PREVALENCE OF METHICILLIN RESISTANT STAPHYLOCOCCUS AUREUS (MRSA) AND ANTIMICROBIAL SUSCEPTIBILITY PATTERNS AT A PRIVATE HOSPITAL IN SANA'A, YEMEN

Methicillin resistant Staphylococcus aureus (MRSA) strains are common causes of nosocomial infections and are associated with increased morbidity and mortality. In addition, the antibiotic resistance for MRSA is a major concern in clinical practice. To study the prevalence and the antimicrobial susceptibility pattern of MRSA isolates from patients in a private hospital. The study was performed at a private hospital in Sana’a, Yemen. All the patients' samples from January, 2017 to December, 2017 were included. All isolates from inpatients’ clinical specimens (mainly respiratory secretion, pus, urine, and blood) were collected and standard isolation procedures were applied to all the samples. The records were taken from the microbiology department. Antibiotic susceptibility testing was done by Kirby Bauer's Disc diffusion technique, following the national committee for clinical laboratory standards. The antimicrobial susceptibility patterns of all the MRSA strains were studied against several antibiotics. A total of 2079 samples were gathered during the study period. Among them, 199 strains of Staphylococcus aureus were isolated. A majority of Staphylococcus aureus isolates were from pus specimen (n=81/199; 40.7%). MRSA prevalence among the patients was 17.6 % (n=35/199; 17.6). Highest proportion of MRSA was in ICU and surgical departments about 29% of all MRSA isolates. A majority of MRSA isolates were from sputum specimen (n=18/35; 51.4%). The study findings showed that MRSA isolates had the highest frequency of resistant (100%) to levofloxacin and amoxicillin/ clavulinic acid, followed by ciprofloxacin 97%, gentamicin and ampicillin/ sulbactam 94 %, cefuroxime 91 %, moxifloxacin 76%, erythromycin 71.5%, clindamycin 70.5%, and imipenem 55%. The highest frequency of sensitivity (100%) was observed with linezolide, vancomycin, and trimethoprim/ sulfamethoxazole. MRSA isolates were highly susceptible to newer drugs such as linezolid and to vancomycin which is not a commonly prescribed drug due to the higher nephrotoxic antibiotic. A more careful monitoring for use of broad-spectrum antibiotics should be instituted.


INTRODUCTION
Staphylococcus aureus is one of the most important pathogens affecting humans. Penicillin is the drug of choice to treat infections caused by Staphylococcus aureus. Because of the penicillin was commonly prescribed drugs, the prevalence of penicillin resistant strains increased, especially in hospitals 1 . MRSA was first described in 1961. It was reported after one year of introduction of methicillin. Recently, it was defined as one of the most important nosocomial pathogens. New studies suggest that the infection due to MRSA is not only hospital-acquired but community acquired as well 2 . Also MRSA has become a major public health problem worldwide 3 . Some large outbreaks of MRSA have been documented from different regions of the world, where it had caused severe infections including septicemia, endocarditis and meningitis 4 . A study by Dickinson in England and Wales has approved an increase in the death rate of MRSA infection 5  antibiotics abuse, prolonged hospitalization, intravascular instrumentation and hospitalization in an intensive care unit. There is significant variation in prevalence of clinical infections among units, hospitals and countries 4 . MRSA is of serious concern because of resistance to many other antimicrobials that are commonly used in hospitals. Another important element was that emergence of resistance to vancomycin, although at a low level has been documented 6 . Significantly, the Netherlands and countries in the Scandinavian region, had maintained low rates of MRSA infection due to strict infection control practices 7 . The problem of MRSA continues to increase and the rising colonization rates lead to the increasing of infection rates in the public and in hospitals. This can increase hospital stays and greater costs of the health care system 8 . Appropriate selection of empiric antibiotics for infections, depending on local circulating pathogens, can lead to good patient outcomes. So, epidemiologic information collected through ongoing surveillance is important to support clinicians and infection control committees to prevent and treat infection 7 .The hospitals of different sizes were facing the problem of MRSA. In addition, the problem of MRSA appears to be increasing regardless the size of hospital and its control measures for MRSA 8 . The aim of the present study is the detection of MRSA from various clinical departments at a local private hospital in Sana'a, Yemen and determining their susceptibility to some antimicrobial agents.

