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Methicillin-resistant Staphylococcus aureus nasal carriage among primary school-aged children from Jordan: prevalence, antibiotic resistance and molecular characteristics

Alzoubi, Hamed M.a; Aqel, Amin A.a; Al-Sarayreh, Sameeh A.b; Al-Zayadneh, Enasc

The Journal Of The Egyptian Public Health Association: December 2014 - Volume 89 - Issue 3 - p 114–118
doi: 10.1097/01.EPX.0000454671.83406.e8
Original articles

Background and aim Colonization with methicillin-resistant Staphylococcus aureus (MRSA) increases the risk for subsequent infections with an increased mortality and morbidity. Children were suggested to be a major asymptomatic reservoir for community-associated (CA) MRSA with an ability to quickly spread the MRSA within community. Therefore, the availability of epidemiological and antibiotic susceptibility data of CA-MRSA will be useful for the infection control and management policies. This study aimed to assess the nasal carriage, molecular characteristics and antibiotic susceptibility of MRSA in primary school-aged children from Jordan.

Patients and methods A total of 210 nasal swabs were collected from children aged 6–11 years. Isolated MRSA and its SCCmec typing, Spa type and PVL (Panton-Valentine Leukociden) toxin were identified following culture, biochemical and PCR. Antibiogram was determined by the disc diffusion method.

Results The prevalence of CA-MRSA was 7.1%. Allergic rhinitis and recent antibiotic exposure were the only significant risk factors for MRSA nasal carriage among children. Resistance to erythromycin, trimethoprim–sulfamethoxazole and tetracycline was 33.4, 20 and 13.4%, respectively. All isolates were susceptible to the remaining non-β-lactam antibiotics used in this study, in particular linezolid and mupirocin. All MRSA isolates were SCCmec type IV and PVL toxin negative and the majority were Spa type t223.

Conclusion and recommendations This is the first study to assess the MRSA prevalence among children aged 6–11 years in Jordan. The prevalence in community children is within the range compared with other studies in other countries. The antibiogram, SCCmec and Spa types of the isolated MRSA are much similar to what was found previously in Jordan. However, all isolates were PVL toxin negative. The study recommends increasing the public awareness of MRSA and the proper antibiotics dispensing. Future studies to follow-up on the changing epidemiology of the CA-MRSA in Jordan are also recommended.

Departments of aMicrobiology and Immunology

bBiochemistry and Molecular Biology

cPediatrics, Faculty of Medicine, Mu’tah University, Al-Karak, Jordan

Correspondence to Hamed M. Alzoubi, MD, PhD, Department of Microbiology and Immunology, Faculty of Medicine, Mu’tah University, PO Box 7, Al kark, Jordan Tel: +962 799 238 884; fax: +962 323 97180; e-mail:

Received February 8, 2014

Accepted March 17, 2014

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Methicillin-resistant Staphylococcus aureus (MRSA) was first recognized in the early 1960s soon after introduction of methicillin 1. Since then, it has been confined to healthcare settings. However, since the 1990s, the number of MRSA infections in populations that lack exposure to healthcare settings has increased. This has led to the recognition of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) strains 2–4.

CA-MRSA strains are mainly distinguished from healthcare-associated methicillin-resistant Staphylococcus aureus (HA-MRSA) using molecular techniques 4. The SCCmec in CA-MRSA belongs to type IV or V and is smaller than that of HA-MRSA SSCmec, which belongs to type I, II or III. In addition, CA-MRSA strains frequently carry the gene for PVL (Panton-Valentine Leukociden) toxin, which is rarely found in HA-MRSA 4,5.

CA-MRSA infections can affect healthy young patients leading to serious community infections with increased mortality and morbidity such as skin and soft tissue infections, necrotizing pneumonia and sepsis 3,5. Colonization with MRSA has been suggested to increase the risk of developing subsequent infections 6. Therefore, such colonization represents a threat to the carrier and community as infections caused by MRSA can be difficult and more expensive to treat compared with those caused by methicillin-sensitive Staphylococcus aureus strains 7.

The risk factors for carriage of CA-MRSA in the community are not fully defined 8. However, it was found that the carriage rate of CA-MRSA among children is higher than in adults 9. In addition, adults living with young children are at increased risk for MRSA colonization 10. Therefore, it was suggested that children could be a major asymptomatic reservoir for CA-MRSA with an ability to quickly spread MRSA within other different community populations 8,11,12. As a result, studying the nasal carriage of MRSA is a necessary sentinel approach to assess the MRSA burden and its risk factors in a population.

The aim of this study was to assess the nasal carriage of MRSA among primary school-aged children in Al-Karak province, Jordan. It also aimed to describe the risk factors for colonization, antibiotic resistance and molecular characteristics of CA-MRSA. This may serve as a guide for infection control and management policies of CA-MRSA infections.

