Correlation of methicillin resistance and virulence genes of Staphylococcus aureus with infection types and mode of acquisition in Sofia, Bulgaria

1*Gergova, R. T., 1Tsitou, V. S., 2Gergova, I. I., 1Muhtarova, A. A., and 1Mitov, I. G.
1Department of Medical Microbiology, Faculty of Medicine, Medical University of Sofia, 2 Zdrave str., 1431-Sofia, Bulgaria
2Department of Military Epidemiology and Hygiene, Military Medical Academy, Sofia, Bulgaria *Correspondence to:; +35929172547

Background: Infections due to methicillin resistant Staphylococcus aureus (MRSA) which is the most virulent species among the staphylococci have become a global health challenge. The aim of this study was to assess the correlation of genes encoding virulence and methicillin resistance in invasive and non-invasive isolates from inpatients/outpatients with staphylococcal infections in Sofia, Bulgaria. Materials and methods: Non-duplicate S. aureus isolates were recovered from clinical samples obtained from a total of 368 in-patients with healthcare-associated infections and outpatients with community acquired infections, following overnight cultures of samples on Columbia agar with 5% sheep blood at 35°C. The isolates were presumptively identified by colony and Gram stain morphology, positive catalase reaction and plasma-coagulase test. Isolates were screened for methicillin resistance by the cefoxitin disk method according to the European Committee on Antimicrobial Susceptibility Testing (EUCAST) protocol. The mecA and mecC, and 12 staphylococcal virulence genes were detected by a combination of monoplex and multiplex polymerase chain reaction (PCR) assays.
Results: The prevalence of MRSA based on carriage of mecA gene was 12%; 7.7% for outpatients and 16.2% for inpatients (p<0.05). The frequency of toxin genes detection in the staphylococcal isolates were as follows; sei (72.6%), seb (59.8%), seh (41.3%), sec (38.3%), seg (37.5%), sej (32.3%), sea (26.6%), sed (10.3%), tst (6.5%), and see (4.3%). The virulence genes, tst, sea, seb, sec, seg, seh and sei were more frequently associated with MRSA than methicillin sensitive (MSSA) strains (p<0.05). About one-third of the clinical S. aureus isolates harbored seven virulence genes; sea, seb, sec, see, seg, seh and sei, that were detected significantly more among the invasive isolates (p<0.05).
Conclusions: This study shows the occurrence of highly virulent staphylococcal isolates in our geographical region.

Key words: Staphylococcus aureus, virulence, methicillin resistance

Received May 17, 2019; Revised June 5, 2019; Accepted June 7, 2019 Copyright 2019 AJCEM Open Access. This article is licensed and distributed under the terms of the Creative Commons Attrition 4.0 International License (//, which permits unrestricted use, distribution and reproduction in any medium, provided credit is given to the original author(s) and the source.

Corrélation des gènes de résistance à la méthicilline et de virulence de Staphylococcus aureus avec les types d’infection et le mode d’acquisition à Sofia, Bulgarie

1*Gergova, R. T., 1Tsitou, V. S., 2Gergova, I. I., 1Muhtarova, A. A., et 1Mitov, I. G.
1Département de microbiologie médicale, Faculté de médecine,
Université médicale de Sofia, 2, Zdrave str., 1431-Sofia, Bulgarie
2Département d’épidémiologie et d’hygiène militaires, Académie de médecine militaire, Sofia, Bulgarie *Correspondence à:; +35929172547


