Mannheimia haemolytica

M. haemolytica is responsible for causing contagious bovine pleuropneumonia, a bacterial disease which causes pneumonia and inflammation of the lung membranes. It is a Gram-negative coccobacillus (elongated, rod spheres) which shows mild haemolysis when plated on blood agar plates. This species comprises of 12 capsular serotypes (of which some are more responsible for disease than others). Serotype A2 is associated with sheep pneumonia.

Typically, diseases associated with M. haemolytica present themselves as fever, along with nasal discharge, coughing, inappetance and weight loss. Death associated with this bacteria is typically due to acute fibrous pleuropneumonia. Pathological observations would reveal an obstruction of the bronchioles with a fibrous exudate and an accumulation of neutrophils and fibrin in the alveoli. There will also be thrombosis and distension of the lymphatic vessels.

Mannheimia haemolytica as an Opportunistic Pathogen

M. haemolytica resides in the nasopharynx and tonsils of healthy cattle. A balance between the host and bacteria, under good environmental conditions maintains homeostasis between the host and bacteria (i.e. no disease). Should environmental conditions alter however, the balance may be tipped in favour of the bacteria, removing the status of homeostasis and resulting in disease. Key inciting events for disease include; weaning, adverse weather conditions, dehorning, feed changes and transportation i.e. causing stress. Stress provokes the bacteria in the nasal/tonsillar mucosa allowing them to be inhaled in to the lungs.

In healthy cattle clearance of the bacteria from the lungs is efficient enough to prevent disease. This suggests that stress induced alterations in immune functions can lead to host susceptibility and thus development of disease.

Close contact with other animals increases exposure to microorganisms which upsets the natural balance of commensal bacteria within the host. Changes in the natural microflora can cause M. haemolytica to revert to a pathogenic state. Slight changes in the environment are responsible for creating a favourable situation for M. haemolytica to colonies the lung and thus cause disease.

Mannheimia haemolytica Virulence Factors

Virulence factors of M. haemolytica include:

• Adhesins – Required for initial colonization and adherence to host cells
• Capsular polysaccharide – Helps prevent opsonisation and thus has anti-phagocytic properties
• Sialoglycoprotease – An enzyme which cleaves IgG and thus reduces opsonisation of the bacteria
• Neuraminidase – An enzyme which reduces the viscosity of respiratory mucus, reducing the chance of the bacteria being excreted via mucus
• Iron-binding proteins – Removes iron from the host, for the benefit of the bacterium
• Leukotoxin – A primary toxin involved in the pathogenesis of M. haemolytica

Mannheimia haemolytica Leukotoxin

Leukotoxin (LKT) is an actively secreted exotoxin which targets leukocytes, primarily neutrophils. The toxin is encoded for by four genes (lkt A-D) which can be found in the RTX toxin operon.

• lktA encodes for the active toxin
• lktB and lktD encode for the secretion of the toxin
• lktC encodes for proteins responsible in the transportation and activation of the toxin

LKT help bacteria survive by allowing them to evade phagocytes. It is a type 2 exotoxin (membrane-damaging) which binds to leukocytes via the cell surface receptor CD18 (an integrin). It forms pores in the cell membrane of leukocytes which leads to an influx of K⁺/Ca²⁺ ions. This promotes swelling of the leukocyte and ultimately lysis of the cell.

Neutrophils affected by the toxin also become overly active, they overproduce certain mediators, reactive oxygen species and proteases, all of which promote further cellular and tissue damage within the host. Lesions develop which are filled with fibrous exudate and thrombosis of lymphatic vessels occurs. The alveolar epithelium also becomes damaged which is believed to be associated with neutrophil infiltration.

Controlling Mannheimia haemolytica Infections

Methods which could be implemented to reduce cases of M. haemolytica infections include:

• Livestock management – Reduce stress of cattle by effectively managing when and how calves are weaned, sold and transported
• Antibiotic use – Injectable antibiotic regimes are extensively used, especially in the intensive farming seen in large feedlots. However evidence shows that M. haemolytica are developing resistance to many of the common antibiotics used, such as; penicillin, ampicillin, tetracycline and sulphonamide
• Vaccines – Both killed and live attenuated vaccines are used as well as cell-free supernatants such as the leukotoxin or capsular polysaccharide
• Genetically Modified plants – A suggestion has been made for the use of GM crops to feed cattle in the future. The GM plants would produce M. haemolytica antigens which would essentially act as edible vaccines

These control methods could help to restore balance to the situation (previously unbalanced by negative environmental factors). Thus homeostasis between host and bacteria would be restored and disease prevented.

