Mycoplasma and Gram Positive Bacteria

Question 1

Briefly describe the aetiology of the non-haemotropic Mycoplasma infections seen in cats (and dogs)

Answer 1

• Mycoplasma sppare the smallest known free living prokaryote
• They have a relatively small genome, meaning they rely on an environment that can nourish the cells
  
  • They primarily exist on the mucosal surfaces of the respiratory and urogenital tract
• They lack a cell wall and are fragile outside of the host
• They are not susceptible to lysozyme and antibacterials that target the cell wall such as the penicillins
• They are able to survive variable time depending on environmental conditions off the host
  
  • From 7-14 days in dry conditions at 30 degrees

Question 2

Discuss the array of clinical disease that could be caused by Mycoplasma felis in cats

Answer 2

• Conjunctivitis
  
  • Most often seen in young cats
  • Most often seen in groups of housed kittens soon after weaning (loss of maternal antibodies)
  • More commonly isolated by PCR from feline conjunctivitis than FHV or chlamydophila
  • Serous discharge followed by sticky mucoid discharge
  • Hyperemic to oedematous conjunctiva
  • Rarely involves the cornea
• Respiratory Infections
  
  • Can be isolated from both healthy and ill cats from group-living households (ie. not all show signs)
  • Highly likely to be pathogenic in cats when positive and signs are present
  • Been isolated in the face of suppurative bronchitis and pneumonia
  • Rare reports of mycoplasma associated pyothorax have been reported
• Genitourinary infections
  
  • Unable to survive the osmotic conditions of normal feline urine
  • May infect the genital region
• Systemic infection - rare and would require immunosuppression
• MSK infection
  
  • Rare reports of mycoplasma polyarthritis with immunodeficiency
  • Single report of mycoplasmal arthritis after bite wound

Question 3

Discuss how to confirm a mycoplasma is involed in an infectious process

Answer 3

• Often commensals, so interpretation in mixed infection can be difficult
• Mycoplasma are not seen on routine microscopy
  
  • Inflammatory exudates are typically primarily non-degenerate neutrophils and non-odiferous
• Special culture media are required due to the fragile nature of the organism
  
  • Hayflick’s formula / broth
  • Characteristic fried egg appearance when grown in colonies
• PCR can be used to identify the specific organism in culture

Question 4

Discuss the aetiology of streptococcal infection in dogs and cats

Answer 4

• Streptococci Spp are gram positive, facultatively anaerobic bacteria
• They cause pyogenic localised to systemic infection
• The various strains are differentiated by their antigenic differences in cell wall carbohydrates
  
  • Alternatively, they are differentiated based on their ability to lyse erythrocytes in culture
• Generally, the pathogenic strains are beta-haemolytic
• Steptococcus most commonly contributes to various infections of the upper respiratory tract infections, urinary tract, and skin.
• Necrotising fasciitis and toxic shock syndrome is the most relevant clinical disease, most often caused by S canis

Question 5

Provide an overview of the most clinically relevant infections caused by Group C Streptococcal bacteria

Answer 5

• Group C Streptococcus have been identified in respiratory tract washings from healthy and sick dogs
  
  • When present, more severe lower respiratory tract signs were present
  • Acute haemorrhagic and purulent pneumonia has been reported - most often S. equi ssp. zooepidemicus
  • ​Histopathology findings from severely affected animals suggests an exuberant inflammatory response similar to toxic shock

Question 6

Provide an overview of the most clinically relevant infections caused by Group G Streptococcal bacteria

Answer 6

Cats:

• Group G Streptococcal (most are b-haemolytic)
  
  • Can cause severe infections in kittens, opportunistic in older cats
  • Abscess, pharyngitis, pneumonia, lymphadenitis, diskospondylitis, osteomyelitis, arthritis can all occur
  • Dermal infection can be seen concurrently to the more severe signs
  • Necrotizing fasciitis and toxic shock like syndrome can be seen with Group G b-haemolytic strep.
  • Embolic disease due to bacteremia is common

Dogs:

• Most isolates are S. canis
• Common commensal and can cause a vast array of clinical infections
• Neonatal infection formas a part of the fading puppy syndrome
• Haematogenous spread can lead to meningoencephalomyelitis, polyarthritis, endocarditis etc.
• The primary cause of necrotizing fasciitis in dogs and toxic shock syndrome
• Generally susceptible to Penicillins, erthromycin and clindamycin

