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Thomas J. Inzana - One of the best experts on this subject based on the ideXlab platform.

  • The Role of luxS in Histophilus somni Virulence and Biofilm Formation.
    Infection and immunity, 2021
    Co-Authors: Yu Pan, Aloka B. Bandara, Indra Sandal, Shivakumara Siddaramappa, Allan Dickerman, Thomas J. Inzana
    Abstract:

    ABSTRACT S-Ribosylhomocysteinase (LuxS) is required for the synthesis of the autoinducer-2 (AI-2) quorum-sensing signaling molecule in many Gram-negative bacteria. The bovine (and ovine) opportunistic pathogen Histophilus somni contains luxS and forms a biofilm containing an exopolysaccharide (EPS) in the matrix. Since biofilm formation is regulated by quorum sensing in many bacteria, the roles of luxS in H. somni virulence and biofilm formation were investigated. Although culture supernatants from H. somni were ineffective at inducing bioluminescence in the Vibrio harveyi reporter strain BB170, H. somniluxS complemented the biosynthesis of AI-2 in the luxS-deficient Escherichia coli strain DH5α. H. somni strain 2336 luxS was inactivated by transposon mutagenesis. RNA expression profiles revealed that many genes were significantly differentially expressed in the luxS mutant compared to that in the wild-type, whether the bacteria were grown planktonically or in a biofilm. Furthermore, the luxS mutant had a truncated and asialylated lipooligosaccharide (LOS) and was substantially more serum sensitive than the wild-type. Not surprisingly, the luxS mutant was attenuated in a mouse model for H. somni virulence, and some of the altered phenotypes were partially restored after the mutation was complemented with a functional luxS. However, no major differences were observed between the wild-type and the luxS mutant in regard to outer membrane protein profiles, biofilm formation, EPS production, or intracellular survival. These results indicate that luxS plays a role in H. somni virulence in the context of LOS biosynthesis but not biofilm formation or other phenotypic properties examined.

  • The role of luxS in Histophilus somni virulence and biofilm formation.
    Infection and immunity, 2021
    Co-Authors: Yu Pan, Aloka B. Bandara, Indra Sandal, Shivakumara Siddaramappa, Allan Dickerman, Thomas J. Inzana
    Abstract:

    S-ribosylhomocysteinase (LuxS) is required for the synthesis of the autoinducer-2 (AI-2) quorum-sensing signalling molecule in many Gram-negative bacteria. The bovine (and ovine) opportunistic pathogen Histophilus somni contains a luxS, and forms a biofilm containing an exopolysaccharide (EPS) in the matrix. Since biofilm formation is regulated by quorum sensing in many bacteria, the role of luxS in H. somni virulence and biofilm formation was investigated. Although culture supernatants from H. somni were ineffective at inducing bioluminescence in the Vibrio harveyi reporter strain BB170, H. somni luxS complemented the biosynthesis of AI-2 in the luxS-deficient Escherichia coli strain DH5α. H. somni strain 2336 luxS was inactivated by tranposon mutagenesis. Comparison of RNA exression profiles revealed that many genes were significantly differentially expressed in the luxS mutant compared to the wildtype, whether the bacteria were grown planktonically or in a biofilm. Furthermore, the luxS mutant had a truncated and asialylated lipooligosaccharide (LOS), and was substantially more serum-sensitive than the wildtype. Not surprisingly, the luxS mutant was attenuated in a mouse model for H. somni virulence, and some of the altered phenotypes were partially restored after the mutation was complemented with a functional luxS However, no major differences were observed between the wildtype and the luxS mutant in regard to outer membrane protein profiles, biofilm formation, EPS production, or intracellular survival. These results indicate that luxS plays a role in H. somni virulence in the context of LOS biosynthesis, but not biofilm formation or other phenotypic properties examined.

