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Veterinary Microbiology Mar 2018Mannheimia haemolytica and Pasteurella multocida are two bacterial species implicated in the bovine respiratory disease complex (BRDC) that is costly to the beef and...
Mannheimia haemolytica and Pasteurella multocida are two bacterial species implicated in the bovine respiratory disease complex (BRDC) that is costly to the beef and dairy cattle industries. Both bacterial species are thought to occupy a similar niche as commensals in the upper respiratory tract. Many bacteria are thought to exist as biofilms in their hosts, perhaps in close proximity with other bacterial species. We previously showed that M. haemolytica forms biofilm on bovine respiratory epithelial cells in vitro. We are interested in the possibility that M. haemolytica and P. multocida co-exist as biofilms in the upper respiratory tract of cattle. In this study, we begin to explore this possibility by assessing the ability of M. haemolytica and P. multocida to form a biofilm on bovine respiratory epithelial cells in vitro. We found that M. haemolytica and P. multocida are separately able to form biofilms on bovine respiratory epithelial cells, but mutually inhibit one another when incubated together as a biofilm. Both the biofilm matrix (crystal violet stain) and bacterial numbers (CFU and PCR) were reduced when M. haemolytica and P. multocida were incubated together on fixed epithelial cells. This inhibition does not appear to result from a soluble factor, as neither conditioned medium nor separation of the two species by a transwell filter membrane reproduced the effect. We infer that when located in close proximity on the epithelial surface, M. haemolytica and P. multocida mutually regulate one another.
Topics: Animals; Antibiosis; Biofilms; Bronchi; Cattle; Epithelial Cells; Mannheimia haemolytica; Pasteurella multocida
PubMed: 29519520
DOI: 10.1016/j.vetmic.2018.02.015 -
Journal of Microbiological Methods Apr 2019Disruptive innovations in long-range, cost-effective direct template nucleic acid sequencing are transforming clinical and diagnostic medicine. A multidrug resistant...
Disruptive innovations in long-range, cost-effective direct template nucleic acid sequencing are transforming clinical and diagnostic medicine. A multidrug resistant strain and a pan-susceptible strain of Mannheimia haemolytica, isolated from pneumonic bovine lung samples, were sequenced at 146× and 111× coverage, respectively with Oxford Nanopore Technologies MinION. De novo assembly produced a complete genome for the non-resistant strain and a nearly complete assembly for the drug resistant strain. Functional annotation using RAST (Rapid Annotations using Subsystems Technology), CARD (Comprehensive Antibiotic Resistance Database) and ResFinder databases identified genes conferring resistance to different classes of antibiotics including β-lactams, tetracyclines, lincosamides, phenicols, aminoglycosides, sulfonamides and macrolides. Resistance phenotypes of the M. haemolytica strains were determined by minimum inhibitory concentration (MIC) of the antibiotics. Sequencing with a highly portable MinION device corresponded to MIC assays with most of the antimicrobial resistant determinants being identified with as few as 5437 reads, except for the genes responsible for resistance to Fluoroquinolones. The resulting quality assemblies and AMR gene annotation highlight the efficiency of ultra-long read, whole-genome sequencing (WGS) as a valuable tool in diagnostic veterinary medicine.
Topics: Animals; Anti-Bacterial Agents; Cattle; Drug Resistance, Bacterial; Genome, Bacterial; Mannheimia haemolytica; Microbial Sensitivity Tests; Nanopore Sequencing; Pneumonia of Calves, Enzootic; Sequence Analysis, DNA; Whole Genome Sequencing
PubMed: 30849421
DOI: 10.1016/j.mimet.2019.03.001 -
Veterinary Microbiology Oct 2015Pure cultures of non-haemolytic Mannheimia haemolytica, were cultivated from pleural effusion fluid and blood from a 1-month old Belgian Blue bull calf that was...
Pure cultures of non-haemolytic Mannheimia haemolytica, were cultivated from pleural effusion fluid and blood from a 1-month old Belgian Blue bull calf that was presented with apathy and anorexia. The isolates were identified as M. haemolytica by 16S rRNA gene sequencing and MALDI-TOF-MS. Since haemolysis on blood agar plates is considered a hallmark of M. haemolytica we wanted to elucidate the unusual phenotype of the isolated strain. Therefore the leukotoxin operon (lktCABD), responsible for the haemolytic phenotype of M. haemolytica and regarded as the most important virulence factor, was completely sequenced. The leukotoxin operon of the isolated strain showed a deletion in the lktA gene, resulting in a truncated LktA protein. The absence of a complete LktA protein is responsible for the non-haemolytic phenotype of the strain. To the best of our knowledge, this is the first report of a well-characterized non-haemolytic M. haemolytica isolate causing disease in cattle.