Question 7

Discuss the pathogenesis of necrotizing fasciitis and myosytis

Answer 7

• Most cases in dogs are caused by b-haemolytic group Streptococcus spp.
• Alterations in the M-protein gene associated with surface fibrillar material increases invasiveness
  
  • This alteration inhibts phagocytosis by neutrophils and macrophages
• The bacteria produces a family of highly mitogenic superantigens
  
  • These antigens simultaneously bind to MHC II and T cell receptors
  • This binding results in sudden high cytokine levels
  • Early use of enrofloxacin may induce bacteriophage lysis of S canis
• Most causes of NFM start with a localised infection - bite wound, urinary tract or respiratory tract infection
• Marked, rapidly progressive cellulitis is the initial sign

Question 8

Describe the clinical disease, necrotizing fasciitis and myositis caused by Streptococcus canis

Answer 8

• Initial infection / wound
• Local severe, rapidly progressive cellulitis
  
  • Fever (cats may be afebrile)
  • Left shift with variable leukocyte count.
  • CK often elevated in cats due to myositis
  • Bacteria are readily identifiable in the fluid associated with the cellulitis
• Pulmonary interstitial densities may be seen on radiographs
  
  • Septic or hypercoagulable emboli
• Cytology and culture are essential early to help guide initial antibiotic therapy
• Surgical debridement of the wounds / fascia is sometimes recommended
• Toxic shock syndrome can develop rapidly after the onset of the cellulitis.
• Treatment involves:
  
  • Intensive fluid therapy
  • Parenteral antibiotics - penicillins, erythromycin, clindamycin
    
    • Clindamycin is a potent inhibitor of bacterial toxin synthesis, inhibits M-protein synthesis and supresses LPS induced monocytes synthesis of TNF
  • Pain relief - opiates
  • Plasma therapy when DIC or hypocoagulability is present
  • Heparin therapy may be of benefit with DIC

Question 9

Discuss the aetiology of Staphylococcus infection in dogs and cats

Answer 9

• Staphylococcus spp are facultatively anaerobic, gram positive, catalase positive bacteria that tend to grow in clusters
• Common commensal organism and classically an opportunistic pathogen
  
  • Large percentage of normal animals carry Staph.
  • Important cause of disease
• Staphylococcus pseudointermedius, S. aureus and S. schleriferi ssp coagulans are most common

Question 10

Discuss and describe the epidemiology of Staphylococcus pseudointermedius infection in dogs

Answer 10

• Isolated from 31-68% of healthy adults and up to 100% of puppies
• Likely transmitted from the dam to puppies within 8 hours of parturition
• Colonises many body sites including the nasal passage, oral cavity, skin and perinum
• Colonisation may be prolonged to permanent or transient and recurrent
• Disease due to S. pseudointermedius occurs at a low percentage and generally requires a predisposing cause
• Methicillin-resistant SP is an emerging and now common problem
  
  • While colonisation with MRSP does not cause disease more readily that non-MRSP, treatment options are limited
  • Risk factors include recent surgery, prior antibiotic exposure and recent hospitalisation all increase risk

Question 11

Discuss the pathogenesis of Staphylococcal spp infection in dogs and cats.

Discuss the various known virulence factors

Answer 11

• Staphylococcus sppare common commensals
• Numerous virulence factors have been identified that enable colonisation, allowing the organism to await an opportunity to cause disease
• Virulence factors: mostly locally acting
  
  • Adhesins - numerous described
    
    • Staph. aureus - protein A binds to IgG and enables evasion of the host immune response
  • Clumping factor
  • Fibronectin binding proteins
  • Collagen binding proteins
  • Various enzymes are produced - can enhance host tissue breakdown during invasion
    
    • eg. hyaluronidase
  • Toxins
    
    • Alpha toxin - damage to host cell membranes
  • Proteases - can cleave Ig, defensins and platelet derived microbicidal peptides
• Distant acting virulence factors include
  
  • Enterotoxin
  • Exfoliative toxins - responsible for scalded skin syndrome and can cause toxic shock syndrome
• Can form biofilms
  
  • extracellular polysaccharide network that helps bacteria evade the host immune response

Question 12

Describe the mechanism by which Staphylococcus spp acquire or develop

Answer 12

• Staphylococcus spp have the ability to acquire resistance factors and mutate during a course of treatment
• Resitance can develop after horizontal transfer of chromosome, plasmids or transposons
• Resistance can occur in vivo due to genetic mutation
• Methicillin-resistant Staphylococcus are resistant to beta-lactams
  
  • Produce an altered form of the penicillin-binding-protein in the cell wall
  • Mediated through the mecA gene on the chromosomal casette
• Multi-drug resistance can also develop
  
  • May be due to the production of drug-efflux pumps
  • Enzymes that cleave the drug
  • Proteins that can bind and protect the drug target site

Question 13

Discuss the aetiology of gram negative bacterial infections in dogs and cats

Answer 13

• The major gram negative bacteria of clinical relevance include
  
  • E coli, Proteus spp and Klebsiella spp.
    
