Clonal diversity of New Delhi metallobetalactamase-1 producing Enterobacteriaceae in a tertiary care centre.

PURPOSE: New Delhi metallobetalactamase-1 (NDM-1) production is a major mechanism of resistance to carbapenems among the Enterobacteriaceae and is a cause for concern in the field of microbial drug resistance. This study was performed to detect NDM-1 in Enterobacteriaceae and to determine the clonal relatedness of NDM-1 producing Escherichia coli and Klebsiella pneumoniae isolated from patients admitted in a tertiary care centre. MATERIALS AND METHODS: A total of 111 clinically significant Enterobacteriaceae isolates, resistant to cephalosporin subclass III were screened for carbapenemase production by the modified Hodge test. Minimum inhibitory concentration to imipenem and meropenem was determined and interpreted according to Clinical Laboratory Standards Institute 2011 criteria. Presence of bla NDM-1 was detected by polymerase chain reaction. To ascertain clonal relatedness, random amplification of polymorphic deoxyribonucleic acid (RAPD) was carried out for representative NDM-1 producers. RESULTS: bla NDM-1 was detected in 64 study isolates, of which 27 were susceptible to carbapenems. RAPD revealed a high degree of clonal diversity among NDM-1 producers except for a small clustering of isolates in the neonatal intensive care unit. CONCLUSION: There is extensive clonal diversity among the NDM-1 producing E. coli and K. pneumoniae. Hence, antibiotic selection pressure rather than horizontal transfer is probably an important operating factor for the emergence of NDM-1. This calls for increased vigilance, continuous surveillance and strict enforcement of antibiotic policy with restricted use of inducer drugs.

Detection of Amp C genes encoding for beta-lactamases in Escherichia coli and Klebsiella pneumoniae.

PURPOSE: Amp C beta-lactamase are Ambler class C enzymes that confer resistance to extended spectrum cephalosporins and are not inhibited by beta-lactamase inhibitors. Their detection is crucial, since the phenotypic tests are not standardised leading to ambiguity in interpretation of results. This study was done to detect the types of Amp C prevalent in Escherichia coli and Klebsiella pneumoniae by multiplex polymerase chain reaction (PCR). MATERIALS AND METHODS: Seventy-seven consecutive cefoxitin resistant clinical isolates of E. coli (n = 25) and K. pneumoniae (n = 52) were included in the study. Antibiotic susceptibility testing to various classes of antibiotics was performed by disc diffusion using Clinical Laboratory Standards Institute (CLSI) guidelines. Minimum inhibitory concentration (MIC) to cefoxitin, imipenem and meropenem were determined by broth microdilution method. Isolates were screened for production of Extended Spectrum Beta-Lactamase (ESBL). Multiplex PCR was performed for the detection of Amp C genes after phenotypic testing (Hodge test and inhibitor based test). RESULTS: Cefoxitin Hodge test was positive in 40 isolates which included 20 E. coli and 20 K. pneumoniae. There was zone enhancement with boronic acid in 55 isolates, of which 36 were K. pneumoniae and 19 were E. coli. Multiplex PCR detected Amp C in 11/25 E. coli and 12/52 K. pneumoniae isolates. The Amp C genes detected were CIT (Amp C origin - Citrobacter freundii), DHA (Dhahran Hospital, Saudi Arabia), ACC (Ambler class C), EBC (Amp C origin - Enterobacter cloacae) groups. ESBL was co-produced in 54 isolates. CONCLUSIONS: Amp C was detected in 29.87% of the study isolates. Majority of them co-produced ESBL. The most common Amp C was the CIT family. Screen tests for cefoxitin resistance may be falsely positive due to production of carbapenamases.

OXA beta-lactamase-mediated carbapenem resistance in Acinetobacter baumannii.

OBJECTIVES: Acinetobacter baumannii is a significant pathogen in health care settings. In recent years, an increase in carbapenem resistance among A. baumannii due to Ambler class B metallo-beta-lactamases or class D OXA carbapenamases has been reported. In this study we detected the presence of OXA carbapenamases and coproduction of metallo-beta-lactamases (blaVIM and blaIMP ) by phenotypic and genotypic methods in carbapenem resistant clinical isolates of Acinetobacter baumannii. MATERIALS AND METHODS: A total of 116 consecutive, non-duplicate carbapenem resistant A. baumannii isolated from various clinical specimens were included in the study. The modified Hodge test and inhibitor potentiated disk diffusion tests were done for the screening of carbapenamase and metallo-beta-lactamase production, respectively. Polymerase chain reaction (PCR) was performed for the detection of OXA (blaOXA 23 like, blaOXA 24 like, blaOXA-51 like and blaOXA-58 like genes) and metallo-beta-lactamases (blaVIM and blaIMP ) genes. Gene sequencing was performed for representative isolates. RESULTS: Among 116 A. baumannii, OXA genes were detected in 106 isolates. BlaOXA 51 like (n = 99) and blaOXA -23 like (n = 95) were the most common and they coexisted in 89 isolates. blaOXA-24 like gene was detected in two isolates of which one also carried blaOXA-51 like and blaOXA-58 like genes. The modified Hodge test was positive in 113 isolates. The metallo-beta-lactamase screening test was positive in 92 isolates. blavim was detected in 54 isolates of which 1 also carried the blaIMP gene. CONCLUSIONS: blaOXA-23 like and bla OXA 51 like genes are the most common types of OXA carbapenamases while the blaVIM type is the most common type of metallo-beta-lactamase contributing to carbapenem resistance in clinical isolates of A. baumannii. The coproduction of OXA and metallo-beta-lactamases is not an uncommon phenomenon in A. baumannii.