  • Polymicrobial Biofilm Interaction Between Histophilus somni and Pasteurella multocida.
    Frontiers in microbiology, 2020
    Co-Authors: Briana Petruzzi, Allan Dickerman, Kevin K. Lahmers, William K. Scarratt, Thomas J. Inzana
    Abstract:

    Histophilus somni and Pasteurella multocida are two of multiple agents responsible for bovine respiratory disease (BRD) in cattle. Following respiratory infection of calves with H. somni, P. multocida may also be isolated from the lower respiratory tract. Because H. somni may form a biofilm during BRD, we sought to determine if P. multocida can co-exist with H. somni in a polymicrobial biofilm in vitro and in vivo. Interactions between the two species in the biofilm were characterized and quantified by fluorescence in situ hybridization (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 multocida and H. somni were evenly distributed in the in vitro biofilm, 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 300 P. multocida genes 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 following H. somni challenge. Encapsulated P. multocida isolates not capable of forming a substantial biofilm enhanced an in vitro polymicrobial biofilm with H. somni, indicating they contributed to the polymicrobial biofilm matrix. Indirect evidence indicated that encapsulated P. multocida also contributed to a polymicrobial biofilm in vivo. Only the EPS of H. somni could be detected by FTL staining of bovine tissues following challenge with H. 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 encapsulated P. multocida may take advantage of the H. somni biofilm to persist in the host during chronic BRD. These results may have important implications for the management and prevention of BRD.

  • Identification of Histophilus somni by a nanomaterial optical fiber biosensor assay.
    Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians Inc, 2018
    Co-Authors: Aloka B. Bandara, Ziwei Zuo, Kelly Mccutcheon, Siddharth Ramachandran, James R. Heflin, Thomas J. Inzana
    Abstract:

    Histophilus somni is an opportunistic pathogen responsible for respiratory and systemic diseases of cattle and sheep. Rapid and accurate detection of H. somni is essential to distinguish H. somni f...

  • DS1_JVDI_10.1177_1040638718803665 – Supplemental material for Identification of Histophilus somni by a nanomaterial optical fiber biosensor assay
    2018
    Co-Authors: Aloka B. Bandara, Ziwei Zuo, Kelly Mccutcheon, Siddharth Ramachandran, James R. Heflin, Thomas J. Inzana
    Abstract:

    Supplemental material, DS1_JVDI_10.1177_1040638718803665 for Identification of Histophilus somni by a nanomaterial optical fiber biosensor assay by Aloka B. Bandara, Ziwei Zuo, Kelly McCutcheon, Siddharth Ramachandran, James R. Heflin and Thomas J. Inzana in Journal of Veterinary Diagnostic Investigation

Øystein Angen - One of the best experts on this subject based on the ideXlab platform.

  • Taxonomy of Histophilus somni.
    Current topics in microbiology and immunology, 2015
    Co-Authors: Øystein Angen
    Abstract:

    Histophilus somni was proposed in 2003 as a common name for bacteria that earlier had been called “Haemophilus somni”, “Haemophilus agni”, “Haemophilus somnifer”, and “Histophilus ovis”. The species is clearly separated from other species and genera within the family Pasteurellaceae. The species is phenotypically variable, but highly uniform regarding the 16S rDNA sequence. Whole-genome sequencing has revealed distinct genetic differences between a commensal and a pathogenic strain, particularly in regard to putative virulence factors. However, broad generalizations regarding the genetics of H. somni cannot be applied to the entire species until the genomes of additional strains are sequenced.

  • Antimicrobial susceptibility of Haemophilus parasuis and Histophilus somni from pigs and cattle in Denmark.
    Veterinary microbiology, 2004
    Co-Authors: Frank Møller Aarestrup, Anne Mette Seyfarth, Øystein Angen
    Abstract:

    Abstract A total of 52 Haemophilus parasuis and 80 Histophilus somni isolates were tested for antimicrobial susceptibility by MIC-determinations. None of the isolates were resistant to ampicillin, ceftiofur, ciprofloxacin, erythromycin, florphenicol, penicillin, spectinomycin, tetracycline, tiamulin, or tilmicosin. Two H. parasuis isolates were resistant to trimethoprim+sulfamethoxazole. Six H. parasuis isolates had reduced susceptibility (0.06–0.5 μg/ml) to ciprofloxacin and 10 reduced susceptibility to TMP+sulfamethoxazole (1–2 μg/ml). This study showed that Danish isolates of H. parasuis and H. somni in general are fully susceptible to antimicrobial agents currently used for treatment of infections with these pathogens.

