Nasal Carriage Rate, Antimicrobial Susceptibility Patterns and Associated Risk Factors of Methicillin-resistant Staphylococcus Aureus Among Prisoners in Dessie Town, Northeast Ethiopia

Background: The prevalence of Methicillin-resistant S. aureus (MRSA) within the general public has caused outbreaks in groups of people in close quarters such as military barracks, gyms, day care centres and correctional facilities. Correctional facilities are of particular importance for spreading MRSA, as inmates are often in close proximity and have limited access to hygienic products and clean clothing. In Ethiopia, there is a limited number of studies about methicillin-resistant S. aureus in the prison setting. Therefore, this study aimed to determine nasal carriages rate, antimicrobial susceptibility patterns, and associated risk factors of MRSA among prisoners in Dessie town, Northeast Ethiopia. Methods: An institution-based cross-sectional study was conducted from January 1 to March 30/2020 on 329 prisoners. Nasal swabs were collected and processed on blood agar and Mannitol salt agar plates. The isolates were identied as S. aureus based on morphology, coagulase test, and mannitol salt agar fermentation. Disk diffusion antibiotic susceptibility tests were performed according to the guidelines of the Clinical and Laboratory Standards Institute. Methicillin-resistance by S. aureus was conrmed by using cefoxitin (30μg). Data entry and statistical analysis was done using SPSS Version 20. Results: Out of the 329 prisoners, 115(35%) carried S. aureus, of which 35/115 (30.4%) were MRSA. Therefore, 10.64% of all prison inmates were identied as MRSA carriers. MRSA carriage rate was highest among the age groups of 24-34 and 35-45, which was 12 (34.3). Penicillin showed the highest rate of resistance among MRSA and MSSA isolates (100%). The multi-drug resistance rate was very high (50.4%). Among 115 S. aureus isolates, about 2(7.4%) showed inducible Clindamycin resistance. Conclusion: The high rate of nasal MRSA carriage among prisoners found in this study is alarming and highlights the need for adjusted infection control measures to prevent MRSA transmission within inmates or to the community during their discharge.

Data released by the U.S. Centers for Disease Control and Prevention (CDC) in 2019 showed that S. aureus infections are a major problem in the United States, with 119,000 infections and almost 20,000 deaths in 2017. In Louisiana, it is estimated that out of a 4,500,000 population, 1,500,000 are carriers of S. aureus and 45,000 are carriers of MRSA [6]. According to the report of the Pan-European Institute, MRSA was estimated to affect more than 150,000 patients annually in the European countries and is responsible for the expenditure of 380 million Euros in extra costs for healthcare systems [7]. Methicillin-resistant S. aureus is widely prevalent in most Asian countries too. For instance, multidrug-resistant MRSA are endemic in most hospitals, with an estimated prevalence from 28% in Hong Kong and Indonesia to greater than 70% in Korea [8].
In Africa, the burden of MRSA has remarkable variations across countries due to a lack of advanced laboratories, effective and systematic routine surveillance systems. Due to this, the prevalence of methicillin resistance in S. aureus isolates rises up to 82% (34). South Africa and central Africa are among regions where a high rate of MRSA prevalence reported with 24.4% and 40.4% respectively [9].
In Ethiopia, MRSA is spreading quickly both in hospital and community settings at a very alarming rate [10]. The study conducted in Jimma town, among prisoners, depicted that the overall prevalence of MRSA was 48% [11]. A study conducted among health workers of Dessie Referral Hospital showed the prevalence of MRSA was 12.7% [12]. Another study conducted in Gondar town, among urban and rural elementary school children, showed that the prevalence of MRSA was 9.79% [13]. The spread of MRSA in the health care system is a serious opportunity for transmission of MRSA outside the health care system. However, most of the studies related to MRSA colonization are limited to health care institutions. This is a relevant issue, since prison characteristics vary in different countries. Moreover, since those institutions harbor closed populations, it is important to know whether the prevalence of MRSA from those prisons similar to the prevalence in the hospitals or in the community. Therefore, this study will ll this gap and determine the prevalence of carriage, antimicrobial susceptibility patterns, and risk factors of MRSA among prisoners in Dessie town.

