PREVALENCE, ANTIMICROBIAL SUSCEPTIBILITY PATTERN AND RISK FACTORS OF MRSA ISOLATED FROM CLINICAL SPECIMENS AMONG MILITARY PATIENTS AT 48 MEDICAL COMPOUND IN SANA'A CITY-YEMEN

Objective: Methicillin-resistant strains of S. aureus evolved in the 1970 and have troubled hospitals worldwide with persistent infections in patients. The objectives of this study were to determine the prevalence, antimicrobial susceptibility pattern and risk factors of MRSA isolated from clinical specimens among military patients at 48 Medical Compound in Sana'a city - Yemen. Methods: The study included 233 patients of whom suffering from Staphylococcus aureus infections. Specimens and data collected from November 2016 to November 2017. Standard methods of isolation and identifications were used to isolate bacteria in pours culture then Staphylococcus aureus were identifying using standard cultural techniques. MRSA was determined by the disc diffusion method to oxacillin and antimicrobial susceptibility testing was performed by the disc diffusion method for selected antibiotics. Results: The prevalence rate of MRSA was 19.3% and there was significant association between MRSA and older age patients, and surgical site infections. There was higher rate of antibiotics resistant for tested antibiotics in MRSA isolates comparing with lower rate of antibiotics resistant in MSSA. 60% of the MRSA isolates were resistant to vancomycin. They were also susceptible to erythromycin and rifampicin (100%), but showed resistance to Cotrimoxazole and Gentamycine. Conclusion: In conclusion, the emergence of S. aureus isolates resistant to vancomycin and other wide range of antibiotics have raised MRSA in Yemen into a multi-drug-resistant ‘Superbug”, making it more and more dangerous than ever in hospital environments. Regular surveillance of hospital associated infections and monitoring antibiotic sensitivity pattern and strict drug policy for antibiotics used within and outside the hospital environments are recommend.


INTRODUCTION
Staphylococcus aureus (S. aureus) is a bacterium of significant importance because of its ability to cause a wide range of diseases and capacity to adapt to diverse environmental forms 1 . The organism colonizes skin, skin glands and mucous membrane, causing infections both in human and animals such as rashes, inflammations of bones and the meninges as well as septicaemia 2 . At present, the majority of S. aureus strains are resistant to most penicillin derivatives and ordinary antimicrobial agents like drugs from the family of aminoglycosides, macrolides, chloramphenicol, tetracyclines and fluoroquinolones 3 . The global spread of MRSA constitutes one of the most serious contemporary challenges to the treatment of hospitalacquired infections 4 . MRSA carries a uniquely effective antibiotic resistance mechanism that can protect the microorganisms against all members of β-lactam antibiotics. This makes infections caused by these pathogens very difficult to manage and costly to treat 5, 6 . The objectives of this study were to determine the prevalence, antimicrobial susceptibility pattern and risk factors of MRSA isolated from clinical specimens among military patients at 48 Medical Compound in Sana'a city-Yemen.

SUBJECTS AND METHODS Ethical Consideration
Ethical clearance for the study was taken from the Faculty of Medicine and Health Sciences Research Review Committee. A written permission was also taken from the administrative Manager of the 48 Medical Compound, Sana'a city, Yemen. Informed Consent was taken from the patients before the questionnaire was filled.  18-24h. The blood cultures were performed using an automated blood culture system, followed by bacterial growth inspection. The identification of isolates was made according to standard methods 9 for any potential clinically significant growth appear on the culture media on the base of quantity, feature of growth, source and site of specimens. The primary identification was made with basic microbiological methods using colony morphology, Gram staining, catalase and coagulase tests 7 . All the Staphylococcus aureus isolates were tested for the susceptibility to 5μg Methicillin disc and 1μg Oxacillin disc provided by Difco using the disk diffusion method as described by NCCLS. The resistance breakpoints were ≥14mm to ≤ 10mm for 5μg Methicillin, and ≥ 12 mm to ≤ 10mm for 1 μg Oxacillin. The ability of other antibiotic disc to inhibit MRSA or MSSA were estimated according to the guidelines provided by NCCLS using commercially available discs which include: azithromycine (AZM, 15µg), chloramphenicol (C, 30µg), gentamycine (GN, 10µg), trimethoprimsulfamethoxazole (SXT, 1, 25/23, 75µg), Oxacillin (OX, 1µg), erythromycin (E, 15µg), cefoxitin (FOX, 30µg), nitrofurantoin (NIT, 300µg), tetracycline (T, 30µg), teicoplanin (TE, 30µg), clindamycin (CC, 2µg), rifampicin (RA, 5µg), ciprofloxacin (5 µg, Ci), vancomycin (V, 30µg) and others listed in Table 4.