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Patients and methods

Study design and data collection

This cross-sectional study was carried out from November 2012 to May 2013 at Al-Karak province in Jordan. Approval for the study was obtained from the scientific and the ethics committees at the faculty of medicine, University of Mu’tah, Jordan. Parents gave written informed consent.

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The sample

The sample size for this study was calculated using Kish’s formula for cross-sectional studies 13 and at a prevalence of community MRSA of ∼13% in Jordan and a neighboring country 14,15. On the basis of that, the minimum sample size required for this study was 174 samples. However, 210 samples were collected from 210 children aged 6–11 years attending the University-Model school at Al-Karak, Jordan. A questionnaire was filled in with information on illnesses, having a family member who is a healthcare worker, antibiotic consumption and hospital admission over the last 3 months. Hospital admission was an exclusion criterion in this study.

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Bacterial detection, antimicrobial susceptibility testing and molecular characterization of methicillin-resistant Staphylococcus aureus isolates

This was carried out as previously described 16. Briefly, nasal swabs were inoculated on Mannitol salt agar (MSA) (BBL Microbiology System; Becton Dickinson Company, Sparks, Maryland, USA) and incubated at 35±1°C and examined for growth after 24–48 h. Single distinctive morphotype of a mannitol-positive colony was selected from an MSA plate, subcultured to a nutrient agar (BBL Microbiology Systems; Becton Dickinson) and incubated overnight at 37°C in a humidified incubator. Colonies growing on nutrient agar were identified as S. aureus by Gram staining, anaerobic utilization of glucose and mannitol, catalase production and tube coagulase test. Identifying the MRSA isolates was then carried out using 30 μg/ml cefoxitin disc in Mueller-Hinton agar.

Antimicrobial susceptibility to fusidic acid, erythromycin, mupirocin, gentamicin, linezolid, teicoplanin, ciprofloxacin, trimethoprim–sulfamethoxazole, tetracycline, rifampicin and cefoxitin was performed by disk diffusion, according to the European Committee on Antimicrobial Susceptibility Testing for fusidic acid and according to the Clinical Laboratory Standards Institute guidelines for the remaining antibiotics 17,18. S. aureus ATCC 25923 was used as control strain. Discs were purchased from Oxoid (Hampshire, UK).

Methicillin-resistance genes (mecA and mecC) and the PVL gene (Luk-PV) were detected by multiplex PCR as previously published 19; isolates Spa-typing was carried out as described by Harmsen et al.20, and the SCCmec typing was performed according to Milheiriço et al. 21.

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Statistical analysis

The statistical analysis was conducted using STATA10 (Stata Statistical Software, Release 10; Stata Corp LP, College Station, Texas, USA) to evaluate the significance of potential risk factors. A P value of less than 0.05 was considered as significant.

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The study population included 109 male patients and 101 female patients. A total of 15 MRSA were isolated from seven male patients and eight female patients. The prevalence of MRSA nasal carriage was therefore 7.1% (15/210). The distribution of the isolated MRSA according to different variables and statistical significance is shown in Table 1. The only significant variables for MRSA nasal carriage are receiving antibiotics over the last 3 months (P=0.037) and suffering from allergic rhinitis (P=0.022).

Table 1

Table 1

The results also show that 33% (5/15) and 67% (10/15) of the 15 isolates were distributed among age groups 6 to below 9 and 9 to below 12 years, respectively. However, age was not found to be statistically significant (P=0.25) for MRSA nasal carriage in the current study. Similarly, sex (P=0.67) and having a family member working as a healthcare worker (P=0.12) were not statistically significant for MRSA nasal carriage among study population.

Antibiotic susceptibility pattern is shown in Table 2. None of the 15 MRSA isolates (0%) were susceptible to cefoxitin, whereas 66.6, 80 and 86.6% of the isolates were susceptible to erythromycin, trimethoprim-sulfamethoxazole and tetracycline, respectively. All isolates were susceptible to the remaining antibiotics used in this study as shown in Table 2.

Table 2

Table 2

The MecA gene was detected in all MRSA isolates, whereas none of them were positive for the mecC and PVL gene (Luk-PV). Nine isolates were of t223 Spa type, followed by t214, t386 and t012, which were detected in three, two and one isolates, respectively. The IVa-SCCmec lineage was detected in 14 MRSA isolates and IVc-SCCmec was detected in one isolate.

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MRSA infections are globally emerging as an escalating infection in the community and can no longer be considered as a healthcare-associated infection only. Therefore, hospital-based infection control measures alone are not enough to fight the increasing MRSA infections in the community 6. CA-MRSA have been shown to infiltrate hospital settings 22. Epidemiological and antibiotic susceptibility data for CA-MRSA can be of paramount importance for practitioners to guide their infection control and management policies.

