Detection of New Delhi metallo beta lactamase-1 (NDM-1) carbapenemase in Pseudomonas aeruginosa in a single centre in southern India.

Background & objectives: New Delhi metallo β-lactamase-1 (NDM-1) producing Pseudomonas aeruginosa isolates are potential threat to human health. This study was conducted to detect the presence of blaNDM-1 in carbapenem resistant P. aeruginosa in a tertiary care center in southern India. Methods: Sixty one carbapenem resistant clinical isolates of a total of 212 P. aeruginosa isolates cultured during the study period were screened for the presence of NDM-1by PCR. Clinical characteristics of the NDM-1 positive isolates were studied and outcome of the patients was followed up. Results: Of the 61 isolates, NDM-1 was detected in four isolates only. These were isolated from patients in the intensive care units and chest medicine ward. The source specimens were pus, sputum, bronchoalveolar lavage and endotracheal aspirate. The NDM-1 producers were susceptible only to polymyxin B. Only one patient responded to polymyxin B therapy, while the others succumbed to the infection. Conclusion: These findings reveal that NDM-1 is not a major mechanism mediating carbapenem resistance in P. aeruginosa in this centre. However, continuous surveillance and screening are necessary to prevent their dissemination.

Identification of natural compounds which inhibit biofilm formation in clinical isolates of Klebsiella pneumoniae.

Klebsiella pneumoniae, an important opportunistic pathogen, exists as a biofilm in persistent infections and in-dwelling medical devices. With the objective of identifying natural compounds inhibiting biofilm formation in K. pneumoniae, 35clinical isolates were screened,out of which 7 strong biofilm producers were identified. Six natural compounds were tested for their inhibitory effects on bacterial growth and biofilm formation by determining the minimum inhibitory concentration and minimum concentration for biofilm inhibition (MBIC) for each compound. The results show that reserpine followed by linoleic acid, were the most potent biofilm inhibitors. Reserpine, an efflux pump inhibitor was effective at biofilm inhibition at a concentration of 0.0156 mg/mL, 64-fold lower concentration than its MIC. Linoleic acid, an essential fatty acid was effective as a biofilm inhibitor at 0.0312 mg/mL, which is 32-fold lower than its MIC. Berberine, another plant derived antimicrobial, chitosan and eugenol had an MBIC value of 0.0635 mg/mL. Curcumin, a natural phenolic compound was effective at biofilm inhibition at a concentration of 0.25 mg/mL, which is 50 fold less than its MIC. Notably, the MIC and MBIC data on these 6 natural compounds was reproducible in all seven high biofilm forming isolates of K. pneumoniae. The present report is a comprehensive comparative analysis of the dose dependent inhibition of various natural compounds on biofilm formation in K. pneumoniae.

Detection of Amp C beta lactamases production in Escherichia coli & Klebsiella by an inhibitor based method.

BACKGROUND & OBJECTIVE: Detection of AmpC-mediated resistance in Gram-negative organisms poses a problem due to misleading results in phenotypic tests. There are no recommended guidelines for detection of this resistance mechanism and there is a need to address this issue as much as the detection of extended spectrum beta lactamases (ESBLs) since both may co-exist and mask each other. Though resistance to cefoxitin is used as a screening test, it does not reliably indicate Amp C production. This study was undertaken to detect Amp C beta lactamases in certain Gram-negative bacteria employing an inhibitor base test using boronic acid. METHODS: A total number of 76 consecutive non repetitive clinical isolates of Escherichia coli (n=67) and 9 Klebsiella pneumoniae (n=9) obtained over a period of two months, were screened for amp C production by disc diffusion method using cefoxitin (30 microg) dics and confirmed by inhibitor based test using boronic acid as inhibitor. RESULTS: A total of 36 of 76 isolates (47.3%) screened harboured amp C enzymes, of which a majority 31 (86.1%) co-produced ESBL enzymes. Pure ampC production was seen in 7 (9.2%) of isolates only. INTERPRETATION & CONCLUSION: Most of the amp C producers also produced ESBL enzymes. The inhibitor based test was useful in identifying cefoxitin susceptible amp C producers and could also effectively differenciate ESBL from amp C producing isolates.

Renal abscess.

This study analyses the pre-disposing factors, diagnostic modalities, therapeutic options and prognostic factors involved in 7 subjects with renal abscess. Most often they presented with high fever and flank pain. USG and CT were used to establish the diagnosis. Urine culture yielded organisms in five cases and hence empirical antibiotic therapy for Gram-negative organism was offered. Complicated abscess in our series carried high mortality especially in immuno suppressed and cachectic patients.