Study Setting, Design, and Period
An institution-based cross-sectional study was conducted in Dessie town prison center from January 1 to March 30/2020. Dessie town is located in Amhara Regional State, Northeast Ethiopia, 401 km far from Addis Ababa, and 471 km far from Bahir Dar. Based on the 2013 national census conducted by the Central Statistical Agency of Ethiopia (CSA) [14], Dessie town has a total population of 151,174, of whom 72,932 are men and 78,242 women [15]. During the study, about 1350 prisoners were jailed.

Sample size and Sampling technique
The sample size was determined by using a single population proportion formula by considering the prevalence of 48% [11], with a 95% con dence interval, a 5% margin of error, and with 10% no-response rate.
A systematic random sampling technique was used to recruit 329 study percipients from the sampling frame formed by the registration number of prisoners. Prisoners with active upper respiratory tract infection, fever, recent nasal surgery, and use of nasal antimicrobial therapy were excluded.

Data collection procedures
A well-structured questionnaire was rst prepared in English and then translated to Amharic language (local language). The second version of the questionnaire was retranslated into the original one by language experts to evaluate its consistency. The pre-tested nal version of the questionnaire was used to collect data from study participants. Face-to -face interview technique was applied to collect data related to socio-demographic, clinical, and treatment-related variables as well as hygiene related factors.

Sample collection and transportation
Nasal swabs were collected by a trained laboratory technologist as per the collection and transportation procedures. A single sterile swab was used for each nostril. The sterile cotton swab was pre-moistened with sterile normal saline (0.85% NaCl) before swabbing. The moistened cotton swab was introduced to the nostrils 2-3 cm and rotated 4-5 times both clockwise and anticlockwise before the withdrawal. The collected specimens were placed into Amies transport media and transported with an icebox to Amhara Public Health Institute, Dessie branch for further processing within 2 hours of collection [16,17].

Bacterial isolation and identi cation
The nasal swab specimens were inoculated onto Sheep Blood Agar (Oxoid, LTD, UK) and Mannitol Salt Agar (Oxoid, England). All plates were incubated at 37ºC for 24 hours under aerobic conditions. Preliminary identi cation of S.aureus was done based on the colony morphology on Mannitol and blood agar plates. Further con rmatory identi cation was done by Gram staining, catalase, mannitol fermentation, and coagulase tests following standard procedures [18].

Antimicrobial susceptibility testing
The antimicrobial susceptibility test of the isolates was determined according to the Kirby Bauer disc diffusion technique as illustrated by the Clinical Laboratory Standard Institute 2018 [19]. The bacterial suspension was made from pure colony of the test organism and matched with 0.5 McFarland standards. Then, standardized culture suspension was swabbed with lawn technique onto the surface of Mueller-Hinton agar (MHA) plate. After 3 minutes, a set of 8 standard antimicrobial discs were then placed aseptically on the inoculated Mueller-Hinton agar plates and allowed to stand at room temperature for 15 minutes. All inoculated media were then incubated at 37°C aerobically. After 24hrs of incubation, the diameter of the zone of inhibition around each disk was measured with a ruler. Based on the zone of inhibition (mm) of each antimicrobial disc, isolates were classi ed as resistant, intermediate, or susceptible following the standard interpretive chart of CLSI 2018 [19]. S. aureus ATCC 25923 (oxacillin-sensitive, and ATCC 33591 (oxacillin-resistant,) were used as controls in all experiments.
Methicillin-resistance by S. aureus was detected using surrogate antibiotics of 30μg cefoxitin discs by the Kirby-Bauer disk diffusion method. All the isolates were subjected to cefoxitin disc. Similarly, 0.5 McFarland standard matched suspension of the isolates was prepared and inoculated on Mueller Hinton agar plate.
Data management and quality control Data quality control The training was given for the data collector about data collection procedures and interviewing techniques for one day. The principal investigator, together with the supervisor, supervised the data collection. The pretest was carried out at the Kombolcha prison Centre.

Laboratory quality control
Pre-analytical, analytical, and post-analytical quality assurance was maintained [18].

Data processing and analysis
Initially, data were checked for completeness and coded. After coding, data were entered using EPI-info version 3.5.1 and exported to SPSS version 20 for further analysis. Frequency analysis was carried out to assess the prevalence of MRSA. Logistic regression analyses were done to determine factors associated with the carriage rate of MRSA. A variable with p-value ≤ 0.2 in bivariate logistic regression was included in multivariate analysis. Crude and adjusted odds ratios were calculated to quantify the strength of association between MRSA carriage rate and risk factors. The 95% con dence interval was used and risk factors with p-value < 0.05 in multivariate analyses were considered as statistically signi cant.

