Oral commensal streptococci are primary colonizers of the oral cavity. These streptococci produce many adhesins, metabolites, and antimicrobials that modulate microbial succession and diversity within the oral cavity.

Pseudomonas aeruginosa dominates the complex polymicrobial cystic fibrosis (CF) airway and is a leading cause of death in persons with CF. Oral streptococcal colonization has been associated with stable CF lung function. However, no studies have demonstrated how Streptococcus salivarius, the most abundant streptococcal species found in individuals with stable CF lung disease, potentially improves lung function or becomes incorporated into the CF airway biofilm. By utilizing a two-species biofilm model to probe interactions between S. salivarius and P. aeruginosa, we discovered that the P. aeruginosa exopolysaccharide Psl promoted S. salivarius biofilm formation. Further, we identified a S. salivarius maltose-binding protein (MalE) that is required for promotion of biofilm formation both in vitro and in a Drosophila melanogaster co-infection model. Finally, we demonstrate that promotion of dual biofilm formation with S. salivarius is common among environmental and clinical P. aeruginosa isolates. Overall, our data supports a model in which S. salivarius uses a sugar-binding protein to interact with P. aeruginosa exopolysaccharide, which may be a strategy by which S. salivarius establishes itself within the CF airway microbial community.

Commensal streptococci regulate health and homeostasis within oral polymicrobial communities. Remarkably, high salivary nitrite concentrations have also been associated with improved health in the oral cavity. We previously demonstrated that nitrite assists hydrogen peroxide-producing oral commensal streptococci in regulating homeostasis via the generation of reactive nitrogen species (RNS), which have antimicrobial activity on oral pathogens. However, it is unknown how nitrite and commensal streptococci work in concert to influence the metabolome of oral polymicrobial communities. In this study, we report that nitrite aids commensal streptococci in the inhibition of multi-kingdom pathogens that reside in distinct oral niches, which supports commensal dominance. More importantly, we show that commensal streptococci utilize nitrite to drive the metabolic signature of multispecies biofilms in a manner that supports commensal metabolism and resistance to RNS, and restricts metabolic processes that are required for pathogen virulence. Taken together, our study provides insight into how commensal streptococci use nitrite to trigger shifts in the oral polymicrobial metabolome to support health and homeostasis.

Voltage-dependent L-type Ca2+ channels (L-VDCCs) and the RhoA/Rho kinase pathway are two predominant intracellular signaling pathways that regulate renal microvascular reactivity. Traditionally, these two pathways have been thought to act independently; however, recent evidence suggests that these pathways could be convergent. We hypothesized that Rho kinase inhibitors can influence L-VDCC signaling. The effects of Rho kinase inhibitors Y-27632 or RKI-1447 on KCl-induced depolarization or the L-VDCC agonist Bay K8644 were assessed in afferent arterioles using an in vitro blood-perfused rat juxtamedullary nephron preparation. Superfusion of KCl (30–90 mM) led to concentration-dependent vasoconstriction of afferent arterioles. Administration of Y-27632 (1, 5, and 10 µM) or RKI-1447 (0.1, 1, and 10 µM) significantly increased the starting diameter by 16–65%. KCl-induced vasoconstriction was markedly attenuated with 5 and 10 µM Y-27632 and with 10 µM RKI-1447 ( P < 0.05 vs. KCl alone). Y-27632 (5 µM) also significantly attenuated Bay K8644-induced vasoconstriction ( P < 0.05). Changes in intracellular Ca2+ concentration ([Ca2+]i) were estimated by fura-2 fluorescence during KCl-induced depolarization in cultured A7r5 cells and in freshly isolated preglomerular microvascular smooth muscle cells. Administration of 90 mM KCl significantly increased fura-2 fluorescence in both cell types. KCl-mediated elevation of [Ca2+]i in A7r5 cells was suppressed by 1–10 µM Y-27632 ( P < 0.05), but 10 µM Y-27632 was required to suppress Ca2+ responses in preglomerular microvascular smooth muscle cells. RKI-1447, however, significantly attenuated KCl-mediated elevation of [Ca2+]i. Y-27632 markedly inhibited Bay K8644-induced elevation of [Ca2+]i in both cell types. The results of the present study indicate that the Rho kinase inhibitors Y-27632 and RKI-1447 can partially inhibit L-VDCC function and participate in L-VDCC signaling.

Chronic infections in the cystic fibrosis (CF) airway are composed of both pathogenic and commensal bacteria. However, chronic Pseudomonas aeruginosa infections are the leading cause of lung deterioration in individuals with CF. Interestingly, oral commensals can translocate to the CF lung and their presence is associated with improved lung function, presumably due to their ability to antagonize P. aeruginosa. We have previously shown that one commensal, Streptococcus parasanguinis, produces hydrogen peroxide that reacts with nitrite to generate reactive nitrogen intermediates (RNI) which inhibit P. aeruginosa growth. In this study, we sought to understand the global impact of commensal-mediated RNI on the P. aeruginosa transcriptome. RNA sequencing analysis revealed that S. parasanguinis and nitrite-mediated RNI dysregulated expression of denitrification genes in a CF isolate of P. aeruginosa compared to when this isolate was only exposed to S. parasanguinis. Further, loss of a nitric oxide reductase subunit (norB) rendered an acute P. aeruginosa isolate more susceptible to S. parasanguinis-mediated RNI. Additionally, S. parasanguinis-mediated RNI inactivated P. aeruginosa aconitase activity. Lastly, we report that P. aeruginosa isolates recovered from CF individuals are uniquely hypersensitive to S. parasanguinis-mediated RNI compared to acute infection or environmental P. aeruginosa isolates. These findings illustrate that S. parasanguinis hinders the ability of P. aeruginosa to respond to RNI, which potentially prevents P. aeruginosa CF isolates from resisting commensal and host-induced RNI in the CF airway.