Detection of metallo betalactamase producing Pseudomonas aeruginosa in hospitalized patients.

BACKGROUND & OBJECTIVE: Metallo beta-lactamase (MBL)-mediated resistance to carbapenems is an emerging threat in hospital isolates of Pseudomonas aeruginosa. Though there are several screening methods to detect this enzyme production, the National Committee for Clinical Laboratory Standards (NCCLS) does not have performance standards documented so far. There is not enough information from the Indian subcontinent regarding the prevalence and the screening methods for these enzymes. The present study was undertaken to detect MBL in nosocomial isolates of P. aeruginosa by two screening methods. METHODS: Fifty consecutive P. aeruginosa isolates obtained from hospitalized patients were subjected to susceptibility testing to antipseudomonal drugs by disc diffusion, and minimum inhibitory concentration (MIC) of imipenem was determined. The production of MBL was detected by 4-fold reduction in MIC with imipenem-ethylene diamine tetraacetic acid (EDTA) and the zone size enhancement with EDTA impregnated imipenem and ceftazidime discs. RESULTS: Sixteen per cent of the isolates tested were resistant to imipenem by disc diffusion method of which 87.5 per cent exhibited a zone size enhancement with EDTA impregnated imipenem and ceftazidime discs as well as a 4-fold reduction in MIC with imipenem EDTA. The imipenem susceptible isolates (84%) had normal MIC values and exhibited no zone diameter enhancement with EDTA impregnated antibiotic discs. INTERPRETATION & CONCLUSION: MBL-mediated imipenem resistance in P. aeruginosa is a cause for concern in the therapy of critically ill patients. The two confirmatory methods i.e., zone diameter enhancement with EDTA impregnated imipenem and ceftazidime discs and 4-fold reduction in MIC with imipenem EDTA combination are equally effective for their detection.

Increase in minimum inhibitory concentration to quinolones and ceftriaxone in salmonellae causing enteric fever.

Multidrug resistance among Salmonella typhi is well known. Reports of treatment failure in enteric fever with Ciprofloxacin made us undertake this study to determine the antibiotic susceptibility pattern of S. typhi and S. paratyphi A isolated from typhoid bacteremia cases, by disc diffusion and MIC by broth dilution method. A total of 50 strains were tested, 48 of Salmonella typhi and 2 of S. paratyphi A. The disc diffusion method was done using ampicillin, chloramphenicol, cotrimoxazole, tetracycline, ciprofloxacin, ofloxacin, cefuroxime and ceftriaxone as antibiotics. The MIC was performed using ciproloxacin, ofloxacin and ceftriaxone based on standard procedure. ACCOT resistance as determined by disc diffusion method was seen in 68% of isolates. All the strains remained susceptible to flouroquinolones cephalosporins and aminoglycosides. The MIC of ciprofloxacin, ofloxacin and ceftriaxone were in the recommended range of susceptibility as given by NCCLS, 14 (28%) strains had MIC of ciprofloxacin greater than 0.5 ug/ml with 4 strains having an MIC of 1.56 ug/ml; 25 (50%) strains had MIC of ofloxacin greater than 0.5 ug/ml and 20 (40%) strains had MIC of ceftriaxone greater than 0.5 ug/ml. The high levels of MIC of ciprofloxacin may account for treatment failure cases. The rising levels of MIC of ofloxacin and ceftriaxone in S. typhi and S. paratyphi is also of concern. We document here the emergence of high levels of MIC not only to ciprofloxacin, but also ofloxacin and ceftriaxone in S. typhi and S. paratyphi A. We recommend that MIC levels of ofloxacin and ceftriaxone should be monitored along with ciprofloxacin in treatment failure cases of enteric fever.

Blastocystis hominis--an emerging cause of diarrhoea in renal transplant recipients.

Blastocystis hominis is an intestinal protozoan that is emerging as an important cause of diarrhea in the immunosuppressed population. We report two cases of diarrhea caused by this organism in renal transplant recipients. The infection was diagnosed promptly by careful stool examination and treated successfully with metronidazole. These case report highlights the fact that unusual parasites like Blastocystis hominis should be looked for and treated in cases of diarrhea occurring in renal transplant recipients. This reduces the rate of post-transplant morbidity and mortality.