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Trevor W. Alexander - One of the best experts on this subject based on the ideXlab platform.

  • development of bacterial therapeutics against the bovine respiratory pathogen Mannheimia Haemolytica
    Applied and Environmental Microbiology, 2019
    Co-Authors: Samat Amat, Edouard Timsit, Danica Baines, Jay Yanke, Trevor W. Alexander
    Abstract:

    ABSTRACT Bovine respiratory disease (BRD) is a major cause of morbidity and mortality in beef cattle. Recent evidence suggests that commensal bacteria of the bovine nasopharynx have an important role in maintaining respiratory health by providing colonization resistance against pathogens. The objective of this study was to screen and select bacterial therapeutic candidates from the nasopharynxes of feedlot cattle to mitigate the BRD pathogen Mannheimia Haemolytica. In a stepwise approach, bacteria (n = 300) isolated from the nasopharynxes of 100 healthy feedlot cattle were identified and initially screened (n = 178 isolates from 12 different genera) for growth inhibition of M. Haemolytica. Subsequently, selected isolates were evaluated for the ability to adhere to bovine turbinate (BT) cells (n = 47), compete against M. Haemolytica for BT cell adherence (n = 15), and modulate gene expression in BT cells (n = 10). Lactobacillus strains had the strongest inhibition of M. Haemolytica, with 88% of the isolates (n =33) having inhibition zones ranging from 17 to 23 mm. Adherence to BT cells ranged from 3.4 to 8.0 log10 CFU per 105 BT cells. All the isolates tested in competition assays reduced M. Haemolytica adherence to BT cells (32% to 78%). Among 84 bovine genes evaluated, selected isolates upregulated expression of interleukin 8 (IL-8) and IL-6 (P  IMPORTANCE Bovine respiratory disease (BRD) is a significant animal health issue impacting the beef industry. Current BRD prevention strategies rely mainly on metaphylactic use of antimicrobials when cattle enter feedlots. However, a recent increase in BRD-associated bacterial pathogens that are resistant to metaphylactic antimicrobials highlights a pressing need for the development of novel mitigation strategies. Based upon previous research showing the importance of respiratory commensal bacteria in protecting against bronchopneumonia, this study aimed to develop bacterial therapeutics that could be used to mitigate the BRD pathogen Mannheimia Haemolytica. Bacteria isolated from the respiratory tracts of healthy cattle were characterized for their inhibitory, adhesive, and immunomodulatory properties. In total, 6 strains were identified as having the best properties for use as intranasal therapeutics to inhibit M. Haemolytica. If successful in vivo, these strains offer an alternative to metaphylactic antimicrobial use in feedlot cattle for mitigating BRD.

  • susceptibility to tulathromycin in Mannheimia Haemolytica isolated from feedlot cattle over a 3 year period
    Frontiers in Microbiology, 2013
    Co-Authors: Trevor W. Alexander, Cassidy L. Klima, Shaun R. Cook, E Topp, T A Mcallister
    Abstract:

    Mannheimia Haemolytica isolated from feedlot cattle were tested for tulathromycin resistance. Cattle were sampled over a three-year period, starting 12 months after approval of tulathromycin for prevention and treatment of bovine respiratory disease. Nasopharyngeal samples from approximately 5,814 cattle were collected when cattle entered feedlots (N = 4) and again from the same cattle after ≥ 60 d on feed. The antimicrobial use history for each animal was recorded. Mannheimia Haemolytica was isolated from 796 (13.7%) entry samples and 1,038 (20.6%) ≥ 60 d samples. Of the cattle positive for M. Haemolytica, 18.5%, 2.9%, and 2.4% were administered therapeutic concentrations of tulathromycin, tilmicosin, or tylosin tartrate, respectively. In addition, 13.2% were administered subtherapeutic concentrations of tylosin phosphate in feed. In years one and two, no tulathromycin-resistant M. Haemolytica were detected, whereas 5 isolates (0.4%) were resistant in year three. These resistant isolates were collected from three cattle originating from a single pen, were all serotype 1, and were genetically related (≥ 89% similarity) according to pulsed-field gel electrophoreses patterns. The five tulathromycin-resistant isolates were multi-drug resistant also exhibiting resistance to oxytetracycline, tilmicosin, ampicillin, or penicillin. The macrolide resistance genes erm(42), erm(A), erm(B), erm(F), erm(X) and msr(E)-mph(E), were not detected in the tulathromycin-resistant M. Haemolytica. This study showed that tulathromycin resistance in M. Haemolytica from a general population of feedlot cattle in western Canada was low and did not change over a three-year period after tulathromycin was approved for use in cattle.

