Bordetella bronchiseptica Diguanylate Cyclase BdcA Regulates Motility and Is Important for the Establishment of Respiratory Infection in Mice

Abstract

Bacteria can be motile and planktonic or, alternatively, sessile and participating in the biofilm mode of growth. The transition between these lifestyles can be regulated by a second messenger, cyclic dimeric GMP (c-di-GMP). High intracellular c-di-GMP concentration correlates with biofilm formation and motility inhibition in most bacteria, including Bordetella bronchiseptica, which causes respiratory tract infections in mammals and forms biofilms in infected mice. We previously described the diguanylate cyclase BdcA as involved in c-di-GMP synthesis and motility regulation in B. bronchiseptica; here, we further describe the mechanism whereby BdcA is able to regulate motility and biofilm formation. Amino acid replacement of GGDEF with GGAAF in BdcA is consistent with the conclusion that diguanylate cyclase activity is necessary for biofilm formation and motility regulation, although we were unable to confirm the stability of the mutant protein. In the absence of the bdcA gene, B. bronchiseptica showed enhanced motility, strengthening the hypothesis that BdcA regulates motility in B. bronchiseptica. We showed that c-di-GMP-mediated motility inhibition involved regulation of flagellin expression, as high c-di-GMP levels achieved by expressing BdcA significantly reduced the level of flagellin protein. We also demonstrated that protein BB2109 is necessary for BdcA activity, motility inhibition, and biofilm formation. Finally, absence of the bdcA gene affected bacterial infection, implicating BdcA-regulated functions as important for bacterium-host interactions. This work supports the role of c-di-GMP in biofilm formation and motility regulation in B. bronchiseptica, as well as its impact on pathogenesis. IMPORTANCE Pathogenesis of Bordetella spp., like that of a number of other pathogens, involves biofilm formation. Biofilms increase tolerance to biotic and abiotic factors and are proposed as reservoirs of microbes for transmission to other organs (trachea, lungs) or other hosts. Bis-(3′-5′)-cyclic dimeric GMP (c-di-GMP) is a second messenger that regulates transition between biofilm and planktonic lifestyles. In Bordetella bronchiseptica, high c-di-GMP levels inhibit motility and favor biofilm formation. In the present work, we characterized a B. bronchiseptica diguanylate cyclase, BdcA, which regulates motility and biofilm formation and affects the ability of B. bronchiseptica to colonize the murine respiratory tract. These results provide us with a better understanding of how B. bronchiseptica can infect a host.

Bordetella spp. utilize the type 3 secretion system to manipulate the VIP/VPAC2 signaling and promote colonization and persistence of the three classical Bordetella in the lower respiratory tract

“…Measuring bacterial burden at day 7 post-infection is defined as the peak of Bordetella spp. infection, and it also supported by previously published methods used to assess the minimal required dose of bacteria for successful infection (Belhart et al, 2019). Thus, from these values, 50% of the bacterial inoculum required to efficiently infect the mice used for these experiments (also referred to as ID 50 ) was calculated.…”

“…utilize VIP/VPAC axis to promote intracellular survival in macrophages ( Askar et al., 2020 ), suggesting that VIP/VPAC axis might influence infection dynamics and persistence. C57BL/6J and VPAC2 -/- mice were intranasally inoculated with increasing bacterial concentrations of B. pertussis, B. parapertussis , and B. bronchiseptica , and euthanized at day 7 post-infection to enumerate colonies from the respiratory tract ( Belhart et al., 2019 ).…”

“…Similarly, B. parapertussis colonized C57BL/6J ( Weyrich et al., 2012 ) at a lower infectious dose than the VPAC2 -/- mice ( Figure 3B ). Finally, very low doses of B. bronchiseptica RB50 were required to colonize the C57BL/6J lungs efficiently ( Weyrich et al., 2012 ; Belhart et al., 2019 ; Gestal et al., 2019 ) ( Figure 3C ). However, at least 1 x 10 4 CFUs of RB50 were required to colonize the lungs of VPAC2 -/- mice.…”

Bordetella bronchiseptica diguanylate cyclase BdcB inhibits the type three secretion system and impacts the immune response

“…However, during these conditions, the flagellar apparatus is expressed, and the bacteria exhibit motility 9 . We have previously described that the second messenger bis-(3′-5′)-cyclic dimeric guanosine monophosphate (c-di-GMP) regulates biofilm and motility in B. bronchiseptica 10 , 11 . C-di-GMP is extensively associated with the transition between planktonic and biofilm lifestyles in bacteria; when c-di-GMP levels are high a biofilm-forming phenotype is present in most bacteria 12 .…”

Outer Membrane Vesicles Coating Nano-Glycyrrhizic Acid Confers Protection Against Borderella bronchiseptica Through Th1/Th2/Th17 Responses

“…Herein, we chose Bordetella bronchiseptica (Bb) as a model pathogen, which is the cause of infectious respiratory disease in dogs, atrophic rhinitis in pigs, snuffles in rabbits, and pneumonia in immuno-compromised individuals. 14 , 15 In addition, the effects of GAN-OMV were preliminarily investigated on the immunological function of macrophages and compared to that of the single OMV. Following the in vitro experiments, GAN-OMV was subcutaneously injected into mice, which subsequently elicited a mixed Th1/Th2/Th17 response in the vaccinated animals.…”