  • proposal of Histophilus somni gen nov sp nov for the three species incertae sedis haemophilus somnus haemophilus agni and Histophilus ovis
    International Journal of Systematic and Evolutionary Microbiology, 2003
    Co-Authors: Øystein Angen, Peter Ahrens, Peter Kuhnert, Henrik Christensen, Reinier Mutters
    Abstract:

    Earlier investigations have shown that ‘Haemophilus somnus’, ‘Haemophilus agni’ and ‘Histophilus ovis’ represent the same species. In the present investigation, the taxonomic position of this species is explored further by sequencing the 16S rRNA and rpoB genes of strains that were investigated previously by DNA–DNA hybridization. These results clearly support the allocation of this species to a novel genus within the family Pasteurellaceae. The phenotypic separation of Histophilus somni gen. nov., sp. nov. from other members of the family can, for most strains, be based on capnophilia, yellowish pigmentation and indole production. However, due to phenotypic variation, the use of a species-specific PCR test based on the 16S rRNA gene is included in the species description. This is justified by the high sequence similarity of the 16S rRNA gene within the species and the fact that the highest sequence similarity to any other taxon within the family is 93·4 %. The type strain, 8025T=ATCC 43625T=CCUG 36157T, was isolated in the USA from a bovine brain with lesions of thromboembolic meningoencephalitis.

  • proposal of Histophilus somni gen nov sp nov for the three species incertae sedis haemophilus somnus haemophilus agni and Histophilus ovis
    International Journal of Systematic and Evolutionary Microbiology, 2003
    Co-Authors: Øystein Angen, Peter Ahrens, Peter Kuhnert, Henrik Christensen, Reinier Mutters
    Abstract:

    Earlier investigations have shown that 'Haemophilus somnus', 'Haemophilus agni' and 'Histophilus ovis' represent the same species. In the present investigation, the taxonomic position of this species is explored further by sequencing the 16S rRNA and rpoB genes of strains that were investigated previously by DNA-DNA hybridization. These results clearly support the allocation of this species to a novel genus within the family PASTEURELLACEAE: The phenotypic separation of Histophilus somni gen. nov., sp. nov. from other members of the family can, for most strains, be based on capnophilia, yellowish pigmentation and indole production. However, due to phenotypic variation, the use of a species-specific PCR test based on the 16S rRNA gene is included in the species description. This is justified by the high sequence similarity of the 16S rRNA gene within the species and the fact that the highest sequence similarity to any other taxon within the family is 93.4 %. The type strain, 8025(T)=ATCC 43625(T)=CCUG 36157(T), was isolated in the USA from a bovine brain with lesions of thromboembolic meningoencephalitis.

Lynette B. Corbeil - One of the best experts on this subject based on the ideXlab platform.

  • Effect of Histophilus somni on Heart and Brain Microvascular Endothelial Cells
    Veterinary pathology, 2017
    Co-Authors: Donal O’toole, R. Hunter, T. Allen, B. Zekarias, Jason S. Lehmann, Kwang Sik Kim, Dennis J. Grab, Lynette B. Corbeil
    Abstract:

    Histophilus somni is a pathogenic gram-negative bacterium responsible for pneumonia and septicemia in cattle. Sequelae include infectious thrombotic meningoencephalitis (ITME), myocarditis, arthrit...