Contexte: Les infections dues à Staphylococcus aureus résistant à la méthicilline (SARM) l’espèce la plus virulente parmi les staphylocoques, sont devenues un problème de santé mondial. Le but de cette étude était
Methicillin resistance and virulence genes of S. aureus Afr. J. Clin. Exper. Microbiol. 2019; 20 (4): 280-288
d’évaluer la corrélation des gènes qui codant pour la virulence et la résistance à la méthicilline avec des isolats invasifs ou non invasifs de patients hospitalisés/ambulatoires patients infectés par le staphylocoque dans Sofia, Bulgarie. Matériels et méthodes: Des isolats de S. aureus non dupliqués ont été récupérés à partir d’échantillons cliniques prélevés chez 368 patients atteints d’infections associées aux soins de santé et de patients ambulatoires présentant des infections acquises en communauté, après avoir effectué des cultures pendant la nuit d’échantillons sur de la gélose Columbia contenant 5% de sang de mouton à 35°C. Les isolats ont été présumés identifiés par la morphologie de la colonie et de la coloration de Gram, la réaction positive à la catalase et le test plasma-coagulase. Les isolats ont été criblés pour la résistance à la méthicilline par la méthode de la céfoxitine selon le protocole du Comité européen sur les tests de sensibilité aux antimicrobiens (EUCAST). Les gènes mecA et mecC, ainsi que 12 gènes de virulence staphylococcique ont été détectés par une combinaison de tests de réaction en chaîne de la polymérase (PCR) monoplex et multiplex. Résultats: La prévalence de SARM basée sur le portage du gène mecA était de 12%; 7,7% pour les patients ambulatoires et 16,2% pour les patients hospitalisés (p<0,05). La fréquence de détection des gènes de toxines dans les isolats de staphylocoques était la suivante: sei (72,6%), seb (59,8%), seh (41,3%), sec (38,3%), seg (37,5%), sej (32,3%), sea (26,6%), sed (10,3%), tst (6,5%) et see (4,3%). Les gènes de virulence, tst, sea, seb, sec, seg, seh et sei étaient plus fréquemment associés à SARM que les souches sensibles à la méthicilline (MSSA) (p<0,05). Environ un tiers des isolats cliniques de S. aureus portaient sept gènes de virulence; sea, seb, sec, see, seg, seh et sei, qui ont été détectés significativement plus parmi les isolats invasifs (p<0,05). Conclusion: Cela crée un risque de propagation d’isolats très virulents dans la région géographique

Mots-clés: Staphylococcus aureus, virulence, résistance à la méthicilline

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Correlation of methicillin resistance and virulence genes of Staphylococcus aureus with infection types and mode of acquisition in Sofia, Bulgaria

Pattern of resistance to vancomycin and other antimicrobial agents in staphylococcal isolates in a university teaching hospital

BO Olayinka, AT Olayinka, JA Onaolapo, PF Olurinola



Multidrug resistance has been reported in clinical isolates of both coagulase-negative staphylococci (CoNS) and Staphylococcus aureus that are most often resistant to oxacillin/methicillin. Vancomycin, a glycopeptide is the drug of choice for infections caused by such multidrug resistant strains. This study determined the pattern of resistance to vancomycin and other antimicrobial agents in staphylococcal isolates from a University Teaching Hospital. Staphylococcal isolates from clinical specimens submitted to the diagnostic medical microbiology laboratory of the Ahmadu Bello University Teaching Hospital, Zaria (over a three-month period) were characterized using standard microbiological procedures and their susceptibility to vancomycin and other commonly used antimicrobial agents determined by Kirby-Bauer-NCCLS modified disc diffusion technique. A total of 56 of the 97 (57.7%) staphylococcal isolates characterized were resistant to vancomycin 30μg, showing a zone of inhibition less than 15mm. Most of these isolates were from urine (27.3%), wound (21.8%) and pleural aspirate (12.8%). The 56 staphylococcal isolates were made up of 75% (41/56) Staphylococcus aureus and 25% (14/56) coagulase-negative staphylococci. Majority of the isolates, 60.7% (34/56) produced β-lactamase enzyme. Resistance pattern to other antimicrobial agents was benzyl penicillin G (92.9%); tetracycline (69.6%); cefuroxime (60%); chloramphenicol (54.5%); oxacillin (49.1%); erythromycin (35.7%); gentamicin (25%) and ciprofloxacin (16.1%). Analysis of the multiple antibiotic resistance index (MARI) showed that majority (91.1%) were resistant to 3 to 7 of the other antimicrobial agents tested. No isolate was resistant to all the tested antimicrobial agents. A very high proportion of the staphylococcal isolates were resistant to vancomycin, a glycopeptide that is not commonly used in this environment. Ciprofloxacin and gentamicin appear to be the only agents that will be effective in treating infections by these isolates. The high proportion of isolates with MARI of 0.3 and above, suggest that the isolates originated from an environment where antibiotics are often used. There is need for constant, on-going antimicrobial resistance surveillance in important and commonly isolated clinically significant pathogens to form the basis for developing and implementing measures that will reduce the burden of antimicrobial resistance.

Key Words: vancomycin, methicillin resistance, Staphylococcus aureus, coagulase-negative staphylococci, antimicrobial agents

Afr. J. Clin. Exper. Microbiol. Vol.6(1) 2005: 21-27