METHODS
The current retrospective study based on electronic laboratory records of Staphylococcus aureus isolates and MRSA isolates from clinical specimens analyzed at microbiology laboratory of university of sciences and technology (UST) hospital in Sana'a, Yemen. The records were taken from the microbiology department. Staphylococcus strains were identified based on Gram's stain morphology, colony characteristics, and biochemical identification tests. All isolates were identified as S. aureus according to standard methods 9 . A total of 2079 isolates were collected from the hospitalized patients in hospital of UST from January 2017 to December 2017. These isolates were obtained through conventional clinically oriented ordered cultures. From 2079 isolates, a total of 199 isolates were S. aureus strains. Methicillin resistance was determined using methicillin screening. All isolates were from inpatients' clinical specimens (mainly respiratory secretion, pus, urine, and blood) and Standard isolation procedures were applied to all the samples. Antibiotic susceptibility testing was done by Kirby Bauer's Disc diffusion technique, following Clinical and Laboratory Standards Institute (CLSI) 9 . The antimicrobial susceptibility patterns of all the MRSA strains were determined against the following antibiotics: vancomycin, linezolide, imipenem, levofloxacin, ciprofloxacin, cotrimoxzole, Erythromycinamoxicillin/ clavulinic Acid, cefuroxime, doxycycline, clindamycin, ampicillin/sulbactam, gentamicin, moxifloxacin, and lincomycin. Full ethical clearance was obtained from the qualified authorities who approved the study design. All data were analyzed using SPSS Statistics 21. Data was presented in tables and graphs.

RESULTS
The Figure 1 showed highest numbers of Staphylococcus aureus were obtained from pus (n=81/199; 40.7%) and least from urine (n=11/199; 5.5%). In this study, overall MRSA prevalence was 17.6 % (n=35/199; 17.6).  The study results showed that the most of the MRSA isolates prevalence in age group between 46 to 60 years about 12(34%), followed by the age more than 60 years in second rank about 8 (23%), and finally the age between 1 to 15 or years only about 2(6%). The Figure  3 showed that the most of the MRSA isolates were in male about 29(83%), whereas the female participants only about 6(17%). According to the study findings, ICU and surgical departments had the same and most prevalence of MRSA about 10(29%), followed by the medical and laboratory departments with rank about 6(17%), and finally the neurological department only about 3(8 %).
The Figure 5 showed that more than half of MRSA isolates from sputum culture about 18 Table 2 showed there were no statistical significant relationship between the usage of vancomycin and linezolid age group (P value=0.74, 0.44). The Table 3 showed there were no statistical significant relationship between the sensitivity pattern of vancomycin and linezolid hospital department (Pvalue = 0.63, 0.35).

Figure 3: Distribution of MRSA isolates according
to gender Also the study findings showed that there was no statistical significant relationship between the sensitivity pattern of vancomycin and linezolide and sample type (P value=0.80, 0.23).