In the current study, the prevalence of CA-MRSA nasal carriage among children aged 6–11 years was 7.1%. This is nearly similar to the 7.4% prevalence rate recently found among healthy Jordanian adult volunteers 23. However, in a study carried out by Shehabi et al. 14, the prevalence of MRSA among Jordanian infants was 13.2%, which is significantly higher than what was found in the current study. In another study among children aged 1.8–5.5 years in Gaza, the prevalence of CA-MRSA was 12% 15. Such higher rate of prevalence might be explained by factors such as difference in study population and exposure to different potential risk factors. Another important factor can be the difference in the age of the studied population. Our study is the first in Jordan to study the primary school-age children, whereas the previously mentioned studies 14,15 focused on infants and adults. Therefore, the results of the current study covers an important age span, which should, together with the previous studies, provide a more comprehensive picture about CA-MRSA prevalence among different ages. In addition, this study should form a start to follow-up on the changing epidemiology of the CA-MRSA in Jordan and, probably, the region.

Variable prevalence rates of 8.8, 10.8 and 4% among primary school-age children were previously found in different countries 24–26. Therefore, the prevalence of 7.1% found in our study is still within the range of what was found in some other countries.

Factors such as age, sex and having members of the family working in healthcare setting were not found to be statistically significant for MRSA carriage in this study. In contrast, recent antibiotic exposure and having allergic rhinitis significantly increased the risk for MRSA nasal carriage.

Antibiotic exposure was previously suggested to be associated with CA-MRSA carriage among children 27. Antibiotic abuse and misuse is believed to be common in the community where this study was carried out. This is may be due to absence of restriction on antibiotic prescriptions and the proper antibiotics prescription guidelines. However, this aspect is still hypothetical and it needs further investigations. The presence of allergic rhinitis was found to be a significant factor for CA-MRSA carriage in the studied children. This is in agreement with the recently published study by Çevik et al.28, which revealed that MRSA nasal carriage was significantly higher among allergic rhinitis patients compared with control healthy people. The carriage of MRSA among allergic rhinitis patients should be considered as a factor that may increase the comorbidities and complications; therefore, proper diagnosis and antibiotic selection is necessary to manage community infections such as skin and soft tissue infection in this group of patients.

Resistance to erythromycin, trimethoprim-sulfamethoxazole and tetracycline was 33.4, 20 and 13.4%, respectively. The resistance to these antibiotics was nearly similar to some of the previous studies in Jordan 14,16. There was no resistance for the remaining antibiotics that were used in this study, in particular to linezolid, which was also not much different from what was found by some other studies in Jordan 16,23. Such narrow antibiotic resistance profile is consistent with the characteristic of the CA-MRSA 29.

All isolated MRSA in the current study belonged to the IV-SCCmec lineage, which is consistent with MRSA of community origin that usually harbour type IV or V 4, and is nearly similar to what was found previously in the region 14,15,23. The Spa type t223 was predominant among all MRSA isolates and represented 60% of all detected Spa types. This type was found as the second commonest type by Al-Bakri et al.23, where it represented 14.7% preceded by the Spa type t9519, which was detected in 76% in the same study. The significance of Spa types is still debatable with some suggestions of an association between the Spa type and CA-MRSA invasiveness. However, such suggestions have not been confirmed experimentally and further research is needed to assess that 4,30. Unexpectedly, the PVL toxin was not detected in all CA-MRSA isolates of this study. The PVL-positive MRSA isolates in studies carried out recently in Jordan by Shehabi et al.14 and Al-Bakri et al. 23 were 28 and 5%, respectively. Rare occurrence of PVL toxins in the CA-MRSA was previously reported 31. However, the absence of PVL toxin in our isolates is reported for the first time in Jordan and the neighboring countries according to the best of our knowledge and literature search. A proposed hypothesis for such finding is that the PVL genes might be less common in isolates causing asymptomatic colonization compared with those causing clinically apparent infection 4. However, further investigations might be needed in the future.

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Conclusion and recommendations

This study showed a prevalence rate of MRSA nasal carriage among primary school-aged children in Jordan comparable with those in other countries. The antibiotic susceptibility pattern is not alarming and is approximately consistent with CA-MRSA. All MRSA isolates belonged to SCCmec type IV and 60% were Spa type t223. The absence of PVL toxin in all MRSA isolates is a finding that is worth further studying. Further studies are required to sample children for MRSA nasal carriage in the future to follow-up on the epidemiology pattern and antibiotic susceptibility.

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The authors acknowledge Mu’tah University/Jordan for the financial support of this work. They also acknowledge Ahmad Khazar Zayed Makableh, medical student, at the faculty of medicine Mu’tah University, for his help in the practical part of this study.

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Conflicts of interest

There are no conflicts of interest.

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community-associated methicillin-resistant Staphylococcus aureus; Jordan; prevalence; risk factors for methicillin-resistant Staphylococcus aureus; school-aged children

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