  • Histophilus somni Surface Proteins.
    Current Topics in Microbiology and Immunology, 2015
    Co-Authors: Lynette B. Corbeil
    Abstract:

    The pathogen surface is usually the first site of interaction with the host. Histophilus somni was earlier thought to only have an outer membrane on its surface. Now it is known that the surface is composed of many virulence factors, including outer membrane proteins, lipooligosaccharide or endotoxin, a fibrillar network, and an exopolysaccharide. Outer membrane blebs, endotoxin, the fibrillar network, and the exopolysaccharide are also shed from the surface. This review will focus on the surface proteins of this pathogen that may colonize the mucosal surface of ruminants as a commensal or may cause pneumonia, septicemia, myocarditis, thrombotic meningoencephalitis, arthritis, and/or abortion. The major outer membrane protein has been well studied. Since its size and epitopes vary from strain to strain, it may be useful for typing strains. Iron-regulated OMPs have also received much attention because of their role in iron uptake for in vivo growth of H. somni. Other OMPs may be protective, based on passive immunization with monospecific antibodies and active immunization experiments. The surface and shed fibrillar network has been shown to be an immunoglobulin-binding protein in that it binds bovine IgG2 by the Fc portion. Two repeat domains (DR1 and DR2) have cytotoxic Fic motifs. Vaccine studies with recombinant DR2 are promising. Studies of the bacterial genome as well as comparison of surface proteins of different strains from the various H. somni syndromes and carrier states will be discussed and have provided much insight into pathogenesis and protection.

  • Host Immune Response to Histophilus somni.
    Current topics in microbiology and immunology, 2015
    Co-Authors: Lynette B. Corbeil
    Abstract:

    Histophilus somni is known to cause several overlapping syndromes or to be found in genital or upper respiratory carrier states in ruminants. Vaccines have been used for decades, yet efficacy is controversial and mechanisms of protective immunity are not well understood. Since H. somni survives phagocytosis, it has sometimes been considered to be a facultative intercellular parasite, implying that cell-mediated immunity would be critical in protection. However, H. somni not only inhibits phagocyte function, but also is cytotoxic for macrophages. Therefore, it does not live for long periods in healthy phagocytes. Protection of calves against H. somni pneumonia by passive immunization is also evidence that H. somni is more like an extracellular pathogen than an intracellular pathogen. Several studies showed that bovine IgG2 antibodies are more protective than IgG1 antibodies. Even the IgG2 allotypes tend to vary in protection. Of course, antigenic specificity also determines protection. So far, there is most evidence for protection by a 40 K outer membrane protein and by Immunoglobulin binding protein A fibrils. Serology and immunohistochemistry have both been used for immunodiagnosis. Many evasive mechanisms by H. somni have been defined, including decreased phagocyte function, antibodies bound by shed antigens, decreased immune stimulation, and antigenic variation. Interaction of H. somni with other bovine respiratory disease organisms is another layer of pathogenesis. Studies of bovine respiratory syncytial virus (BRSV) and H. somni in calfhood pneumonia revealed an increase in IgE antibodies to H. somni, which were associated with more severe disease of longer duration than with either agent alone. Innate immune mechanisms at the epithelial cell level are also affected by dual infection by BRSV and H. somni as compared to either pathogen alone. Although much more work needs to be done, the complex mechanisms of H. somni immunity are becoming clearer.

  • Antibody responses of calves to Histophilus somni recombinant IbpA subunits.
    Comparative immunology microbiology and infectious diseases, 2012
    Co-Authors: Richard A. Kimball, Jason S. Lehmann, Laurel J. Gershwin, Carolyn A. Worby, Lynette B. Corbeil
    Abstract:

    Histophilus somni causes bovine pneumonia and septicemia, but protective immune responses are not well understood and immunodiagnostic methods are not well defined. We previously showed that antibody to a new virulence factor, IbpA, neutralizes cytotoxicity and immunization with a recombinant IbpA domain protects calves against experimental H. somni pneumonia. To further define immune responses to IbpA, we determined isotypic serum antibody responses to three IbpA domains (IbpA3, an N-terminal coiled coil region; IbpA5, a central region of 200 bp repeats and IbpA DR2, a C-terminal cytotoxic domain). ELISA was used to quantitate IgG1 or IgG2 antibodies to each of the IbpA subunits as well as H. somni whole cells (WCs) or culture supernatant (SUP). Calves experimentally infected with H. somni and monitored for up to 10 weeks had the least "0 time" (background) antibody levels to IbpA5, as well as the earliest and highest responses of greatest duration to the IbpA5 subunit. Responses of these calves were high to WC or SUP antigens but with higher "0 time" (background) antibody levels. We concluded that IbpA5 may be a useful immunodiagnostic antigen. Calves immunized with H. somni WC vaccine had antibody responses to WC antigens, but not to IbpA subunits before challenge. After challenge with H. somni, vaccinated calves had slight anamnestic responses to IbpA3 and IbpA5, but not to IbpA DR2. Since IbpA DR2 is a protective antigen, the data suggest the IbpA DR2 would be a useful addition to H. somni vaccines.