Data analysis
The analysis of data was done by Epi Info version 6 statistical program (CDC, Atlanta, USA), where the chi-square (χ 2 ) and probability value (p) was calculated for the test of significance. In addition, Odd's ratio (OR), confidence interval (CI) were added to estimate the risk factors of contracting MRSA. Table 1, Table 2, and Table 3 show the prevalence of MSSA and MRSA among isolated coagulase positive (S. aureus), the potential associated factors of MRSA in patients with S. aureus infection and the association between prevalence of MRSA and type of hospital wards. Also, the susceptibility pattern of S. aureus strains is presented in isolates comparing with lower rate of antibiotics resistant in MSSA. Also, 60% of the MRSA isolates were resistant to vancomycin. They were also susceptible to erythromycin and rifampicin (100%), but showed resistance to Cotrimoxazole and Gentamycine.   13 . This variation can be explained by that the biology of CA-MRSA appears to differ from that of HA-MRSA and CA-Methicillin-susceptible S. aureus (MSSA), perhaps allowing CA-MRSA to cause disease other than that expected from MSSA 14, 15 . As HA-MRSA emerged, it likely did not merely replace HA-MSSA but led to an overall increase in S. aureus infections in healthcare settings 16, 17 . In addition nearly all the researchers say the same thing that in-patients and outpatients have higher S. aureus/ MRSA infections than S. aureus/MSSA because of wide spread of MRSA in community and hospitals environment 17,18,19 . When we considered age as risk factor of MRSA in current study, the highest incidence of MRSA in current study's cases were in age groups of >median (24 years). The risk age group of in-patients and outpatients in current study was different from that reported previously in several studies in which roughly equal rates of MRSA in the different age groups 20 . In current study when it was considered that the association between prevalence of MRSA and sites of infections (type of clinical specimens), the highest prevalence MRSA was 33.3% in surgical site infection (SSI) with associated odds ratio equal to 2.2 (P>= 0.05) ( Effective antimicrobial activity as well as cost effectiveness should be considered in drugs prescribed for MRSA infections. Oral dosing options for antibiotics can allow earlier discharge of hospitalized patients and minimize the chances of VRSA emergence. Good hospital infection control measures prove to be the main stay against these infections because antibiotics can never be an effective alternative to good medical practice. Also, medium sensitivity percentages were found to gentamycine, trimethoprimsulfamethoxazole and tetracycline (Table 4). Different sensitivity percentages have been reported to these three antibiotics worldwide 28,29,30 . These differences might be due to prolonged antibiotic treatment, age, type of infection and geographical variation.

CONCLUSION
In conclusion, the results of this study showed the importance of regular surveillance of hospital associated infections including monitoring antibiotic sensitivity pattern and strict drug policy for antibiotics used within and outside the hospital environments. Moreover, in-vitro susceptibility testing of every isolate of MRSA in the clinical laboratories may be helpful for reducing the incidence of these infections.

ACKNOWLEDGMENTS
Authors acknowledge the support of Sana'a University and 48 Medical compounds, Yemen.

AUTHOR'S CONTRIBUTION
The manuscript was carried out, written, and approved in collaboration with all authors.