Ethical considerations
Ethical clearance was obtained from the School of Biomedical and Laboratory Sciences, University of Gondar Ethical Review Committee. Ethical permission was obtained from Dessie prison centre o ce. Each study participant was informed about the purpose, methods of collection, anticipated bene t, and risk of study. Written informed consent was obtained from each study participant. Study subjects were identi ed using codes rather than using the actual name and unauthorized persons had no access to the collected data.

Socio demographic characteristics of study subjects
Altogether, 329 prisoners were involved in this study. Of these, 288 (87.5%) were males. The age of study participants was ranged from 17 to 77 years with a mean age of 35.27 ± 10.72 years. Among the study subjects, about 145/329 (44.1%), completed primary school (Table 1).     Multidrug resistance pattern of S.aureus The overall multidrug resistance (MDR) rate of bacterial isolates in this study was 50.4%. About 16.5% of the bacterial isolates showed MDR to four antimicrobials followed by 7.8% to ve antimicrobials [ Table 6].   Table 7].

Discussion
S. aureus is a common bacterium that can colonize many parts of the body, mostly the nasal cavity. Approximately 20-30% of the world's human population is persistently colonized by S. aureus. Colonization rates of S. aureus are different in different risk groups. However, the highest frequency of carriage is observed among high-risk groups, like children and inmates in correctional institutions [21,22]. Studies have indicated that nasal colonization of S. aureus has a key role in developing subsequent infections [23].
In the present study, the overall prevalence of S. aureus colonization among prisoners was 35%. The nasal carriage rate of S.aureus observed in this study was smaller than the ndings of the study conducted in USA maximum-security prisons (54.4%) and Jimma prison center, Ethiopia (50%) [24,11]. However, the carriage rate of S. aureus in the present study was higher than studies done in Brazil (16.5%) (30). The differences might re ect the variations in isolation techniques, the sample size, time of study period, population characteristics, geographical distribution, prevention and control policies, and duration of stay in correctional facilities. This study is in line with a study conducted in Texas county jail inmates (28.5%) [11].
The overall nasal carriage rate of MRSA among prisoners was 10.64%. Of the total 115 S. aureus isolates, the prevalence of MRSA was 30.4% which was similar to the result of a study conducted in South Africa (30.9%) [25] and Saud Arabia (32%) [26]. However, this nding was lower than the reports of other studies, such as MRSA carriage rate of 41% in the manila city jail, Philippine [11], 54% in the USA [27], and 48% in Jimma town correctional facility, Ethiopia [28]. The smaller carriage rate of MRSA in the present study might be due to the variation in isolating techniques, study subjects, population characteristics, and length of study time.
In this study, the high nasal colonization of MRSA among prisoners was attributed to the contribution of different risk factors. The risk factors mainly identi ed from this study after doing multivariate logistic regression analysis were; having a history of super cial skin lesions, a history of antibiotic use, sharing of personal items, and ways of cleaning nostrils. According to the results of the present study, the nasal colonization of MRSA was predominant in participants with a history of super cial skin lesion in the previous one year (56.8%) than prisoners without a history of super cial skin lesions (17.9%). The data showed that it was 6 times more likely for prisoners with a history of super cial skin lesions to have carriage of MRSA than prisoners without it, (p < 0.001).The ndings of this study were comparable with the report of the study conducted in the Los Angeles County Jail (p = 0.001) [29]. Result was comparable with ndings reported in Bahir Dar [32] and Pakistan [33].
In the present study, the frequency of inducible clindamycin resistance was 2/115 (1.74%). This nding was inconsistent with ndings reported in Brunei Darussalam (15.7%) [34]. This difference may be resulted in variation of geographical area.

Conclusion
Nasal colonization with MRSA is the most common source for the occurrence of subsequent infection of the host. Con dentiality was maintained at all levels of the study. In addition, study participants involvement was based on a voluntary basis and participants who were unwilling to take part in the study and those who need to quit their participation at any stage were informed to do so without any restriction. Consent to participate was obtained from the study participants.

Consent for publication
All authors read the manuscript and have provided their consent to publish.