  • Diagnostic Exercise: Myocarditis Due to Histophilus somni in Feedlot and Backgrounded Cattle
    Veterinary pathology, 2009
    Co-Authors: Donal O’toole, R. Hunter, T. Allen, Lynette B. Corbeil
    Abstract:

    A backgrounding operation for calves in Wyoming identified a disease syndrome presenting as lethargy, fever, and death between November and January each year. An unfixed heart was submitted for examination, along with samples of lung. There was focal red discoloration in papillary muscle of the left ventricular myocardium. Histologically, the lesion corresponded to acute necrotizing myocarditis with myriad intravascular and intralesional Gram-negative coccobacilli. Histophilus somni was detected by bacterial culture and immunohistochemistry. Focal myocarditis due to H. somni occurs in fall-placed cattle in western provinces and states of North America, and it can be an appreciable source of death loss. Gross lesions are readily detected in affected hearts. The presence of such changes in papillary muscles of left ventricular myocardium in feedlot or backgrounded cattle should prompt a differential diagnosis of H. somni myocarditis.

Reinier Mutters - One of the best experts on this subject based on the ideXlab platform.

  • proposal of Histophilus somni gen nov sp nov for the three species incertae sedis haemophilus somnus haemophilus agni and Histophilus ovis
    International Journal of Systematic and Evolutionary Microbiology, 2003
    Co-Authors: Øystein Angen, Peter Ahrens, Peter Kuhnert, Henrik Christensen, Reinier Mutters
    Abstract:

    Earlier investigations have shown that ‘Haemophilus somnus’, ‘Haemophilus agni’ and ‘Histophilus ovis’ represent the same species. In the present investigation, the taxonomic position of this species is explored further by sequencing the 16S rRNA and rpoB genes of strains that were investigated previously by DNA–DNA hybridization. These results clearly support the allocation of this species to a novel genus within the family Pasteurellaceae. The phenotypic separation of Histophilus somni gen. nov., sp. nov. from other members of the family can, for most strains, be based on capnophilia, yellowish pigmentation and indole production. However, due to phenotypic variation, the use of a species-specific PCR test based on the 16S rRNA gene is included in the species description. This is justified by the high sequence similarity of the 16S rRNA gene within the species and the fact that the highest sequence similarity to any other taxon within the family is 93·4 %. The type strain, 8025T=ATCC 43625T=CCUG 36157T, was isolated in the USA from a bovine brain with lesions of thromboembolic meningoencephalitis.

  • proposal of Histophilus somni gen nov sp nov for the three species incertae sedis haemophilus somnus haemophilus agni and Histophilus ovis
    International Journal of Systematic and Evolutionary Microbiology, 2003
    Co-Authors: Øystein Angen, Peter Ahrens, Peter Kuhnert, Henrik Christensen, Reinier Mutters
    Abstract:

    Earlier investigations have shown that 'Haemophilus somnus', 'Haemophilus agni' and 'Histophilus ovis' represent the same species. In the present investigation, the taxonomic position of this species is explored further by sequencing the 16S rRNA and rpoB genes of strains that were investigated previously by DNA-DNA hybridization. These results clearly support the allocation of this species to a novel genus within the family PASTEURELLACEAE: The phenotypic separation of Histophilus somni gen. nov., sp. nov. from other members of the family can, for most strains, be based on capnophilia, yellowish pigmentation and indole production. However, due to phenotypic variation, the use of a species-specific PCR test based on the 16S rRNA gene is included in the species description. This is justified by the high sequence similarity of the 16S rRNA gene within the species and the fact that the highest sequence similarity to any other taxon within the family is 93.4 %. The type strain, 8025(T)=ATCC 43625(T)=CCUG 36157(T), was isolated in the USA from a bovine brain with lesions of thromboembolic meningoencephalitis.

