MALDI-TOF-MS based identification and molecular characterization of food associated methicillin-resistant Staphylococcus aureus

Abstract

Food-borne methicillin resistant Staphylococcus aureus (MRSA) is involved in two-fold higher mortality rate compared to methicillin susceptible S. aureus (MSSA). Eventhough Mysuru recognized as cleanest city in the world, prevalence of food contamination is not detailed. The aim is to screen food samples from Mysuru area and to characterize MRSA strain, employing MALDI-Biotyper, multiplex PCR to distinguish between MRSA and MSSA by PCR-coupled single strand conformation polymorphism (PCR-SSCP). Of all the food-borne pathogens, S. aureus contamination accounts for 94.37 ± 0.02% (P < 0.01), strains characterized by means of nuc genes, followed by species specific identification by Coa, Eap and SpA genes and multiplex PCR to confirm the presence of three methicillin resistant staphylococcal species simultaneously using nuc and phoP genes. Amplification of mecA gene in 159 isolates confirmed that all strains are methicillin resistant, except UOM160 (MSSA) and multi-drug resistant (MDR) in 159 isolates confirmed by 22 sets of β-lactam antibiotics. MSSA and MDR-MRSA were discriminated by PCR-SSCP using nuc gene for the first time. From the present studies, compared to conventional methods MALDI-Biotyper emerged as an effective, sensitive (>99%), robust (<2 min), and alternative tool for pathogen identification, and we developed a PCR-SSCP technique for rapid detection of MSSA and MRSA strains.

Investigating Antifungal Susceptibility in Candida Species With MALDI-TOF MS-Based Assays

“…Therefore, the first step would be, as it was already done for some bacteria (Manukumar and Umesha, 2017), to detect specific peaks associated with the resistance or the susceptibility to an antifungal drug (Posteraro et al, 2013). This aim is promising since typing with MALDI-TOF MS was shown to be able to cluster C. glabrata isolates according to their fluconazole susceptibility profile (Dhieb et al, 2015) but no resistance nor susceptibility peaks has yet been identified.…”

The Continuing Threat of Methicillin-Resistant Staphylococcus aureus

“…Protein A typing ( spa typing) is an inexpensive method, which is based on the sequence analysis of variable number tandem repeats (VNTRs) in the encoding gene of this protein ( spa ), while also taking the number of repeat variations and point mutations into account (e.g., t011, t899) [118,119,120]. As a novelty, some studies have reported in the use of MALDI-TOF MS for the rapid discrimination of epidemic clones of MRSA, based on the association of their measured protein peaks (for an excellent review on the background of mass spectrometry, see [121]) and their spa types [118,122]. Whole-genome sequencing (WGS) is currently considered to be the gold standard as a molecular typing tool for the investigation of MRSA outbreaks [123].…”

Overview of phenotypic methods used for differentiation ofStaphylococcus aureus

“…It is based on examining the protein profile of the isolate, and it creates a 'fingerprint' unique for a given species. Furthermore, it is simple to execute and is automated [27][28][29][30]. Moreover, its use significantly shortens the time of identification of the tested microorganism -after bacteria isolation and preparation of sample, only a few minutes are needed for examination [27,29].…”

“…The minimum number of bacterial cells necessary for correct identification of a species is about 10 5 [27]. Additionally, it was noticed that the concentration of bacterial cells in the sample significantly influences the efficiency of the method [28].…”