Due to its ability to form biofilms on medical devices, Staphylococcus epidermidis has emerged as a major pathogen of nosocomial infections. In this study, we investigated the role of the two-component signal transduction system ArlRS in regulating S. epidermidis biofilm formation. An ArlRS-deficient mutant, WW06, was constructed using S. epidermidis strain 1457 as a parental strain. Although the growth curve of WW06 was similar to that of SE1457, the mutant strain was unable to form biofilms in vitro . In a rabbit subcutaneous infection model, sterile disks made of polymeric materials were implanted subcutaneously followed with inoculation of WW06 or SE1457. The viable bacteria cells of WW06 recovered from biofilms on the embedded disks were much lower than that of SE1457. Complementation of arlRS genes expression from plasmid in WW06 restored biofilm-forming phenotype both in vivo and in vitro . WW06 maintained the ability to undergo initial attachment. Transcription levels of several genes involved in biofilm formation, including icaADBC , sigB , and sarA , were decreased in WW06, compared to SE1457; and icaR expression was increased in WW06, detected by real-time reverse-transcription PCR. The biofilm-forming phenotype was restored by overexpressing icaADBC in WW06 but not by overexpressing sigB , indicating that ArlRS regulates biofilm formation through the regulation of icaADBC . Gel shift assay showed that ArlR can bind to the promoter region of the ica operon. In conclusion, ArlRS regulates S. epidermidis biofilm formation in an ica -dependent manner, distinct from its role in S. aureus .

Staphylococcus aureus is a common pathogen in chronic rhinosinusitis (CRS) patients, the pathogenesis of which involves the ability to form biofilms and produce various virulence factors. Tobacco smoke, another risk factor of CRS, facilitates S. aureus biofilm formation; however, the mechanisms involved are unclear. Here, we studied the effect of nicotine on S. aureus biofilm formation and the expression of virulence-related genes. S. aureus strains isolated from CRS patients and a USA300 strain were treated with nicotine or were untreated (control). Nicotine-treated S. aureus strains showed dose-dependent increases in biofilm formation, lower virulence, enhanced initial attachment, increased extracellular DNA release, and a higher autolysis rate, involving dysregulation of the accessory gene regulator (Agr) quorum-sensing system. Consequently, the expression of autolysis-related genes lytN and atlA, and the percentage of dead cells in biofilms was increased. However, the expression of virulence-related genes, including hla, hlb, pvl, nuc, ssp, spa, sigB, coa, and crtN was downregulated and there was reduced bacterial invasion of A549 human alveolar epithelial cells. The results of this study indicate that nicotine treatment enhances S. aureus biofilm formation by promoting initial attachment and extracellular DNA release but inhibits the virulence of this bacterium.

Methicillin-resistant Staphylococcus aureus (MRSA), also known as oxacillin-resistant S. aureus, is a leading cause of community and hospital associated infections globally. In this work, we found that deletion of the arlRS two-component system genes in the USA300 and USA500 strains resulted in increased susceptibilities to oxacillin (8–16-fold decrease in minimal inhibitory concentrations). In USA300Δ arlRS, transcriptional levels of mecA or blaZ showed no obvious change, while mRNA levels of spx showed a 4-fold decrease at 4 h and a 6.3-fold decrease at 10 h. Overexpression of spx in Δ arlRS restored oxacillin resistance to a similar level in USA300. In addition, gel shift assay showed that the recombinant ArlR bound to spx promoter region. Furthermore, silencing of spx led to a significant increase of oxacillin susceptibility in multiple MRSA isolates. Our results indicate that ArlRS plays a strong role in regulating oxacillin resistance in MRSA strains, which involves direct modulation of spx expression. Moreover, oritavancin showed inhibition to ATPase activity of the recombinant histidine kinase ArlS (IC 50 = 5.47 μM). Oritavancin had synergy effect on oxacillin activity against the MRSA strains in both planktonic and biofilm state. Our data suggest that ArlRS is an attractive target for breaking antimicrobial resistance of MRSA.

Epigallocatechin gallate (EGCG) is a main constituent of green tea polyphenols that are widely used as food preservatives and are considered to be safe for consumption. However, the underlying antimicrobial mechanism of EGCG and the bacterial response to EGCG are not clearly understood. In the present study, a genome-wide transcriptional analysis of a typical spoilage bacterium, Pseudomonas fluorescens that responded to EGCG was performed using RNA-seq technology. A total of 26,365,414 and 23,287,092 clean reads were generated from P. fluorescens treated with or without 1 mM EGCG and the clean reads were aligned to the reference genome. Differential expression analysis revealed 291 upregulated genes and 134 downregulated genes and the differentially expressed genes (DEGs) were verified using RT-qPCR. Most of the DGEs involved in iron uptake, antioxidation, DNA repair, efflux system, cell envelope and cell-surface component synthesis were significantly upregulated by EGCG treatment, while most genes associated with energy production were downregulated. These transcriptomic changes are likely to be adaptive responses of P. fluorescens to iron limitation and oxidative stress, as well as DNA and envelope damage caused by EGCG. The expression of specific genes encoding the extra-cytoplasmic function sigma factor (PvdS, RpoE and AlgU) and the two-component sensor histidine kinase (BaeS and RpfG) were markedly changed by EGCG treatment, which may play important roles in regulating the stress responses of P. fluorescens to EGCG. The present data provides important insights into the molecular action of EGCG and the possible cross-resistance mediated by EGCG on P. fluorescens, which may ultimately contribute to the optimal application of green tea polyphenols in food preservation.

Soil alkalinity is a major abiotic constraint to crop productivity and quality. Wild soybean (Glycine soja) is considered to be more stress-tolerant than cultivated soybean (G. max), and has considerable genetic variation for increasing alkalinity tolerance of soybean. In this study, we analyzed the transcriptome profile in the roots of an alkalinity tolerant wild soybean variety N24852 at 12 and 24 h after 90 mM NaHCO3 stress by RNA-sequencing. Compared with the controls, a total of 449 differentially expressed genes (DEGs) were identified, including 95 and 140 up-regulated genes, and 108 and 135 down-regulated genes at 12 and 24 h after NaHCO3 treatment, respectively. Quantitative RT-PCR analysis of 14 DEGs showed a high consistency with their expression profiles by RNA-sequencing. Gene Ontology (GO) terms related to transcription factors and transporters were significantly enriched in the up-regulated genes at 12 and 24 h after NaHCO3 stress, respectively. Nuclear factor Y subunit A transcription factors were enriched at 12 h after NaHCO3 stress, and high percentages of basic helix-loop-helix, ethylene-responsive factor, Trihelix, and zinc finger (C2H2, C3H) transcription factors were found at both 12 and 24 h after NaHCO3 stress. Genes related to ion transporters such as ABC transporter, aluminum activated malate transporter, glutamate receptor, nitrate transporter/proton dependent oligopeptide family, and S-type anion channel were enriched in up-regulated DEGs at 24 h after NaHCO3 treatment, implying their roles in maintaining ion homeostasis in soybean roots under alkalinity. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis showed “phenylpropanoid biosynthesis” and “phenylalanine metabolism” pathways might participate in soybean response to alkalinity. This study provides a foundation to further investigate the functions of NaHCO3 stress-responsive genes and the molecular basis of soybean tolerance to alkalinity.