Topics: Animals; Cattle; Cattle Diseases; Exotoxins; Gene Deletion; Gene Expression Regulation, Bacterial; Male; Mannheimia haemolytica; Pleuropneumonia; Sepsis
PubMed: 26344042
DOI: 10.1016/j.vetmic.2015.08.019 -
Veterinary Microbiology Jan 2013The primary objective of this study was to determine, at the lung level, whether single or multiple clones of Mannheimia haemolytica are present within a pen during a...
The primary objective of this study was to determine, at the lung level, whether single or multiple clones of Mannheimia haemolytica are present within a pen during a bovine respiratory disease (BRD) episode. A secondary objective was to assess whether M. haemolytica isolates obtained from nasal swabs (NS) are identical to those isolated deeper within the respiratory tract. Sixteen BRD episodes that naturally occurred in 12 pens of eight to 12 bulls (n=112) newly-received at three fattening operations were investigated. One hundred and seventy five M. haemolytica isolates were collected from 239 pairs of trans-tracheal aspirations (TTA) and NS performed during these 16 BRD episodes. M. haemolytica isolates were characterized by pulsed-field gel electrophoresis (PFGE). PFGE types obtained from NS and TTA were then compared. M. haemolytica was isolated during 14 BRD episodes. Two to three different clones of M. haemolytica were recovered during 10 episodes whereas only one clone was recovered in four episodes. A moderate agreement (kappa=0.50) between NS and TTA for M. haemolytica isolation was observed. Identical PFGE types were only observed in 77% of matched NS-TTA pairs. The significant within-pen diversity of M. haemolytica during BRD episodes indicates that the disease is not primarily due to the spread of a single virulent clone among cattle and highlights the importance of predisposing factors that enable the resident flora to overcome the cattle's immune system. The results also demonstrate that isolates recovered from NS are not always representative of the isolates present deeper within the respiratory tract.
Topics: Animal Husbandry; Animals; Cattle; Cattle Diseases; Electrophoresis, Gel, Pulsed-Field; Lung; Male; Mannheimia haemolytica; Meat; Pasteurellaceae Infections; Respiratory Tract Diseases
PubMed: 22901531
DOI: 10.1016/j.vetmic.2012.07.044 -
PloS One 2016Bovine respiratory disease is a common health problem in beef production. The primary bacterial agent involved, Mannheimia haemolytica, is a target for antimicrobial... (Comparative Study)
Comparative Study
Bovine respiratory disease is a common health problem in beef production. The primary bacterial agent involved, Mannheimia haemolytica, is a target for antimicrobial therapy and at risk for associated antimicrobial resistance development. The role of M. haemolytica in pathogenesis is linked to serotype with serotypes 1 (S1) and 6 (S6) isolated from pneumonic lesions and serotype 2 (S2) found in the upper respiratory tract of healthy animals. Here, we sequenced the genomes of 11 strains of M. haemolytica, representing all three serotypes and performed comparative genomics analysis to identify genetic features that may contribute to pathogenesis. Possible virulence associated genes were identified within 14 distinct prophage, including a periplasmic chaperone, a lipoprotein, peptidoglycan glycosyltransferase and a stress response protein. Prophage content ranged from 2-8 per genome, but was higher in S1 and S6 strains. A type I-C CRISPR-Cas system was identified in each strain with spacer diversity and organization conserved among serotypes. The majority of spacers occur in S1 and S6 strains and originate from phage suggesting that serotypes 1 and 6 may be more resistant to phage predation. However, two spacers complementary to the host chromosome targeting a UDP-N-acetylglucosamine 2-epimerase and a glycosyl transferases group 1 gene are present in S1 and S6 strains only indicating these serotypes may employ CRISPR-Cas to regulate gene expression to avoid host immune responses or enhance adhesion during infection. Integrative conjugative elements are present in nine of the eleven genomes. Three of these harbor extensive multi-drug resistance cassettes encoding resistance against the majority of drugs used to combat infection in beef cattle, including macrolides and tetracyclines used in human medicine. The findings here identify key features that are likely contributing to serotype related pathogenesis and specific targets for vaccine design intended to reduce the dependency on antibiotics to treat respiratory infection in cattle.
