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Indian Journal of Medical Microbiology, Vol. 28, No. 2, April-June, 2010, pp. 127-129 Original Article Characterisation of methicillin resistant S. aureus strains and risk factors for acquisition in a teaching hospital in northeast India *JB Sarma, GU Ahmed Department of Microbiology, Northumbria Healthcare NHS Foundation Trust Newcastle-Upon-Tyne, UK Correspondence Address: *Department of Microbiology, Northumbria Healthcare NHS Foundation Trust Newcastle-Upon-Tyne, UK jayanta.sarma@nhs.net Date of Submission: 29-Jul-2009 Code Number: mb10039 PMID: 20404458 DOI: 10.4103/0255-0857.62489 Abstract Purpose: A point prevalence study was carried out in a teaching hospital in Assam to characterise S. aureus strains, establish the rate of colonisation of methicillin resistant S. aureus (MRSA) and associated risk factors for its acquisition. Keywords: HA-MRSA colonisation, antibiotic consumption, risk factors, infection control Introduction MRSA strains are now prevalent worldwide [1] since these first appeared in the UK and Denmark in early sixties soon after the introduction of methicillin. The rapid spread of bacteria resistant to antimicrobials, although a global phenomenon, is perceived to be higher in developing countries attributed mainly to misuse of antibiotics, [2],[3],[4],[5],[6] including use for prophylaxis [7],[8] However, most of these reports are anecdotal and not representative of all countries or different geographic areas within the same country [9] This study was conducted to determine the rate of colonisation of MRSA, associated risk factors for acquisition, particularly exposure to antimicrobials, and to characterise the prevalent strains in a teaching hospital in Assam which had little or no infection control interventions at the time of the study. Material and Methods A point prevalence study was conducted involving 182 male patients in three surgical, one orthopaedic and four medical units in August, 2001, in a teaching hospital in Assam. Baseline demographic data, diagnoses, procedures performed and antimicrobials prescribed with their duration were recorded from the patient's medical record. Dry swabs from nose and throat, and any wound accessible without removing the dressings were collected. The swabs were plated on to Mannitol Salt Agar (MSA) and incubated overnight. The suspect yellow colonies of S. aureus on the MSA plate that agglutinated with Pastorex (Sanofi Diagnostics Pasteur Ltd., Paris, France) were slopped in nutrient agar and shipped in triple containment packaging with labelling and marking meeting United Nation's requirements for transportation of "Biological Substance, Category B" to the microbiology laboratory at the Royal Victoria Infirmary, Newcastle upon Tyne, UK. No other regulatory permission was sought as the only pertinent regulation, Government of India's Guidelines for Exchange of Human Biological Material of 19th November, 1997 for Biomedical Research Purposes, which defines 'Human Material with potential for use in biomedical research', does not include infectious agents. Isolates were tested against a range of antimicrobials using British Society of Antimicrobial Chemotherapy (BSAC) standardized disc sensitivity method using Oxoid® (Oxoid Ltd, Cambridge, CB5 8BZ, UK) discs to generate Antibiogram-Resistogram profile. S. aureus isolates were further characterized by phage typing. A few representative isolates of MRSA with indistinguishable phage pattern were typed by PFGE. The phage and PFGE typing were carried out by the Laboratory of Hospital Infections, PHLS (now HPA), London. Consumption of antimicrobials was calculated in Defined Daily Doses (DDD)/100 bed-days as defined by the WHO Collaborating Centre for Drug Statistics Methodology. A case was defined as a patient colonised with MRSA. Equal numbers of controls (patients not colonised) matched for the duration of hospital stay were selected randomly using SPSS (version 10) from the cohort. The case-control study studied the following risk factors for acquisition of MRSA (1) exposure to third generation cephalosporins, aminoglycosides and fluoroquinolones, (2) undergoing surgery and (3) the duration of hospitals stay. Age was not considered a risk factor as the cohort was relatively young (mean age 34 in orthopaedic/surgery and 38 years in medicine) with no serious co-morbidities. Results Consumption of antimicrobials Overall consumption antimicrobials for systemic use were 92.6, 59.9 and 53.3 DDD/ 100 bed days in the surgical, orthopaedic and medical units, respectively. Consumption of cephalosporins and aminoglycosides were similar across the specialities (22-29 and 8-12 DDD/100 bed-days, respectively), but fluoroquinolone consumption was disproportionately higher in surgery - 41 as opposed to only 7-8 DDD/100 bed-days in orthopaedic and medicine. Overall, the third generation cephalosporins, fluoroquinolone and aminoglycosides constituted the bulk of the total consumption. Older and inexpensive drugs e.g., anti-staphylococcal penicillins, tetracyclines, macrolides, sulfonamides and trimethoprim were found to be either no or little in use. Glycopeptides were not in use, except in the orthopaedic ward (2 DDD/100 bed-days). Phage typing of S. aureus A selection of MRSA (n=21/27) and methicillin sensitive S. aureus, MSSA (n=15/18) isolates were phage-typed. All but two MRSA isolates (n=19) were typable and belonged to phage Group III in