Bordetella bronchiseptica

Bordetella bronchiseptica is an evolutionary progenitor of B. pertussis and is one of the organisms responsible for causing kennel cough in dogs. Kennel cough is the term used to describe a disease which causes coughing in a dog due to inflammation of the trachea and lower airways. Although kennel cough is primarily caused by infection of the airways with B. bronchiseptica it can also be caused by viruses such as canine parainfluenza. Clinical signs of kennel cough include; an intense cough, mucus, nasal discharge and breathing difficulties.

Kennel cough gets its name from the frequency of infections which arise in dogs temporarily kenneled for example whilst the owner is on holiday. A number of dogs can carry the infection and the close contact in the kennels promotes transfer of the infection. Infection is caused either airborne transmission or via direct contact and has an incubation period of 3-10 days. Infected dogs can carry and shed the infection for up to four months after recovering from the disease. Because it require a only a few B. bronchiseptica bacteria to establish and infection in the airways (i.e. low infectious dose), B. bronchiseptica are considered highly infectious.

Bordetella Bronchiseptica Virulence Factors

Adhesins

• Fimbriae – Associated with the initial adherence of the bacteria to epithelia. This form of adhesin allows the bacteria to latch on to host cells and begin to proliferate and form infectious colonies.
• Filamentous Haemagglutinin Adhesin (FHA) – A large, filamentous protein which serves as a dominant attachment factor for adherence to host colliery epithelia cells of the respiratory tract. It is associated with biofilm formation and possesses at least four binding domains which can bind to different cell receptors on the epithelial cell surface.
• Pertactin – An ‘autotransporter protein’ capable of getting to the cell surface without the need of accessory proteins. Pertactin also acts as an adhesin.

Adenylate Cyclase (cyaA)

• Structurally similar to M. haemolytica leukotoxin
• Different function, cyaA enters host cells and catalyses large amounts of cyclic AMP
• This disrupts cell signaling pathways, impairing cellular function and inducing apoptosis whilst allowing the bacterium to avoid phagocytosis
• It also inhibits the expression of interleukin 12 (IL12) an inflammatory cytokine

Dermonecrotic Toxin

• A type 3 exotoxin which acts intracellularly
• Precise mechanism is unknown however the overall effects on the cell include; actin reorganisation, increased DNA synthesis and binulcleation
• Its overall contribution to the pathogenesis of B. bronchiseptica is currently unknown

Not all these virulence factors are active at the same time however, only some are active depending on the temperature. At 37˚C all are active except the flagellum, at at 27˚C none are active except the flagellum. What this shows is that there is a global regulation of gene expression depending on the requirements of the bacteria’s survival.

For example, when B. bronchiseptica is required to colonise the respiratory tract (indicated by a 37˚C temperature i.e. body temperature) it is not going to need to use its flagella for movement so the genes encoding for flagella motion are not expressed. In contrast when B. bronchiseptica is outside of its target destination e.g. the nasal passageway (indicated by a lower temperature) the flagella genes are expressed again as movement will be required to reach the target destination (the airways). Genes encoding for virulence factors will not be expressed as there is no need when the bacterium is not at its target site of pathogenesis.

Treatment of Kennel Cough

Kennel cough is susceptible to many antibiotics (it is however resistant to erythromycin). There are bivalent canine vaccines available which vaccinate against B. bronchiseptica and canine parainfluenza virus. Many of B. bronchiseptica outer proteins are highly immunogenic which makes good vaccines (e.g. fimbriae, FHA, pertactin)

In Summary

M. haemolytica and B. bronchiseptica are two important veterinary pathogens as they both inhabit the same ‘niche’. This is interesting because they also share similar virulence factors. Both diseases result directly from the disruption of the respiratory epithelium and from immunopathology and both diseases are often polymicrobial, involving mutliple agents.

Diagnosis relies on isolation and cultivation of bacteria on agar plates after which the correct antibiotic treatment can be given. Vaccinations are available for both diseases.

These diseases show how the management of animals can influence disease epidemiology.

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