  • Draft Genome Sequence of a Mannheimia Haemolytica Serotype 6 Isolate Collected from the Nasopharynx of a Beef Calf with Bovine Respiratory Disease.
    Genome Announcements, 2013
    Co-Authors: Cassidy L. Klima, Trevor W. Alexander, Shaun R. Cook, Kristen R. Hahn, Steve Hendrick, Kingsley-kwaku Amoako, Tim A. Mcallister
    Abstract:

    ABSTRACT The draft genome of a Mannheimia Haemolytica serotype 6 isolate obtained from the nasopharynx of a feedlot calf with bovine respiratory disease is described.

  • Investigation of Mannheimia Haemolytica bacteriophages relative to host diversity
    Journal of Applied Microbiology, 2013
    Co-Authors: Shaun R. Cook, Cassidy L. Klima, Trevor W. Alexander, L. B. Selinger, Tim A. Mcallister
    Abstract:

    Aims This study aimed to characterize the impact of lytic and temperate bacteriophages on the genetic and phenotypic diversity of Mannheimia Haemolytica from feedlot cattle. Methods and Results Strictly lytic phages were not detected from bovine nasopharyngeal (n = 689) or water trough (n = 30) samples, but Myoviridae- or Siphoviridae-like phages were induced from 54 of 72 M. Haemolytica strains by mitomycin C, occasionally from the same strain. Phages with similar restriction fragment length polymorphism profiles (RFLP ≥70% relatedness) shared common host serotypes 1 or 2 (P 

Patricia E. Shewen - One of the best experts on this subject based on the ideXlab platform.

  • a multivalent Mannheimia bibersteinia vaccine protects bighorn sheep against Mannheimia Haemolytica challenge
    Clinical and Vaccine Immunology, 2011
    Co-Authors: Renuka Subramaniam, Patricia E. Shewen, Sudarvili Shanthalingam, Jegarubee Bavananthasivam, Abirami Kugadas, Douglas C Hodgins, Kathleen A Potter, William J Foreyt, George M Barrington, Donald P Knowles
    Abstract:

    Bighorn sheep (BHS) are more susceptible than domestic sheep (DS) to Mannheimia Haemolytica pneumonia. Although both species carry M. Haemolytica as a commensal bacterium in the nasopharynx, DS carry mostly leukotoxin (Lkt)-positive strains while BHS carry Lkt-negative strains. Consequently, antibodies to surface antigens and Lkt are present at much higher titers in DS than in BHS. The objective of this study was to determine whether repeated immunization of BHS with multivalent Mannheimia-Bibersteinia vaccine will protect them upon M. Haemolytica challenge. Four BHS were vaccinated with a culture supernatant vaccine prepared from M. Haemolytica serotypes A1 and A2 and Bibersteinia trehalosi serotype T10 on days 0, 21, 35, 49, and 77. Four other BHS were used as nonvaccinated controls. On the day of challenge, 12 days after the last immunization, the mean serum titers of Lkt-neutralizing antibodies and antibodies to surface antigens against M. Haemolytica were 1:160 and 1:4,000, respectively. Following intranasal challenge with M. Haemolytica A2 (1 10 5 CFU), all four control BHS died within 48 h. Necropsy revealed acute fibrinonecrotic pneumonia characteristic of M. Haemolytica infection. None of the vaccinated BHS died during the 8 weeks postchallenge observation period. Radiography at 3 weeks postchallenge revealed no lung lesions in two vaccinated BHS and mild lesions in the other two, which resolved by 8 weeks postchallenge. These results indicate that if BHS can be induced to develop high titers of Lkt-neutralizing antibodies and antibodies to surface antigens, they are likely to survive M. Haemolytica challenge which is likely to reduce the BHS population decline due to pneumonia. The bighorn sheep (BHS; Ovis canadensis) population in North America has declined drastically during the last century due to a combination of factors, including loss of habitat, competition for forage with domestic livestock, predation, and disease. Pneumonia is the primary disease that causes significant mortality in BHS (2, 26). Although Mannheimia Haemolytica, Bibersteinia trehalosi, and Pasteurella multocida have been isolated from several pneumonia outbreaks, only M. Haemolytica has been shown to consistently cause fatal pneumonia in BHS under experimental conditions (4, 7, 9, 22). Virulence factors of M. Haemolytica include capsule, outer membrane proteins, neuraminidase, lipopolysaccharide, and a potent exotoxin called leukotoxin (Lkt), which is cytolytic to all subsets of ruminant leukocytes (3, 14). Based on the fact that Lkt deletion mutants cause no mortality in BHS (4) and lower mortality and mild lung lesions in cattle (12, 19, 24), Lkt has been accepted as the major virulence factor of this organism. Although M. Haemolytica causes pneumonia in all ruminants,

