Characterization of Staphylococci and Evaluation of Methods for Detection of Methicillin Resistant Staphylococcus Aureus.

INTRODUCTION : Staphylococci are Gram-positive cocci that belong to the family Micrococcaceae. They are the commonest of all clinical isolates and are responsible for several suppurative types of infections. They have a differential ability to spread and cause outbreaks in hospitals. However, due to development of methicillin resistance in Staphylococcus aureus isolates; treatment of these infections has become problematic. Indiscriminate use of multiple antibiotics, prolonged hospital stay, intravenous drug use, carriage of Methicillin resistant Staphylococcus aureus (MRSA) in nose are important risk factors for MRSA acquisition. Staphylococcus aureus produces a series of enzymes and toxins and is a major pathogen causing skin abscess, wound infections, osteomyelitis, endocarditis, pneumonia, meningitis, bacteremia and toxic shock syndrome. Originally, Penicillin was the drug of choice for treatment of serious Staphylococcus aureus infections. The emergence of its resistance to penicillin was due to acquisition of a plasmid borne genetic element coding for beta lactamase production. Semi synthetic, penicillinase resistant penicillins such as oxacillin and methicillin then became the drug of choice. Gradually, over the years, Staphylococcus aureus has evolved resistance to many antibiotics especially methicillin. Methicillin resistance first appeared among nosocomial isolates of Staphylococcus aureus in 1961 and resistance is due to the presence of altered penicillin –binding protein called PBP2a that results from acquisition of a chromosomal gene called mecA. The incidence of methicillin resistant Staphylococcus aureus (MRSA) in India ranges from 30 to 70%.8 Since MRSA’s are resistant to all β- lactam antibiotics, the therapeutic options are limited significantly. Moreover, the incidence of nosocomial infections caused by MRSA continues to increase world wide, and therefore the importance of their detection, especially for therapeutic and epidemiological purposes arises. Hence methods used to detect MRSA in clinical samples should have high sensitivity and specificity and most importantly the result should be available within a short time. There are many phenotypic methods available for detection of MRSA. Therefore recognition of isolates of Staphylococcus aureus as MRSA continues to be a task that clinical microbiologist should pursue diligently and report as accurately and as rapidly as possible. Renewed emphasis on the prevention of MRSA infections is also necessary. Control of MRSA in the health care setting remains an important means of limiting its spread in the community. AIMS AND OBJECTIVES : 1. To isolate and identify Staphylococcal species from different clinical samples at PSG Hospitals, Coimbatore. 2. To determine the incidence of Methicillin Resistant Staphylococcus aureus (MRSA) and compare between the phenotypic and genotypic methods for detection of MRSA. 3. To compare the antibiotic susceptibility pattern between MSSA and MRSA, community acquired MRSA & hospital acquired MRSA and also to analyse the susceptibility pattern of Coagulase Negative Staphylococci. MATERIALS & METHODS : The study was carried out from May 2006 to August 2007 at PSG Hospitals, Coimbatore. Clinical samples such as sputum, blood, pus, CVP tips, tracheal aspirates, urine etc were processed for isolation of Staphylococci. A Gram stain was performed initially, and then the samples were inoculated on two 10% sheep blood agar (one for aerobic and the other for anaerobic), MacConkey and nutrient agar plates. Blood samples sent in brain heart infusion broths were incubated for 18 hours and then sub cultured onto blood, MacConkey and chocolate agar plates. These plates were then incubated at 37o C for 24 to 48 hours. All the suspected colonies were Gram stained and subjected to various biochemical tests to identify and characterize them. RESULTS : Out of the 24321 samples sent for cultures during the study period, 5919 samples were culture positive and of these 651(10.9%) isolates were staphylococci. Of the 651 Staphylococcus species isolated, 610 were identified as Staphylococcus aureus and 41 as clinically relevant CONS. Among the 610 Staphylococcus aureus, 208 were identified as MRSA (34.1%) based on cefoxitin resistance and 402 as MSSA based on cefoxitin susceptibility. Of the total MRSA’s, 82% were hospital acquired and 18% were community acquired. CONCLUSION : Staphylococcus aureus was isolated from 610 out of 5919 positive samples obtained from different infections in the study period . Methicillin resistant Staphylococcus aureus constituted to 34%(208) out of 610 isolates of Staphylococcus aureus. Elderly patients were significantly more prone for infections with MRSA as compared to children and young adults. In our study, only 8% of infections in neonates were caused by Staphylococcus aureus and almost half of these were MRSA and rest (92%) of the infections were caused by Gram negative bacilli as reported in many other studies. Most cases of MRSA in our hospital were contributed by surgical site wound infection (25%). Methicillin-resistant Staphylococcus aureus (MRSA) have also been isolated with increasing frequency from wound and skin infections in patients with diabetes (24%). Among all infected burns cases,16% were caused by MRSA. We have come to the conclusion that cefoxitin disk diffusion is a better and cheaper method for identification of MRSA and errors in detection of oxacillin resistance can have serious adverse clinical consequences. False susceptibility results may result in treatment failure and the spread of MRSA, if appropriate infection control measures are not applied. Conversely false resistance may increase health care cost following unnecessary isolation precautions and may lead to over use of glycopeptides and later VRSA might emerge