Improved multiplex PCR primers for rapid identification of coagulase-negative staphylococci

Abstract

Coagulase-negative staphylococci (CNS) are opportunistic pathogens that are currently emerging as causative agents of human disease. Though CNS are widespread in the clinic and food, their precise identification at species level is important. Here, using 16S rRNA sequencing, 55 staphylococcal isolates were identified as S. capitis, S. caprae, S. epidermidis, S. haemolyticus, S. pasteuri, S. saprophyticus, S. warneri, and S. xylosus. Although 16S rRNA sequencing is universally accepted as a standard for bacterial identification, the method did not effectively discriminate closely related species, and additional DNA sequencing was required. The divergence of the sodA gene sequence is higher than that of 16S rRNA. To devise a rapid and accurate identification method, sodA-specific primers were designed to demonstrate that species-specific multiplex polymerase chain reaction (PCR) can be used for the identification of CNS species. The accuracy of this method was higher than that of phenotypic identification; the method is simple and less time-consuming than 16S rRNA sequencing. Of the 55 CNS isolates, 92.72% were resistant to at least one antibiotic, and 60% were resistant to three or more antibiotics. CNS isolates produced diverse virulence-associated enzymes, including hemolysin (produced by 69.09% of the isolates), protease (65.45%), lipase (54.54%), lecithinase (36.36%), and DNase (29.09%); all isolates could form a biofilm. Because of the increasing pathogenic significance of CNS, the efficient multiplex PCR detection method developed in this study may contribute to studies for human health.Electronic supplementary materialThe online version of this article (doi:10.1007/s00203-017-1415-9) contains supplementary material, which is available to authorized users.

The Prevalence of antimicrobial resistance in Staphylococcus aureus and coagulase-negative Staphylococci strains isolated from inpatients

“…Among the samples examined in this study, 6 (3.57%) had pvl gene. The occurrence of pvl gene in previous stud ies has been reported from 2 to 35% (26,27). In compari son, the obtained results for pvl gene in the present study were near the lower limit obtained from similar studies.…”

“…The frequency of MRSA and MRCoNS in different wards is shown in Table 1. (26) and, as in this study, demonstrated the importance of rapid detection of CoNS. Among the samples examined in this study, 6 (3.57%) had pvl gene.…”

The Prevalence of antimicrobial resistance in Staphylococcus aureus and coagulase-negative Staphylococci strains isolated from inpatients

“…In this study, Cefoxitin disc can be considered as a very effective method for detecting the presence of mecA gene although it was found that PCR was an easy, fast and inexpensive method for detecting the presence of this gene. Kim et al in China used the 16s rRNA gene PCR to identify different CoNS strains (26) and, as in this study, demonstrated the importance of rapid detection of CoNS. Among the samples examined in this study, 6 (3.57%) had pvl gene.…”

The Epidome - A species-specific Approach to Assess the Population Structure and Heterogeneity of Staphylococcus Epidermidis Colonization and Infection

“…In this paper we present the Epidome, a culture-independent method based on amplicon sequencing of two S. epidermidis-speci c target genes. Similar amplicon-based methods exist to identify staphylococci at genus level [25], and to distinguish between various CoNS [26][27][28].…”