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Browsing by Author "Petruzzi, Briana"

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    Capsular Polysaccharide Interferes with Biofilm Formation by Pasteurella multocida Serogroup A
    Petruzzi, Briana; Briggs, Robert E.; Swords, W. Edward; De Castro, Cristina; Molinaro, Antonio; Inzana, Thomas J. (American Society for Microbiology, 2017-11)
    Pasteurella multocida is an important multihost animal and zoonotic pathogen that is capable of causing respiratory and multisystemic diseases, bacteremia, and bite wound infections. The glycosaminoglycan capsule of P. multocida is an essential virulence factor that protects the bacterium from host defenses. However, chronic infections (such as swine atrophic rhinitis and the carrier state in birds and other animals) may be associated with biofilm formation, which has not been characterized in P. multocida. Biofilm formation by clinical isolates was inversely related to capsule production and was confirmed with capsule-deficient mutants of highly encapsulated strains. Capsule-deficient mutants formed biofilms with a larger biomass that was thicker and smoother than the biofilm of encapsulated strains. Passage of a highly encapsulated, poor-biofilm-forming strain under conditions that favored biofilm formation resulted in the production of less capsular polysaccharide and a more robust biofilm, as did addition of hyaluronidase to the growth medium of all of the strains tested. The matrix material of the biofilm was composed predominately of a glycogen exopolysaccharide (EPS), as determined by gas chromatography-mass spectrometry, nuclear magnetic resonance, and enzymatic digestion. However, a putative glycogen synthesis locus was not differentially regulated when the bacteria were grown as a biofilm or planktonically, as determined by quantitative reverse transcriptase PCR. Therefore, the negatively charged capsule may interfere with biofilm formation by blocking adherence to a surface or by preventing the EPS matrix from encasing large numbers of bacterial cells. This is the first detailed description of biofilm formation and a glycogen EPS by P. multocida. IMPORTANCE Pasteurella multocida is an important pathogen responsible for severe infections in food animals, domestic and wild birds, pet animals, and humans. P. multocida was first isolated by Louis Pasteur in 1880 and has been studied for over 130 years. However, aspects of its lifecycle have remained unknown. Although formation of a biofilm by P. multocida has been proposed, this report is the first to characterize biofilm formation by P. multocida. Of particular interest is that the biofilm matrix material contained a newly reported amylose-like glycogen as the exopolysaccharide component and that production of capsular polysaccharide (CPS) was inversely related to biofilm formation. However, even highly mucoid, poor-biofilm-forming strains could form abundant biofilms by loss of CPS or following in vitro passage under biofilm growth conditions. Therefore, the carrier state or subclinical chronic infections with P. multocida may result from CPS downregulation with concomitant enhanced biofilm formation.
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    Polymicrobial Biofilm Interaction Between Histophilus somni and Pasteurella multocida
    Petruzzi, Briana; Dickerman, Allan W.; Lahmers, Kevin K.; Scarratt, William K.; Inzana, Thomas J. (2020-07-10)
    Histophilus somni and Pasteurella multocida are two of multiple agents responsible for bovine respiratory disease (BRD) in cattle. Following respiratory infection of calves withH. somni,P. multocidamay also be isolated from the lower respiratory tract. BecauseH. somnimay form a biofilm during BRD, we sought to determine ifP. multocidacan co-exist withH. somniin a polymicrobial biofilmin vitroandin vivo. Interactions between the two species in the biofilm were characterized and quantified by fluorescencein situhybridization (FISH). The biofilm matrix of each species was examined using fluorescently tagged lectins (FTL) specific for the exopolysaccharide (EPS) using confocal laser scanning microscopy. Bacterial interactions were determined by auto-aggregation and biofilm morphology.Pasteurella multocidaandH. somniwere evenly distributed in thein vitrobiofilm, and both species contributed to the polymicrobial biofilm matrix. The average biomass and biofilm thickness, and the total carbohydrate and protein content of the biofilm, were greatest when both species were present. Polymicrobial bacterial suspensions auto-aggregated faster than single species suspensions, suggesting physical interactions between the two species. Almost 300P. multocidagenes were significantly differentially regulated when the bacteria were in a polymicrobial biofilm compared to a mono-species biofilm, as determined by RNA-sequencing. As expected, host genes associated with inflammation and immune response were significantly upregulated at the infection site followingH. somnichallenge. EncapsulatedP. multocidaisolates not capable of forming a substantial biofilm enhanced anin vitropolymicrobial biofilm withH. somni, indicating they contributed to the polymicrobial biofilm matrix. Indirect evidence indicated that encapsulatedP. multocidaalso contributed to a polymicrobial biofilmin vivo. Only the EPS ofH. somnicould be detected by FTL staining of bovine tissues following challenge withH. somni. However, both species were isolated and an immune response to the biofilm matrix of both species was greater than the response to planktonic cells, suggesting encapsulatedP. multocidamay take advantage of theH. somnibiofilm to persist in the host during chronic BRD. These results may have important implications for the management and prevention of BRD.