  • Mannheimia Haemolytica and bovine respiratory disease
    Animal Health Research Reviews, 2007
    Co-Authors: J Rice, L Carrascomedina, Douglas C Hodgins, Patricia E. Shewen
    Abstract:

    Mannheimia Haemolytica is the principal bacterium isolated from respiratory disease in feedlot cattle and is a significant component of enzootic pneumonia in all neonatal calves. A commensal of the nasopharynx, M. Haemolytica is an opportunist, gaining access to the lungs when host defenses are compromised by stress or infection with respiratory viruses or mycoplasma. Although several serotypes act as commensals, A1 and A6 are the most frequent isolates from pneumonic lungs. Potential virulence factors include adhesin, capsular polysaccharide, fimbriae, iron-regulated outer membrane proteins, leukotoxin (Lkt), lipopolysaccharide (LPS), lipoproteins, neuraminidase, sialoglycoprotease and transferrin-binding proteins. Of these, Lkt is pivotal in induction of pneumonia. Lkt-mediated infiltration and destruction of neutrophils and other leukocytes impairs bacterial clearance and contributes to development of fibrinous pneumonia. LPS may act synergistically with Lkt, enhancing its effects and contributing endotoxic activity. Antibiotics are employed extensively in the feedlot industry, both prophylactically and therapeutically, but their efficacy varies because of inconsistencies in diagnosis and treatment regimes and development of antibiotic resistance. Vaccines have been used for many decades, even though traditional bacterins failed to demonstrate protection and their use often enhanced disease in vaccinated animals. Modern vaccines use culture supernatants containing Lkt and other soluble antigens, or bacterial extracts, alone or combined with bacterins. These vaccines have 50–70% efficacy in prevention of M. Haemolytica pneumonia. Effective control of M. Haemolytica pneumonia is likely to require a combination of more definitive diagnosis, efficacious vaccines, therapeutic intervention and improved management practices.

  • Construction and analysis of a Mannheimia Haemolytica A1 luxS mutant.
    Veterinary Microbiology, 2005
    Co-Authors: Amanda N. Van Der Vinne, Reggie Y.c. Lo, Patricia E. Shewen
    Abstract:

    Abstract Mannheimia Haemolytica A1 is the causative agent of bovine pneumonic pasteurellosis, a major cause of sickness, death, and economic loss to the feedlot cattle industry. M. Haemolytica A1 produces autoinducer-2 (AI-2) like molecules that are capable of inducing quorum sensing system 2 of Vibrio harveyi. This interspecies quorum sensing system has been shown to regulate the expression of virulence genes in several pathogenic bacteria. The protein central to the production of AI-2 is LuxS. To determine if quorum sensing is involved in the regulation of virulence genes in M. Haemolytica A1, a luxS mutant was constructed by replacing luxS with a cat cassette. This mutant was verified by PCR analysis, Southern hybridization, as well as its inability to induce bioluminescence in the V. harveyi reporter strain. RT-PCR analysis showed there was no difference in leukotoxin (lktC) mRNA levels, however there were increased mRNA levels of putative virulence associated genes, transferrin binding protein B (tbpB), adhesin (ahs) and capsule biosynthesis (nmaA). Electron microscopy showed that the level of encapsulation in the mutant is higher than the parent. Additionally, the mutant was slightly more adherent to bovine tracheal cells than the parent. In vitro competition assays showed the mutant out-competed the parent under iron-restricted conditions. However, in a calf challenge, the parent was the dominant isolate recovered.

