Membrane Vesicles Are the Dominant Structural Components of Ceftazidime-Induced Biofilm Formation in an Oxacillin-Sensitive MRSA.

Xinlong He,Yuzheng Huang,Weijuan Gong,Jiahui Xu,Yinyan Yin,Feng Lu,Jun Cao,Li Qian,Tingting Guo,Yi Yin,Guocai Li,Xiaoqin He,Shuang Li

doi:10.3389/fmicb.2019.00571

Abstract

Methicillin-resistant Staphylococcus aureus (MRSA) has received increasing attention in recent years. However, the characteristics and relevant mechanisms of biofilm formation in oxacillin-sensitive MRSA (OS-MRSA) are poorly understood. This study was designed to characterize biofilm formation in OS-MRSA BWSA15 in response to ceftazidime (TZ) by comparing the methicillin-sensitive S. aureus (MSSA) strain BWSA23 and the oxacillin-resistant MRSA (OR-MRSA) strain BWSA11. The biofilms and biofilm-forming cells were observed by electron microscopy. Biofilms grown on microtiter plates were chemically decomposed and analyzed by Fourier transform infrared spectroscopy. The transcriptional regulation of genes associated with methicillin resistance, surface adhesion, fatty acid biosynthesis, and global regulation (sigma B) was investigated. A significant increase in biofilm formation ability (10.21-fold) and aggregation ability (2.56-fold) was observed in BWSA15 upon the treatment with TZ (16 μg/ml). The TZ-induced biofilm formation in BWSA15 was characterized by a disappearance of polysaccharide-like extracellular substances and an appearance of a large number of intercellular MVs from extracellular matrix. Few MVs were identified in the biofilms formed by BWSA11 and BWSA23. There was a significant upregulation of mecA, sigB, and fatty acid biosynthesis-associated genes and downregulation of icaA, icaD, clfA, clfB, and fnaA in BWSA15 upon the treatment with TZ. The formation of intracellular junctions of MVs in the biofilms of BWSA15 was mediated by a significant increase in the proportion of proteins as well as by an increase in the proportion of non-ionized carboxyl groups in fatty acids. This study demonstrated that beta-lactam antibiotics can induce biofilm formation in OS-MRSA, and the biofilm induction in OS-MRSA can mainly be attributed to exposed MVs with increased hydrophobicity rather than polysaccharide intercellular adhesins, cell wall-anchored surface proteins, and extracellular DNA.

Highlights

MATERIALS AND METHODSStaphylococcus aureus, a pathogenic bacteria, can cause various infections, including skin and soft tissue infections, endocarditis, pneumonia, and even septicemia (Tong et al, 2015)
Since to ceftazidime (TZ) did not trigger significant increase in biofilm forming ability in BWSA 11 and BWSA 23, the enhancement of the biofilm formation phenotype observed in BWSA15 upon treatment with a subinhibitory concentration of TZ is mainly attributed to increased aggregation ability caused by biological response rather than simple physical interaction during cell wall biosynthesis
We found that the differences in the transcriptional expression of clumping factor A (clfA), clumping factor B (clfB), and fnbA between BWSA11 and BWSA15 in the absence of antibiotic stress were significantly correlated with the differences in biofilm formation by these strains, suggesting the involvement of mecA in the regulation of the native expression of the surface adhesin proteins during biofilm formation by methicillin-resistant S. aureus (MRSA) strains

Summary

Introduction

MATERIALS AND METHODSStaphylococcus aureus, a pathogenic bacteria, can cause various infections, including skin and soft tissue infections, endocarditis, pneumonia, and even septicemia (Tong et al, 2015). MRSA is one of the most common pathogens that causes nosocomial infections worldwide (den Heijer et al, 2013; Udobi et al, 2013; Chen and Huang, 2014) Among these clinical MRSA strains, oxacillinsensitive MRSA (OS-MRSA) has received increasing attention due to the latent but highly inducible methicillin resistance exhibited by this strain (Hososaka et al, 2007; Pu et al, 2014; Chung et al, 2016). Microbial surface components recognizing adhesive matrix molecules (MSCRAMM) are commonly found in S. aureus strains (Atshan et al, 2012b) Among these MSCRAMMs, polysaccharide intercellular adhesins (PIAs) or polymeric N-acetyl-glucosamine (PNAG) and surface protein adhesins are considered to be important substances that contribute to the surface adhesion of MSSA and MRSA (Pozzi et al, 2012; McCarthy et al, 2015), respectively.

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