Incidence of ESBL producers amongst Gram-negative bacilli isolated from intra-abdominal infections across India (based on SMART study, 2007 data).

OBJECTIVES: This study was conducted in 9 centers spread over India from January 1 to December 31, 2007 to monitor in vitro susceptibility of Gram-negative bacilli to Group I carbapenem, ertapenem and other antimicrobials in intra-abdominal infections and to identify early changes in susceptibility pattern of community or hospital acquired organisms, with a focus on ESBL producers. MATERIAL AND METHODS: Gram-negative bacilli isolated from intra-abdominal samples of patients with documented intra-abdominal infections were processed for identification by conventional/ automated methods and antimicrobial susceptibility by Micro-Scan (Siemens) MIC panel against 12 antimicrobials (3rd and 4th generation cephalosporins, Groups I and II carbapenems, amikacin, levofloxacin, amoxicillin-clavulanic acid and piperacillin-tazobactam). RESULTS: A total of 588 isolates were identified, of which 351 (60%) were E. coli and 114 (19%) were Klebsiella spp. 79% of E. coli and 70% of Klebsiella spp. were ESBL producers in general. 110 of E. coli and 35 of Klebsiella isolates were from community-acquired intra-abdominal infections. 80% of E. coli and 63% of Klebsiella isolates from community-acquired infections were ESBL producers, against 79% of E. coli and 73% of Klebsiella isolates from hospital-acquired infections. Amongst the ESBL-positive isolates of E. coli, 94% were susceptible in vitro to ertapenem, 96% to imipenem and 76% to piperacillin-tazobactam. For ESBL-positive isolates of Klebsiella spp., the corresponding figures were 80%, 94% and 59% respectively. CONCLUSION: The study showed a high incidence of ESBL-producers amongst Enterobacteriaceae isolates from intra-abdominal infections in both community-acquired and hospital-acquired settings across India. Ertapenem was comparable with imipenem against ESBL-positive E. coli isolates, while imipenem was more effective than ertapenem against ESBL-positive Klebsiella isolates.

Tuberculous peritonitis in a cohort of continuous ambulatory peritoneal dialysis patients.

Among 155 patients who were initiated on continuous ambulatory peritoneal dialysis (CAPD), 4 patients (2 men, 2 women) developed tuberculous peritonitis. They had been on PD for between 2 months and 84 months when they developed the peritonitis. The Mantoux test was negative in all of them. The diagnosis was made by a variety of means in the various cases: demonstration of Mycobacterium tuberculosis in the peritoneal cavity; presence of caseating granuloma in a peritoneal biopsy; Mycobacterium tuberculosis in a cold abscess adjacent to the peritoneal cavity; and demonstration of IS6110 and MPB64 genes of Mycobacterium tuberculosis by polymerase chain reaction (PCR) technique. Two of the patients developed ultrafiltration failure. Among 3 patients who were switched to hemodialysis, 2 died and 1 continues on maintenance dialysis. The last patient, whose catheter was removed, was reimplanted with a new catheter and continues on PD without ultrafiltration failure. Any patient with peritonitis unresponsive to conventional therapy should be investigated for tuberculous peritonitis. Institution of chemotherapy without delay will preserve peritoneal membrane integrity.

Renal aspergillosis giving rise to obstructive uropathy and recurrent anuric renal failure.

A sixty-year-old previously healthy male patient presented with anuric renal failure of sudden onset. He was detected to have Aspergillus fumigatus fungal balls in the renal pelvis, ureters and bladder which were removed and his renal function improved. He was treated with itraconazole and sent home. Three weeks later he again presented with anuria and renal failure. He had recurrence of the obstruction with the same fungus. The fungal ball was removed, a double 'J' stenting was performed and he was treated with amphotericin B and itraconazole. Hence we report a previously healthy patient with no evidence of immunosuppression presenting an obstructive anuric renal failure due to isolated renal aspergillosis.