  • Mannheimia Haemolytica serotype 1 and Pasteurella trehalosi serotype 10 culture supernatants contain fibrinogen-binding proteins.
    Veterinary Immunology and Immunopathology, 2002
    Co-Authors: Heather J Mcneil, Patricia E. Shewen, Reggie Y.c. Lo, Jennifer A. Conlon, Michael W. Miller
    Abstract:

    Abstract Fibrinogen-binding proteins were found in the culture supernatants of Mannheimia Haemolytica serotype 1 (ATCC 43270) and Pasteurella trehalosi serotype 10 (ECO-100). Sheep fibrinogen was biotinylated and shown to bind to proteins in the culture supernatants by modified western blot. Fibrinogen-binding proteins in the culture supernatant may be important virulence factors leading to the characteristic fibrinous pneumonia caused by these organisms and may be critical antigenic targets for immune prophylaxis.

S K Maheswaran - One of the best experts on this subject based on the ideXlab platform.

Bindu Nanduri - One of the best experts on this subject based on the ideXlab platform.

  • Transcriptome profile of a bovine respiratory disease pathogen: Mannheimia Haemolytica PHL213
    BMC Bioinformatics, 2012
    Co-Authors: Joseph S Reddy, James M Watt, Shane C Burgess, Ranjit Kumar, Mark L Lawrence, Bindu Nanduri
    Abstract:

    Background Computational methods for structural gene annotation have propelled gene discovery but face certain drawbacks with regards to prokaryotic genome annotation. Identification of transcriptional start sites, demarcating overlapping gene boundaries, and identifying regulatory elements such as small RNA are not accurate using these approaches. In this study, we re-visit the structural annotation of Mannheimia Haemolytica PHL213, a bovine respiratory disease pathogen. M. Haemolytica is one of the causative agents of bovine respiratory disease that results in about $3 billion annual losses to the cattle industry. We used RNA-Seq and analyzed the data using freely-available computational methods and resources. The aim was to identify previously unannotated regions of the genome using RNA-Seq based expression profile to complement the existing annotation of this pathogen. Results Using the Illumina Genome Analyzer, we generated 9,055,826 reads (average length ~76 bp) and aligned them to the reference genome using Bowtie. The transcribed regions were analyzed using SAMTOOLS and custom Perl scripts in conjunction with BLAST searches and available gene annotation information. The single nucleotide resolution map enabled the identification of 14 novel protein coding regions as well as 44 potential novel sRNA. The basal transcription profile revealed that 2,506 of the 2,837 annotated regions were expressed in vitro , at 95.25% coverage, representing all broad functional gene categories in the genome. The expression profile also helped identify 518 potential operon structures involving 1,086 co-expressed pairs. We also identified 11 proteins with mutated/alternate start codons. Conclusions The application of RNA-Seq based transcriptome profiling to structural gene annotation helped correct existing annotation errors and identify potential novel protein coding regions and sRNA. We used computational tools to predict regulatory elements such as promoters and terminators associated with the novel expressed regions for further characterization of these novel functional elements. Our study complements the existing structural annotation of Mannheimia Haemolytica PHL213 based on experimental evidence. Given the role of sRNA in virulence gene regulation and stress response, potential novel sRNA described in this study can form the framework for future studies to determine the role of sRNA, if any, in M. Haemolytica pathogenesis.

  • Transcriptome profile of a bovine respiratory disease pathogen: Mannheimia Haemolytica PHL213.
    BMC bioinformatics, 2012
    Co-Authors: Joseph S Reddy, James M Watt, Shane C Burgess, Ranjit Kumar, Mark L Lawrence, Bindu Nanduri
    Abstract:

