PCR and random amplified polymorphic deoxyribonucleic acid (RAPD). RESULTS All the isolates were completely resistant to the second and third generation cephalosporins tested as well as carbapenems. Susceptibility profiling of the isolates indicated that 100% retained susceptibility to tigecycline and colistin. Conjugation experiments indicated that blaNDM-1 was transferable and likely through a plasmid-mediated event. All the isolates showed the presence of blaNDM-1 with co association of blaCTX-M-15. REP-PCR, ERIC-PCR and RAPD revealed a single clonal type circulating in NICU environment. CONCLUSION Co-production of NDM-1 with CTX-M-15 in K. pneumoniae isolates was detected for the first time in our NICU. Transmission of plasmid carrying these resistant genes to other members of Enterobacteriaceae will increase the incidence of multidrug resistance. Early detection of these genes will help in prevention and adequate infection control by limiting the spread of these organisms.

First Report of an Enterobacter ludwigii Isolate Coharboring NDM-1 and OXA-48 Carbapenemases

Enterobacter spp. have emerged as pathogens responsible for hospital-acquired infections. Carbapenem resistance is increasingly being reported in this species, which is a matter of concern. Enterobacter spp. can produce abdominal, urinary tract, meningeal, and surgical site infections. In the

Species distribution and antimicrobial resistance pattern of Coagulase-negative Staphylococci at a tertiary care centre

BACKGROUND Coagulase-negative Staphylococci (CoNS), previously dismissed at contaminants, have now emerged as an important cause of nosocomial infections especially in patients with implants and prosthetic devices. They are a well-known cause of bloodstream infections, urinary tract infections, wound infections, prosthetic valve endocarditis and eye infections. This study was conducted with an aim to identify CoNS at the species level from various clinical samples and determine the antimicrobial resistance pattern of these isolates. METHODS This cross sectional study was carried out from September 2011 to February 2014 in which 150 non-repetitive clinical isolates of CoNS were identified at the species level by conventional phenotypic methods. Complete antimicrobial susceptibility profile was also determined by Kirby Bauer disc diffusion method. Susceptibility testing to vancomycin was done by E-test method. RESULTS Only three species of CoNS were isolated, the most common being Staphylococcusepidermidis (60%) followed by Staphylococcussaprophyticus (27.3%) and Staphylococcushemolyticus (12.7%). Most S. epidermidis were isolated from blood and intravascular catheter tip samples, whereas all S. saprophyticus were isolated from urine samples of female patients. All isolates were found to be resistant to penicillin, but were susceptible to glycopeptides and linezolid and showed variable resistance to fluoroquinolones, aminoglycosides and macrolides. CONCLUSION CoNS are emerging nosocomial pathogens and should not always be overlooked as contaminants. However, growth of CoNS from blood cultures and intravascular catheter tips should be clinically correlated and carefully interpreted. As many CoNS strains exhibit drug resistance, antimicrobial susceptibility profile should be determined prior to treatment of these infections.

Presence of a novel variant NDM-10, of the New Delhi metallo-beta-lactamase in a Klebsiella pneumoniae isolate.

This study showed that the prevalence of 3GC resistant Enterobacteraceae and non‐fermenting Gram‐negative Bacilli in patients admitted to ICUs of our hospital was substantial while the prevalence of NDM‐1 was substantially low (3.3%). GI colonisation by 3GC and carbapenem‐resistant pathogens increases the risk of clinical infections by these pathogens and subsequent human to human transmission.[3]