Topics: Animals; Base Sequence; CRISPR-Cas Systems; Cattle; DNA, Intergenic; Genome, Bacterial; Mannheimia haemolytica; Molecular Sequence Data; Phylogeny; Prophages; Sequence Analysis, DNA
PubMed: 26926339
DOI: 10.1371/journal.pone.0149520 -
The Journal of Antimicrobial... Jun 2022
Topics: Anti-Bacterial Agents; Anti-Infective Agents; Drug Resistance, Bacterial; Mannheimia haemolytica; Pasteurella multocida
PubMed: 35434742
DOI: 10.1093/jac/dkac116 -
Current Microbiology Jun 2024Mannheimia haemolytica is recognized as principal pathogen associated with pneumonic pasteurellosis leading to huge economic losses to small ruminant farmers. Even...
Mannheimia haemolytica is recognized as principal pathogen associated with pneumonic pasteurellosis leading to huge economic losses to small ruminant farmers. Even though the disease causes huge economic losses, epidemiology of M. haemolytica is less studied, hindering the formulation of effective control strategies. Current study aimed to highlight molecular characterisation of M. haemolytica strains isolated from ovine pneumonic infection. M. haemolytica 27 isolates with two reference strains were characterised using capsular and virulence gene typing, multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) methods. M. haemolytica serotype A2 recognized as predominant serotype (74%) followed by A6 (11%) and A1 (5%) serotypes. Virulence gene profiling by PCRs showed dominance of all five virulent genes [such as adh and gcp (100% each)] followed by gs60 (88.8%), lktC (85.2%), tbpB (51.9%) and least nmaA gene (14.8%). MLST profiling delineated M. haemolytic isolates into 11 sequence types (STs) with most prevalent being ST37 (27.9%) and ST16 (23%) and nine new STs (ST37, 38, 39, 40, 41, 42, 47, 48, and 49). These new STs did not belong to any of the three clonal complexes (CC4, CC8 and CC28). ST16 was exclusively noted in A1 and A6 serotypes. Amongst 25 isolates, 22 pulsotypes (GD 0.88) recorded indicated variability of the M. haemolytica isolates in PFGE analysis. In conclusion, the study suggested dominance of M. haemolytica serotype A2 harbouring different virulent genes, diverse STs and pulsotypes responsible for pneumonic pasteurellosis frequently encountered in sheep.
Topics: Animals; Mannheimia haemolytica; Sheep; Sheep Diseases; India; Multilocus Sequence Typing; Pasteurellosis, Pneumonic; Serogroup; Electrophoresis, Gel, Pulsed-Field; Virulence Factors; Virulence; Phylogeny
PubMed: 38862704
DOI: 10.1007/s00284-024-03740-7 -
Journal of Applied Microbiology Jun 2013This study aimed to characterize the impact of lytic and temperate bacteriophages on the genetic and phenotypic diversity of Mannheimia haemolytica from feedlot cattle.
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 < 0·0001). Likewise, phages with similar RFLP tended to occur in genetically related host bacteria (70-79% similarity). Host range assays showed that seven phages from host serotypes 1, 2 and 6 lysed representative strains of serotypes 1, 2 or 8. The genome of vB_MhM_1152AP from serotype 6 was found to be collinear with P2-like phage φMhaA1-PHL101.
CONCLUSIONS
Prophages are a significant component of the genome of M. haemolytica and contribute significantly to host diversity. Further characterization of the role of prophage in virulence and persistence of M. haemolytica in cattle could provide insight into approaches to control this potential respiratory pathogen.
SIGNIFICANCE AND IMPACT OF THE STUDY
This study demonstrated that prophages are widespread within the genome of M. haemolytica isolates and emphasized the challenge of isolating lytic phage as a therapeutic against this pathogen.
Topics: Animals; Bacteriophages; Cattle; Enrofloxacin; Fluoroquinolones; Genetic Variation; Host Specificity; Mannheimia haemolytica; Prophages
PubMed: 23489937
DOI: 10.1111/jam.12185 -
The Canadian Veterinary Journal = La... Sep 2022Cattle at high-risk for bovine respiratory disease on entry to western Canadian feedlots are often treated metaphylactically with antimicrobials from the macrolide...