    • ​Natural inhabitants of the intestinal tract of mammals
  • Pseudomonas aeruginosa
    
    • ​Soil, water, decaying vegetation and animals
• Non-spore forming, non-acid fast, facultative anaerobic gram negative rods
• The outer membrane consists primarily of lipopolysaccharide (LPS)
  
  • The lipid component (lipid A) is the active component of endotoxin
  • The polysaccharide portion is the O antigen

Question 14

Discuss the various virulence factors of gram negative bacteria and note how they impact the pathogenicity

Answer 14

• Most gram negative bacteria are opportunistic pathogens
  
  • Some are true pathogens
• Virulence factors include:
  
  • Adhesins
    
    • Small molecules located on bacterial pilli
    • Allow for adhesion to the host cells
  • Toxins - numerous
  • Iron acquisition systems - siderophores
  • Capsules
    
    • Composed of polysaccharides - protect from the membrane attack complex of complement
    • Inhibit attachment of phagocytic cells
• It is the set of virulence factors, rather than the strain that determine pathogenicity

Question 15

Briefly describe the various virulence factors that enable Pseudomonas spp to establish an infection within a host

Answer 15

• Fibronectin coat is damaged by mechanical trauma, infection, maceration or other means
• Pili and exozyme S promote attachment to the epithelial cells
• Extracellular slime of the bacteria is anti-phagocytic
• Siderophores help the bacteria compete for iron stores with the host
• Varied toxins are also produced to promote deeper tissue infection by damaging the normal cellular structure and extracellular matrix
  
  • Collagenase
  • Lipase
  • Proteases
  • Fibrinolysis
• LPS on the bacterial surface is a potent trigger of numerous cytokine cascades and the complement cascade

Question 16

Describe the different virulence factors that lead to the varied presentation of gastrointetinal tract infection cause by E coli.

Answer 16

1. Enteropathogenic E coli (EPEC)
   
   • Adheres to mucosal cells of the SI and LI
   • Causes attaching and effacing lesions to the microvilli
   • A secretion system that stimulates the host cell to attach to the bacteria using its own actin
2. Enterohaemorrhagic E coli (EHEC)
   
   • Similar attachment and effacement as EPEC
   • Preferentially localise in the distal ileum and colon
   • Secrete effectors that encourage formation of a strong bonding “pedestal” below the bacteria
   • Produce Shiga toxin and Shiga-like toxin which are released on bacterial lysis
     
     • Attaches to stx receptors primarily on renal endothelial and GIT epithelial cells
     • Causes haemorrhagic diarrhoea
     • Shiga-like toxin in the enterocyte down regulates the inflammatory response
3. Enterotoxigenic E coli (ETEC)
   
   • Adhere to SI mucosal cells and elaborate toxins
   • Adherance is mediated by colonization factors and adhesins
   • Toxins - heat-stable and heat-labile lead to a secretory type diarrhoea without histopathological change
4. Adherent Invasive E coli (AIEC)
   
   • Contain an adhesin associate gene that enables invasion of the epithelial cells
   • Adherance is primarily in the colon
   • Adherance stimulates production of TNF and INF-g which promote inflammation and further enhacne adherance
   • They can translocate into the macrophages of the lamina propria, stimulting large concentrations of local TNF-a
5. Necrotoxigenic E coli
6. Enteroaggregative E coli
7. Enteroinvasive E coli

Question 17

Describe the virulence factors of uropathogenic E coli that allow them to colonise the urinary tract

What characteristic may contribute to at least some relapsing or recurent infections?

Answer 17

• Major determinant of pathogenicity is the ability to adhere to the urothelium via adhesins such as P fimbriae
  
  • After attachment, pili and endotoxin incite neutrophil migration and an inflammatory response
    
    • Urgency, discomfort, haematuria
• Other factors including exotoxin production, iron acquisition via siderophores, the bacterial capsule and anti-phagocytic systems
• Some UPEC strains can invade the uroepithelial cells and produce a biofilm-like complex within the cell. This may allow the bacteria to survive and evade antibiotics and the host immune system