Streptococcus suis (S. suis) and Haemophilus parasuis (H. parasuis) are two primary pathogens currently affecting the porcine industry. They often cause encephalitis and arthritis. They also frequently co-infect in clinical settings. In the current study, we identified significant correlations between S. suis and H. parasuis. The results from CI versus RIR suggested that S. suis and H. parasuis were competitive in general. Compared to mono-species biofilm, the biomass, bio-volume, and thickness of mixed-species biofilms were significantly higher, which was confirmed using crystal violet staining, confocal laser scanning microscopy, and scanning electron microscopy. Compared to mono-species biofilm, the viable bacteria in the mixed-species biofilms were significantly lower, which was confirmed using the enumeration of colony-forming units (CFU cm−2). The susceptibility of antibiotics in the co-culture decreased in the planktonic state. In contrast, biofilm state bacteria are significantly more difficult to eradicate with antibiotics than in a planktonic state. Whether in planktonic or biofilm state, the expression of virulence genes of S. suis and H. parasuis in mixed culture was very different from that in single culture. Subsequently, by establishing a mixed infection model in mice, we found that the colonization of the two pathogens in organs increased after mixed infection, and altered the host’s inflammatory response. In summary, our results indicate that S. suis and H. parasuis compete when co-cultured in vitro. Surprisingly, S. suis and H. parasuis synergistically increased colonization capacity after co-infection in vivo. This study elucidated the interaction between S. suis and H. parasuis during single infections and co-infections. Future studies on bacterial disease control and antibiotic treatment should consider the interaction of mixed species.

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Streptococcus suis LuxS/AI-2 quorum sensing system regulates biofilm formation, resulting in increased pathogenicity and drug resistance, and diminished efficacy of antibiotic treatment. The remaining peony seed cake after oil extraction is rich in monoterpenoid glycosides, which can inhibit the formation of bacterial biofilm. In this study, we investigated the effect of seven major monocomponents (suffruticosol A, suffruticosol B, suffruticosol C, paeonifloin, albiflorin, trans-ε-viniferin, gnetin H) of peony seed meal on minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of S. suis. The results showed that the MICs of the seven single components were all greater than 200 μg/mL, with no significant bacteriostatic and bactericidal advantages. Crystal violet staining and scanning electron microscope observation showed that the seven single components had a certain inhibitory effect on the biofilm formation ability of S. suis at sub-MIC concentration. Among them, the ability of paeoniflorin to inhibit biofilm was significantly higher than that of the other six single components. AI-2 signaling molecules were detected by bioreporter strain Vibrio harvey BB170. The detection results of AI-2 signal molecules found that at 1/2 MIC concentration, paeoniflorin significantly inhibited the production of S. suis AI-2 signal, and the inhibitory effect was better than that of the other six single components. In addition, molecular docking analysis revealed that paeoniflorin had a significant binding activity with LuxS protein compared with the other six single components. The present study provides evidence that paeoniflorin plays a key role in the regulation of the inhibition of S. suis LuxS/AI-2 system and biofilm formation in peony seed meal.

The multidrug-resistant Salmonella is becoming major health threat due to inappropriate use of antibiotics. It is found that the metabolic changes contributed to bacterial drug resistance. AI-2 quorum sensing (QS) is involved in the regulation of bacterial metabolic activities. However, the role of AI-2 quorum sensing in the metabolic activities and drug resistance of Salmonella remain unclear. In this study, we determined the drug sensitivity of Salmonella typhimurium by 96-well plate and then determined the time-killing curve. Then, the regulation of AI-2 quorum sensing on Salmonella metabolic activity was analyzed by quantitative real‑time PCR and LC-MS metabonomics. The results showed that luxS mutant strain of Salmonella typhimurium lacking AI-2 quorum sensing was at least 4-fold less sensitive to killing by fluoroquinolone antibiotics. The decreased cell killing after the fluoroquinolones challenge was found to involve a low metabolic state presented by up-regulating anaerobic respiration, and LC-MS metabonomics analysis showed that the low metabolic state of luxS deletion strain might be related to the metabolism of amino acids such as leucine and valine. In conclusion, this study indicated that the deletion of luxS gene in Salmonella typhimurium mediated the tolerance to fluoroquinolones by affecting valine metabolism and up-regulating the anaerobic metabolism pathway.

Background: Streptococcus suis is a zoonotic pathogen closely related to the respiratory tract of swine. Paeonol is a traditional Chinese medicine with antibacterial effect. This study demonstrated the effect of paeonol on biofilm formation and the expression of virulence factors of Streptococcus suis. Methods: The minimum inhibitory concentration (MIC) of paeonol on Streptococcus suis was determined by double dilution method. Crystal violet staining method was used to determine the effect of paeonol on the biofilm formation. The effects of paeonol on the morphology and structure of Streptococcus suis biofilm were observed by field emission scanning electron microscopy (FE-SEM). Real-time quantitative PCR (qRT-PCR) was used to detect the expression levels of Streptococcus suis related virulence factors under sub-MIC (1/4 MIC, 1/8 MIC) paeonol. Result: The results showed the effect of 1/4 MIC and 1/8 MIC paeonol on biofilm formation and mature biofilm was significant (*P less than 0.05). Under the action of paeonol, the biofilm structure becomes sparse. qRT-PCR results showed that 1/8 MIC paeonol inhibits bacterial adhesion by up-regulating the expression of mrp virulence genes, thereby inhibiting the formation of Streptococcus suis biofilm.

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