    Computational methods for structural gene annotation have propelled gene discovery but face certain drawbacks with regards to prokaryotic genome annotation. Identification of transcriptional start sites, demarcating overlapping gene boundaries, and identifying regulatory elements such as small RNA are not accurate using these approaches. In this study, we re-visit the structural annotation of Mannheimia Haemolytica PHL213, a bovine respiratory disease pathogen. M. Haemolytica is one of the causative agents of bovine respiratory disease that results in about $3 billion annual losses to the cattle industry. We used RNA-Seq and analyzed the data using freely-available computational methods and resources. The aim was to identify previously unannotated regions of the genome using RNA-Seq based expression profile to complement the existing annotation of this pathogen. Using the Illumina Genome Analyzer, we generated 9,055,826 reads (average length ~76 bp) and aligned them to the reference genome using Bowtie. The transcribed regions were analyzed using SAMTOOLS and custom Perl scripts in conjunction with BLAST searches and available gene annotation information. The single nucleotide resolution map enabled the identification of 14 novel protein coding regions as well as 44 potential novel sRNA. The basal transcription profile revealed that 2,506 of the 2,837 annotated regions were expressed in vitro, at 95.25% coverage, representing all broad functional gene categories in the genome. The expression profile also helped identify 518 potential operon structures involving 1,086 co-expressed pairs. We also identified 11 proteins with mutated/alternate start codons. The application of RNA-Seq based transcriptome profiling to structural gene annotation helped correct existing annotation errors and identify potential novel protein coding regions and sRNA. We used computational tools to predict regulatory elements such as promoters and terminators associated with the novel expressed regions for further characterization of these novel functional elements. Our study complements the existing structural annotation of Mannheimia Haemolytica PHL213 based on experimental evidence. Given the role of sRNA in virulence gene regulation and stress response, potential novel sRNA described in this study can form the framework for future studies to determine the role of sRNA, if any, in M. Haemolytica pathogenesis.

T A Mcallister - One of the best experts on this subject based on the ideXlab platform.

  • comparative analysis of multiple inducible phages from Mannheimia Haemolytica
    BMC Microbiology, 2015
    Co-Authors: Shaun R. Cook, Cassidy L. Klima, Jiaying Wang, Andrew M Kropinski, Dann Turner, T A Mcallister
    Abstract:

    Background Mannheimia Haemolytica is a commensal bacterium that resides in the upper respiratory tract of cattle that can play a role in bovine respiratory disease. Prophages are common in the M. Haemolytica genome and contribute significantly to host diversity. The objective of this research was to undertake comparative genomic analysis of phages induced from strains of M. Haemolytica serotype A1 (535A and 2256A), A2 (587A and 1127A) and A6 (1152A and 3927A).

  • susceptibility to tulathromycin in Mannheimia Haemolytica isolated from feedlot cattle over a 3 year period
    Frontiers in Microbiology, 2013
    Co-Authors: Trevor W. Alexander, Cassidy L. Klima, Shaun R. Cook, E Topp, T A Mcallister
    Abstract:

    Mannheimia Haemolytica isolated from feedlot cattle were tested for tulathromycin resistance. Cattle were sampled over a three-year period, starting 12 months after approval of tulathromycin for prevention and treatment of bovine respiratory disease. Nasopharyngeal samples from approximately 5,814 cattle were collected when cattle entered feedlots (N = 4) and again from the same cattle after ≥ 60 d on feed. The antimicrobial use history for each animal was recorded. Mannheimia Haemolytica was isolated from 796 (13.7%) entry samples and 1,038 (20.6%) ≥ 60 d samples. Of the cattle positive for M. Haemolytica, 18.5%, 2.9%, and 2.4% were administered therapeutic concentrations of tulathromycin, tilmicosin, or tylosin tartrate, respectively. In addition, 13.2% were administered subtherapeutic concentrations of tylosin phosphate in feed. In years one and two, no tulathromycin-resistant M. Haemolytica were detected, whereas 5 isolates (0.4%) were resistant in year three. These resistant isolates were collected from three cattle originating from a single pen, were all serotype 1, and were genetically related (≥ 89% similarity) according to pulsed-field gel electrophoreses patterns. The five tulathromycin-resistant isolates were multi-drug resistant also exhibiting resistance to oxytetracycline, tilmicosin, ampicillin, or penicillin. The macrolide resistance genes erm(42), erm(A), erm(B), erm(F), erm(X) and msr(E)-mph(E), were not detected in the tulathromycin-resistant M. Haemolytica. This study showed that tulathromycin resistance in M. Haemolytica from a general population of feedlot cattle in western Canada was low and did not change over a three-year period after tulathromycin was approved for use in cattle.