Indra Sandal - One of the best experts on this subject based on the ideXlab platform.

  • The role of luxS in Histophilus somni virulence and biofilm formation.
    Infection and immunity, 2021
    Co-Authors: Yu Pan, Aloka B. Bandara, Indra Sandal, Shivakumara Siddaramappa, Allan Dickerman, Thomas J. Inzana
    Abstract:

    S-ribosylhomocysteinase (LuxS) is required for the synthesis of the autoinducer-2 (AI-2) quorum-sensing signalling molecule in many Gram-negative bacteria. The bovine (and ovine) opportunistic pathogen Histophilus somni contains a luxS, and forms a biofilm containing an exopolysaccharide (EPS) in the matrix. Since biofilm formation is regulated by quorum sensing in many bacteria, the role of luxS in H. somni virulence and biofilm formation was investigated. Although culture supernatants from H. somni were ineffective at inducing bioluminescence in the Vibrio harveyi reporter strain BB170, H. somni luxS complemented the biosynthesis of AI-2 in the luxS-deficient Escherichia coli strain DH5α. H. somni strain 2336 luxS was inactivated by tranposon mutagenesis. Comparison of RNA exression profiles revealed that many genes were significantly differentially expressed in the luxS mutant compared to the wildtype, whether the bacteria were grown planktonically or in a biofilm. Furthermore, the luxS mutant had a truncated and asialylated lipooligosaccharide (LOS), and was substantially more serum-sensitive than the wildtype. Not surprisingly, the luxS mutant was attenuated in a mouse model for H. somni virulence, and some of the altered phenotypes were partially restored after the mutation was complemented with a functional luxS However, no major differences were observed between the wildtype and the luxS mutant in regard to outer membrane protein profiles, biofilm formation, EPS production, or intracellular survival. These results indicate that luxS plays a role in H. somni virulence in the context of LOS biosynthesis, but not biofilm formation or other phenotypic properties examined.

  • The Role of luxS in Histophilus somni Virulence and Biofilm Formation.
    Infection and immunity, 2021
    Co-Authors: Yu Pan, Aloka B. Bandara, Indra Sandal, Shivakumara Siddaramappa, Allan Dickerman, Thomas J. Inzana
    Abstract:

    ABSTRACT S-Ribosylhomocysteinase (LuxS) is required for the synthesis of the autoinducer-2 (AI-2) quorum-sensing signaling molecule in many Gram-negative bacteria. The bovine (and ovine) opportunistic pathogen Histophilus somni contains luxS and forms a biofilm containing an exopolysaccharide (EPS) in the matrix. Since biofilm formation is regulated by quorum sensing in many bacteria, the roles of luxS in H. somni virulence and biofilm formation were investigated. Although culture supernatants from H. somni were ineffective at inducing bioluminescence in the Vibrio harveyi reporter strain BB170, H. somniluxS complemented the biosynthesis of AI-2 in the luxS-deficient Escherichia coli strain DH5α. H. somni strain 2336 luxS was inactivated by transposon mutagenesis. RNA expression profiles revealed that many genes were significantly differentially expressed in the luxS mutant compared to that in the wild-type, whether the bacteria were grown planktonically or in a biofilm. Furthermore, the luxS mutant had a truncated and asialylated lipooligosaccharide (LOS) and was substantially more serum sensitive than the wild-type. Not surprisingly, the luxS mutant was attenuated in a mouse model for H. somni virulence, and some of the altered phenotypes were partially restored after the mutation was complemented with a functional luxS. However, no major differences were observed between the wild-type and the luxS mutant in regard to outer membrane protein profiles, biofilm formation, EPS production, or intracellular survival. These results indicate that luxS plays a role in H. somni virulence in the context of LOS biosynthesis but not biofilm formation or other phenotypic properties examined.

