The Family Pasteurellaceae

1. The Family Pasteurellaceae Haemophilus, Actinobacillus and Pasteurella Each genus will be described based on its general characteristics, how the disease is diagnosed, host(s) it infects, specific diseases it causes, and the properties that enable it to cause disease (virulence factors).

2. Gram positive Gram negative Outer membrane Cell wall Cell (inner) membrane

3. Gram negative bacteria

4. The Family Pasteurellaceae • Gram negative, bacilli or cocco-bacilli, 0.4 x 0.4-2.0 m • Non-motile (except Haemophilus) • Pyogenic • Aerobic/facultatively anaerobic • Biochemically active, most are oxidase & catalase positive • Many species have requirement for haemin, B vitamins or particular amino acids • Many species contain extracellular toxins

5. Haemophilus Small (0.5- 1 µm), gram-negative, coccobacilli Facultative anaerobic Capable of fermenting sugars Will not grow on MacConkey agar Fastidious nature - will not grow on some laboratory media, CO2 is sometimes required Haemophilus (meaning "blood loving") based on the requirement for X factor (haemin), or V factor (nicotinamide adenine dinucleotide), or both Found on the mucosal surfaces of animals and humans, and are extremely host-specific

6. Diagnosis Haemophilus spp are fragile and the specimens should be protected from drying and cultured within 24 hours. Most isolates from animals require only NAD, and they will not grow on blood agar, because NAD is destroyed by NADase present in blood. Isolates will thus grow either on chocolate agar or on blood agar with a disc of NAD or streak of Staphylococcus aureus that provides the NAD. Growth appears like the satellite (stars surrounding the moon). Colonies are small, dewdrop-like after 48h.

7. Other species: • - H. influenzae • - H. somnus

8. H. paragallinarum • fowl coryza. • Chickens primarily infected; turkeys and pigeons are resistant to infection. • Clinical signs: inflammation of the turbinates and sinus epithelium, disruption of the turbinates, acute air sacculitis. • infection is usually more severe following viral or mycoplasma infection.

9. H. parasuis • Glasser's disease • fibrinous inflammation of the serous surfaces or joints • may affect the pericardium, pleura, peritoneum, joints, and in severe cases, the meninges • Clinical signs include swollen joints and lameness, fever, pleuritis, and possibly meningitis.

10. H. somnus • spectrum of diseases in cattle • TME: Clinical signs include weakness, fever, staggering, somnolence, dyspnea, paralysis, and sudden death • The lesions consist of a fibrinous meningitis, with thrombosis and necrosis. • pneumonia with pleuritis and arthritis • Reproductive failure may be a result of endometritis, metritis, or late abortion. Mastitis and vaginitis may also occur

11. Virulence Factors Capsule. H. parasuis and H. paragallinarum produce well-defined, cell-associated capsular polysaccharides. Form protective coat around the bacterium, and allows it to evade host defenses. Exotoxins. H. somnus produces a weak haemolysin. Other factors: proteases: destroy mucosal IgA (H. influenzae) outer membrane proteins that bind to the Fc receptor of normal IgG and block binding of specific IgG (H. somnus) iron regulated outer membrane proteins (IROMPs). Under iron-limiting conditions the bacteria produce these outer membrane proteins that are able to obtain iron from the host

12. Actinobacilli Closely related to those in the genus Haemophilus. Gram-negative coccobacilli, pleomorphic, small (0.4 to 1.0 µm). Non-motile, non-acid fast. Fermentative without gas production. Most species will grow on MacConkey agar. The colonies of most species will be very sticky and difficult to remove from the agar. The normal habitat is the respiratory, alimentary, and urogenital tract. They may occur as commensal agents or as pathogens.

14. Diagnosis cells tend to be delicate and are easily destroyed by environmental effects such as drying or by disinfectants usually easy to isolate from lesions of infected material colonies tend to be sticky and difficult to remove from the plate A. pleuropneumoniae is beta haemolytic and CAMP positive. It requires V factor, and can be isolated on chocolate agar or on blood agar with a streak of Staphylococcus

15. Virulence Factors Capsules capsules are only produced by A. pleuropneumoniae Lipopolysaccharide all species contain LPS (endotoxin), with O side chains only the LPS of A. pleuropneumoniae has been documented to contribute to lesions Exotoxins extracellular toxins (RTX toxins) identified and characterized in A. pleuropneumoniae RTX toxins produced in A. lignieresii, A. equuli, and A. suis also, but their role in disease has not been defined

16. A.lignieresii • Slightly sticky non-haemolytic colonies formed on blood agar • Pink colonies on MacConkey agar (LF) • In smears of pus or granules clumps of organisms and club shaped bodies may be seen • Wooden tongue and rumenitis in cattle, skin abscesses in sheep

17. A. lignieresii vs A. bovis

18. A. pleuropneumoniae • Is the cause of contagious pleuropneumonia of swine, and is absolutely specific for swine. • Grossly, fibrinous pleuritis and necrotizing pneumonia. • Lesions may consist of consolidation and congestion, may be very haemorrhagic with necrosis. • Clinical signs include trembling, anorexia, dyspnea, fever, and haemorrhaging from the nose and mouth • Animals may die within 24 hours.

19. Pasteurella • Gram-negative, short to medium-sized coccobacilli (0.5 µm to 2.0 µm), bipolar staining • Most strains are facultatively anaerobic • Catalase and oxidase positive • Ferment glucose, gas is not produced • Only M. haemolytica is ß-hemolytic on blood agar. M.haemolytica and P. trehalosi will grow to some extent on MacConkey agar • Pasteurella species are normal commensals of the mucosal membranes of the respiratory tract of mammals

20. Diagnosis • Isolation is the gold standard • Gram or Giemsastain: bipolar organisms • P. multocida isolates grow best on blood agar, and have a characteristic sweetish odor. • Isolation of Pasteurella from the nasopharynx is of little significance because they are part of the normal flora. Bronchial lavage is more robust • Biochemical tests: • Urease negative • TSI stab results in yellow slant and butt with no gas or H2S

21. P.trehalosi P.haemolytica haemolytica trehalosi multocida Haemolysis ++- Fermentation of xylose +-- Production of indole --+ MacConkey agar Small pink colonies - Differentiation of P. haemolytica, P. trehalosi and P. multocida

22. Fowl cholera P. multocida • Mucoid colonies • Four main capsule types: A, B, D, E • Causes range of diseases: septicaemia, shipping fever in cattle, pleuropneumonia in sheep, atrophic rhinitis in pigs, and fowl cholera in poultry • Type D produces dermonecrotoxin

23. P. haemolytica • Disease usually results following stress, such as transport, inclement weather, castration, or predisposing infections. • Cattle: primary cause of shipping fever. Sheep: enzootic pneumonia. • Type T causes septicaemia • Produce leukotoxin P. trehalosi Causes acute lamb septicemia in autumn and winter

24. Virulence factors Hyaluronic acid capsulePoorly immunogenic, because it resembles the host hyaluronic acid Lipopolysaccharide LPS is thought to play a role by two mechanisms. 1) protect the organism from host defenses through the carbohydrate moiety; 2) induce inflammation through the lipid A component. ExotoxinsA dermonecrotoxin is produced by type D and some type A strains of P. multocida and is considered an important virulence factor in the pathogenesis of atrophic rhinitis. M. haemolytica produces a labile leukotoxin (RTX toxin) important in the pathogenesis of shipping fever and respiratory disease.