Changes in the phenotypic susceptibility of isolates to macrolide antimicrobials during the early feeding period following metaphylactic tulathromycin use in western Canadian feedlot calves.
Cattle at high-risk for bovine respiratory disease on entry to western Canadian feedlots are often treated metaphylactically with antimicrobials from the macrolide class. High levels of resistance to macrolides have been reported in isolates from clinical samples, but it is less clear whether this trend extends to the broader feedlot population. The objective was to describe near-term [< 40 days on feed (DOF)] changes in the recovery and susceptibility of isolates from healthy feedlot calves after metaphylactic exposure to tulathromycin. Eight cohorts of 100 calves ( = 800) were sampled deep nasopharyngeal swab at entry processing (, before metaphylaxis, at 1 DOF) and again at 13 DOF. Ten calves from each cohort ( = 80) were randomly sampled a third time at 36 DOF. Recovery of isolates across all cohorts increased over the study period, from 33% (95% CI: 26.5 to 40.2%) at 1 DOF to 75% (95% CI: 71.4 to 78.3%) at 36 DOF. A significant shift in the minimum inhibitory concentration (MIC) distribution of tulathromycin from 1 DOF (MIC ≤ 8 μg/mL) to 13 DOF (MIC > 64 μg/mL) was observed. A subset of 36 isolates from 13 DOF screened for macrolide resistance genes multiplex polymerase chain reaction all harbored the and genes. Recovery of at 13 and 36 DOF did not decline in response to metaphylactic use of tulathromycin; conversely, we inferred the potential for rapid inter-pen spread of a macrolide-resistant clone by 13 DOF in 6 of 8 pens under selective pressure from antimicrobial use.
Topics: Animals; Anti-Bacterial Agents; Canada; Cattle; Cattle Diseases; Disaccharides; Drug Resistance, Bacterial; Heterocyclic Compounds; Humans; Macrolides; Mannheimia haemolytica
PubMed: 36060481
DOI: No ID Found -
BMC Microbiology Aug 2015Mannheimia 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...
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).
RESULTS
Overall, four P2-like (535AP1, 587AP1, 1127AP1 and 2256AP1; genomes: 34.9-35.7 kb; G+C content: 41.5-42.1 %; genes: 51-53 coding sequences, CDSs), four λ-like (535AP2, 587AP2, 1152AP2 and 3927AP1; genomes: 48.6-52.1 kb; 41.1-41.4 % mol G+C; genes: 77-83 CDSs and 2 tRNAs) and one Mu-like (3927AP2; genome: 33.8 kb; 43.1 % mol G+C; encoding 50 CDSs) phages were identified. All P2-like phages are collinear with the temperate phage φMhaA1-PHL101 with 535AP1, 2256AP1 and 1152AP1 being most closely related, followed by 587AP1 and 1127AP1. Lambdoid phages are not collinear with any other known λ-type phages, with 587AP2 being distinct from 535AP2, 3927AP1 and 1152AP2. All λ-like phages contain genes encoding a toxin-antitoxin (TA) system and cell-associated haemolysin XhlA. The Mu-like phage induced from 3927A is closely related to the phage remnant φMhaMu2 from M. haemolytica PHL21, with similar Mu-like phages existing in the genomes of M. haemolytica 535A and 587A.
CONCLUSIONS
This is among the first reports of both λ- and Mu-type phages being induced from M. haemolytica. Compared to phages induced from commensal strains of M. haemolytica serotype A2, those induced from the more virulent A1 and A6 serotypes are more closely related. Moreover, when P2-, λ- and Mu-like phages co-existed in the M. haemolytica genome, only P2- and λ-like phages were detected upon induction, suggesting that Mu-type phages may be more resistant to induction. Toxin-antitoxin gene cassettes in λ-like phages may contribute to their genomic persistence or the establishment of persister subpopulations of M. haemolytica. Further work is required to determine if the cell-associated haemolysin XhlA encoded by λ-like phages contributes to the pathogenicity and ecological fitness of M. haemolytica.
Topics: Base Composition; DNA, Viral; Mannheimia haemolytica; Molecular Sequence Data; Prophages; Sequence Analysis, DNA; Sequence Homology; Synteny; Virus Activation
PubMed: 26318735
DOI: 10.1186/s12866-015-0494-5