  • Natural competence in Histophilus somni strain 2336.
    Veterinary microbiology, 2014
    Co-Authors: Nehal Shah, Aloka B. Bandara, Indra Sandal, Thomas J. Inzana
    Abstract:

    Histophilus somni is an etiologic agent of shipping fever pneumonia, myocarditis, and other systemic diseases of bovines. Virulence factors that have been identified in H. somni include biofilm formation, lipooligosaccharide phase variation, immunoglobulin binding proteins, survival in phagocytic cells, and many others. However, to identify the genes responsible for virulence, an efficient mutagenesis system is needed. Mutagenesis of H. somni using allelic exchange is difficult, likely due to its tight restriction modification system. Mutagenesis by natural transformation in Haemophilus influenzae is well established and shows a strong bias for fragments containing specific uptake signal sequences (USS) within the genome. We hypothesized that natural transformation may also be possible in H. somni strain 2336 because its genome is over-represented with H. influenzae USS (5'-AAGTGCGGT-3') and contains most of the genes necessary for competence. H. somni strain 2336 was successfully transformed and mutated with genomic linear DNA from an H. somni mutant (738Δlob2a), which contains a kanamycin-resistance (Kan(R)) gene and the USS within lob2A. Although most of the competence genes found in H. influenzae were present in H. somni, comD and the 5' portion of comE were absent, which may account for the low transformation efficiency. The transformation efficiency of strain 2336 was greatest during mid-log growth phase and when cyclic adenosine monophosphate was added to the transformation medium. However, mutants were not isolated when strain 2336 was transformed with genomic DNA containing the same Kan(R) gene from H. somni luxS or uspE mutants, which lack the USS in these specific genes. Shuttle vector pNS3K was also naturally transformed into strain 2336, though at a lower efficiency. However, natural transformation with either H. somni linear DNA (2336Δlob2A) or pNS3K was unsuccessful with H. somni commensal strain 129Pt and several other disease isolates.

  • Identification, structure, and characterization of an exopolysaccharide produced by Histophilus somni during biofilm formation
    BMC microbiology, 2011
    Co-Authors: Indra Sandal, Thomas J. Inzana, Antonio Molinaro, Christina De Castro, Jian Q. Shao, Michael A. Apicella, Andrew D. Cox, Frank St. Michael, Gretchen Berg
    Abstract:

    Background Histophilus somni, a gram-negative coccobacillus, is an obligate inhabitant of bovine and ovine mucosal surfaces, and an opportunistic pathogen responsible for respiratory disease and other systemic infections in cattle and sheep. Capsules are important virulence factors for many pathogenic bacteria, but a capsule has not been identified on H. somni. However, H. somni does form a biofilm in vitro and in vivo, and the biofilm matrix of most bacteria consists of a polysaccharide.

  • A genomic window into the virulence of Histophilus somni.
    Trends in microbiology, 2009
    Co-Authors: Indra Sandal, Thomas J. Inzana
    Abstract:

    Histophilus somni is an obligate inhabitant of the respiratory and genital mucosal surfaces of bovines and ovines. An individual strain can be a primary pathogen, an opportunistic pathogen, or a commensal, but can also move between these classifications if introduced into an appropriate site (e.g. the lungs) under conditions that favor bacterial persistence. H. somni is one of the bacterial agents responsible for bovine respiratory disease complex and can also cause a variety of systemic diseases in cattle and sheep. Isolates from disease sites, such as the lungs, heart, and brain, express a wide array of virulence factors (including biofilm formation) designed to evade host defense mechanisms. By contrast, some isolates from the healthy genital tract often lack many of these virulence factors. The genomic sequences of two bovine isolates, one from pneumonic lung and the other from healthy prepuce, have aided in deciphering the differences in phenotype and virulence between the two strains, and reveal their striking genetic similarity to Haemophilus influenzae and other members of the Pasteurellaceae.