DISCUSSION
Recently, the prevalence of MRSA has steadily increased worldwide 4 . In addition, the antimicrobial susceptibility pattern of MRSA strains should be described. In current study, most S. aureus strains (n=81/199; 40.7%) were isolated from pus specimen. This is consistent with a previous study done in Nairobi 10 .   In a study done to determine the antimicrobial susceptibility pattern of S. aureus strains isolated from hospitalized patients in Iran, most of the isolates were from blood specimens (29%) 11 . Another study done on prevalence and antibiotic susceptibility pattern of S. aureus from clinical isolates in Nigeria showed a majority of the isolates were from urine specimens (76%) 12 . The high number of S. aureus isolated in pus may be attributed to exposure of wounds which makes them more prone to infections and poor hygiene. In this study, overall MRSA prevalence was 17.6 % (n=35/199; 17.6). This prevalence was lower than in previous studies that reported 31.5% 13 and 46.3%14. This difference could be due to various interventions during the study period such as infection control and appropriate antibiotic usage 14, 15 . Also MRSA prevalence was higher in studies done in two private hospitals in Nairobi, Kenya, which showed a 3.8% prevalence 16 . In addition, a study in Eritrea reported 0.03% of prevalence in Dutch hospitals (17) . This approved there is high variance of MRSA prevalence from different countries. The low prevalence of MRSA in private hospitals could be attributed to better infection controls 14 . In contrast to study findings in Nigeria 6 , and Iran 11 showed different specimens were predominant. This variation in prevalence may be because of several factors like healthcare facilities available in the particular hospital, implementation and monitoring of infection control committee, rationale antibiotic usage which varies from hospital to hospital 14, 15 . In this study, 100% of MRSA isolates were sensitive to vancomycin. This finding is similar to a study done on antimicrobial susceptibility of MRSA in hospitalized patients in Iran, two hospitals in India and two private hospitals in Kenya showed 100% susceptibility to vancomycin 17,18,10 . On other hand, studies done in Iran 19 which showed 5% of the MRSA isolates were resistant to vancomycin and in a tertiary care hospital in India 20 and pediatrics and neonatal intensive care patients at Nairobi which respectively showed 3.5 and 1% resistance to vancomycin among MRSA 6 . Recently, many antibiotics with anti-MRSA activity were introduced to the market, some of which are available in Yemen. Linezolid is among these agents which is not widely available and it is not in use in many Yemeni hospitals including the setting of this study. In the present study, 100% of MRSA isolates were sensitive to linezolid. Garcia et al., have reported the first clinical outbreak of linezolid resistant S. aureus (LRSA) from a tertiary teaching university hospital in Madrid, Spain 21 . In their article they reported 12 patients with LRSA from which 6 patients died (with one death ascribed to LRSA infection). In the last two decades prevalence of MRSA has steadily increased worldwide. Therefore, they stated that this increase was associated with nosocomial transmission and extensive use of this antibiotic 22 .  In this study, the results showed that MRSA isolates were resistant to levofloxacin100%, amoxicillin/ clavulinic acid100%, moxifloxacin 76%, gentamicin 94%, clindamycin 70.5%, ciprofloxacin 97%, cefuroxime 91% erythromycin 71.5%, ampicillin/ sulbactam 94%, and imipenem 55%. The highest frequency of sensitivity (100 %) was observed with trimethoprim/ sulfamethoxazole, linezolide, and vancomycin. Multidrug-resistance made the treatment of MRSA more difficult. This may be due to dispensing antibiotics without prescription and inappropriate selection of antibiotics for infections as empirical treatment worldwide. Also the use of broadspectrum antibiotics for treating infections such as imipenem may be increase the rate of MRSA and other resistant pathogens, so a more careful monitoring of antibiotics should be instituted. Vancomycin is the universally accepted drug of choice. We found all MRSA isolates to be susceptible to vancomycin. Also maximum resistance was seen with ampicillinsulbactam, amoxicillin-clavulanic and gentamicin (Table 3). Similar results were obtained in a previous study 29 . In addition, resistance was also seen with cefuroxime (91%), doxycyclin (91.2%), and erythromycin (71.5%). Gentamicin is a most commonly used drug, because of its low cost and synergistic activity with beta-lactam antibiotics. In the present study, 94% of gentamicin resistance was seen, which was slightly higher than a previous study 30 . Most common reason for multidrug resistant MRSA is indiscriminate use of antibiotics without drug sensitivity testing which may be due to lack of advanced laboratory facilities or negligence on the part of medical practitioners or patients poor economic status. Also this is due to the fact that MRSA is often multidrugresistant 6 . Although vancomycin seems to be the only antimicrobial agent who showed 100% sensitivity and may be used as the drug of choice for treating multidrug-resistant MRSA infections, vancomycin is not a commonly prescribed drug, which is almost due to the higher nephrotoxic antibiotic and its unavailability in many parts of the country. The control of MRSA transmission seems to be the only hope to complete eradication of MRSA. The most effective way to control MRSA is good hand hygiene along with environmental cleaning of hospital rooms to reduce nosocomial rates of infection 7 .The results of current study showed that there were highly susceptible to newer drugs such as linezolid. This finding differs from a study done by Arian poor et al., in Iran which showed 5.5% of MRSA isolates were resistant to linezolid 19 .

CONCLUSION AND RECOMMENDATIONS
The present study showed a low level prevalence of MRSA. Also MRSA isolates were highly susceptible to newer drugs such as linezolid and to vancomycin which is not a commonly prescribed drug due to the higher nephrotoxic antibiotic. Also this study showed that ICU and surgical wards had the highest proportion of MRSA isolates of hospital departments. Information from this study may be used in future as a baseline for follow-up to the susceptibility trend of various drugs to be used for the treatment of S. aureus infections. Routine screening of MRSA and regular studies should be conducted to predict the trend of MRSA.

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