two distinct sub-groups, so called, A (n=15) and B (n=2). The remaining two MRSA isolates were distinct strains. In contrast, only nine MSSA isolates were typable; five were distinct strains and four were evenly distributed in two distinct phage groups. Pulse field gel-electrophoresis typing PFGE was done on six representative MRSA isolates from phage subgroups III-A (n=4) and III-B (n=2) which confirmed that each group was distinct representing a single strain. Rate of colonisation of MRSA 34% (n=29) patients in orthopaedics and 18% (n=80) in surgery were colonised with MRSA whereas only ~1% (n=73) in the medical units. Antibiogram-Resistogram Eighteen anti-staphylococcal antimicrobials were tested against MRSA. 100% of MRSA isolates (n=27) were resistant to erythromycin, trimethoprim, ciprofloxacin, gentamicin and tobramycin; 85% to clindamycin and 96% were also resistant to tetracycline, co-trimoxazole, amikacin, netilmicin and neomycin. In contrast, 100% of MSSA isolates (n=18) were sensitive to these antimicrobials (P < 0.001). Resistance to chloramphenicol (MRSA 56%; MSSA 44%) and rifampicin (MRSA 15%; MSSA 44%) did not show any correlation with methicillin sensitivity. 100% of MRSA isolates were sensitive to teicoplanin, vancomycin, synercid, fusidic acid and mupirocin. 18% of MSSA isolates were also found to be sensitive to penicillin. Risk factors Mantel-Haenszel odd ratio was calculated with 95% confidence interval with an associated P value for risk factors studied. The duration of stay was not significantly associated with MRSA carriage (P=0.071). Having a surgery (P=0.024) and exposure to ciprofloxacin (P=0.050) were risk factors for acquisition of MRSA. Discussion MRSA evolved with the acquisition of the mecA carried by SCCmec element.[10] The single clone theory suggests that MRSA evolved from a limited numbers of susceptible clones that acquired the mec element and subsequently generated substantial chromosomal diversity. [11] However, the widely accepted multi-clone theory holds that MRSA evolved several independent times by lateral gene transfer of the mec element into phylogenetically distinct methicillin susceptible cell lines. [12] Irrespective of the evolutionary origin, MRSA strains worldwide belong to only to five distinct clonal complexes, CC5, CC8, CC22, CC30, and CC45. [13] Five major multi-drug resistant clones of hospital acquired-MRSA (the Iberian, Brazilian, Hungarian, New York/Japan, and Paediatric clones) disseminated globally and accounted for majority of the hospital acquired-MRSA infections in several regions. [14] In our study, over 90% isolates were represented by a single dominant strain. The major clone predominant in Asian countries including India, except Korea and Japan, is the Brazilian/Hungarian strain (CC8-MRSA-III). [15],[16] In our study, the high co-relation of resistance to multiple antibiotic classes to methicillin resistance is almost conclusive of the presence of SCCmec type, either II or III, which determines multiresistance while other SCCmec types encode exclusively for resistance to â-lactam antibiotics.[17] The presence of SCCmec III would imply that the strains from this hospital in Assam are evolutionarily related to the predominant Asian clone (CC8-MRSA-III). Besides, these strains probably carry additional resistance genes at other chromosomal sites and on plasmids given the resistance seen to extended range of antibiotic classes. In the current study the colonisation rate was underestimated as enrichment technique was not used. Direct plating of MRSA swab on to MSA plate has a sensitivity of only 79% compared to the most sensitive conventional culture methods of broth (TSB) enrichment. [18] Ciprofloxacin usage was associated with selection of MRSA in other studies. [19] Surgery was a risk factor for acquisition of MRSA. This is probably because surgical patients were exposed to antimicrobials as prophylaxis which in every single case invariably continued irrationally for several days. Duration of hospital stay was not identified as a risk factor. This was despite the fact that almost all MRSA were hospital acquired, yet a vast majority of patients remained free from MRSA implying that that even in an environment where MRSA is endemic and there is little or no infection control interventions allowing uncontrolled cross-transmission, a significant number of patients not exposed to antimicrobials remain free from colonisation. This finding underlines the importance of 'colonisation resistance' against the nosocomial pathogens offered by the commensals flora, which is damaged by exposure to antimicrobials. In hospitals where identification based on microbiological screening of colonised/infected patients and isolation are not feasible, rational empirical antibiotic prescribing and basic infection control measure viz., hand hygiene should constitute the core of a programme to improve outcome and ensure patient safety against healthcare associated infections. Recent healthcare campaigns with demonstrable success have focused not only on hand hygiene but have also included prudent use of antibiotics. [20] eriodic studies like this for a finite period could act as a 'probe' in determining the incidence and prevalence of antimicrobial-drug resistance, associated risk factors as well as the most cost-effective regimen for specific infections in defined patient cohorts. [21] References
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