Brucella spp. Overview

Brucella spp. Overview

• Brucella spp. are aerobic, gram-negative, coccobacilli bacteria, almost cocci-shaped
• The bacteria are capnophilic and zoonotic
• Brucella are intracellular pathogens and subject to regulatory testing in animals
• Brucellosis is a notifiable disease; infection is persistent and lifelong

Brucella spp. Host Preferences

• Different Brucella species show host preferences but can infect other hosts
• B. abortus primarily infects cattle
• B. melitensis affects sheep and goats and has been eradicated from the US
• B. ovis infects sheep
• B. suis infects swine
• B. canis infects dogs
• B. pennipedialis infects seals
• Brucella does not affect birds

Rough vs Smooth Brucella spp.

• Virulence is partially determined by presence of LPS
• Smooth Brucella have O antigen polysaccharides
• Rough Brucella only have core polysaccharide
• The presence of these polysaccharides has implications for diagnostic serology testing

Brucella - Encounter and Transmission

• Brucella can be transmitted between carrier animals
• Modes of transmission include venereal contact
• Also via; placenta, birthing fluids, aborted fetuses, milk, blood, and urine
• Contact with mucous membranes, including ingestion, can also result in infection

Brucella as Intracellular Pathogens

• Brucella replicates in epithelial and endothelial cells
• Other cells include; dendritic cells, macrophages, monocytes, and microglia
• Inflammatory response to Brucella is pyogranulomatous, involving neutrophils and macrophages
• Infection can persist and live quiescently in the reticuloendothelial system

Brucella Pathogenesis

• Brucella is a true multisystemic pathogen
• It initially targets lymphatic tissues
• Brucella spreads systemically via the reticuloendothelial system and blood
• Locations may include uterus/testes, mammary glands, eyes/CNS, intervertebral discs, endocardium, liver, fetal tissues and joints
• Specific lesions depend on the Brucella species and host species
• Erythritol is preferentially used, and found in high concentrations in the placenta, mammary glands, and epididymis

Cellular Invasion and Replication

• Brucella enters cells via a zipper mechanism
• Phagosomes fuse with lysosomes and move to the ER
• Replication occurs in the ER
• Brucella then moves back to the cell membrane to release its payload
• T4SS is involved in directing the process in the ER

Spectrum of Disease by Species

• B. abortus natural hosts include Cattle and Bison
• The natural disease can include abortion, orchitis and epididymitis
• Occasional incidental hosts include; Sheep, goats and pigs, sporadic abortion
• Horses may get bursitis ("Fistulous Withers," "Poll Evil")
• Humans, dogs, and cats may get intermittent fever ("undulant fever”), systemic disease
• B. melitensis natural hosts include; Goats and sheep with abortion, orchitis, arthritis
• Occasional incidental hosts include cattle causing sporadic abortion
• Humans can get severe fever/systemic disease
• B. suis natural hosts include pigs with abortion, orchitis, arthritis, spondylitis and infertility
• Occasional incidental hosts include humans and dogs, causing intermittent fever and systemic disease
• B. ovis natural hosts include sheep with epididymitis and sporadic abortion; natural infection only in sheep, not in humans
• B. canis natural hosts include dogs with abortion, epididymitis, diskospondylitis and infertility
• Occasional incidental hosts include humans with a mild systemic disease

B. Abortus (Bang's Disease)

• Bacteria are carried in macrophages to bloodstream (intermittent bacteraemia)
• Brucellae shed intermittently in milk
• Brucellae localized in spleen, supramammary lymph nodes and lymphatic tissues
• Brucellae will be present in the fetus, placenta, fetal fluids, and uterine discharge in a pregnant animal

B. Abortus in the United States

• There has been a coordinated state-federal program since the 1930s to eradicate
• Ongoing surveillance occurs at slaughter plant-associated labs
• Vaccination of heifers is performed
• There are regulations surrounding interstate transport and change of ownership in the US
• Enhanced surveillance is conducted in the Greater Yellowstone Area due to bison and elk endemicity

Serologic Screening of Bovine Brucellosis (U.S.)

• Serology involves many tests that can be confusing but are highly sensitive and specific
• Testing protocol: perform the Brucella Acidified Plate Antigen (BAPA) test or CARD test [State Lab]
• If samples are NOT NEGATIVE perform a Fluorescent Polarization Assay (FPA) [State Lab]
• If the FPA is NOT NEGATIVE perform a Complement-Fixation Test (CF) [Federal Lab]
• Animals with "NOT NEGATIVE" tests are called "REACTORS"

Brucella Abortus vaccines for cattle

• Use modified live vaccines
• Need TH1 CD4+ cells because of intracellular infection
• Strain 19 is a less virulent mutant
• It may cause abortion in pregnant cattle, orchitis, and arthritis
• It Interferes with serological testing and undermines surveillance testing
• The vaccine is no longer produced but still has approval
• The RB51 vaccine is an even less virulent mutant
• It does not interfere with testing (DIVA) and is a ROUGH mutant
• The vaccine rapidly clears from the body, usually within 2 weeks

Vaccination of Cattle

• Heifer calves need to be 4-12 months old to be vaccinated
• Cattle need to be permanently identified with an ear tattoo of Tamper proof ear tag
• Vaccination must be given by a federally accredited veterinarian

Treatment of Cattle and Other Food Animals

• Treatment of cattle and other food animals infected with brucella is impossible
• Negative status can not be assured
• Drugs are expensive and illegal to use
• Herd depopulation is necessary

Human Brucella Infection

• Human Brucella infection is known as "Undulant fever" or "Malta fever"
• A cyclic, flu-like febrile syndrome is caused with; chills, sweats, hepatosplenomegaly and lymphadenitis
• Veterinarians and abattoir workers are most affected
• Modified live vaccines and needle stick injuries can cause infection through exposure
• The disease is almost eradicated in the US and a major concern in developing countries

B. Canis

• The pathogenisis is the same as B. abortus
• The colony morphology is rough
• Higher prevalence associated with puppy mills in central U.S.
• It is notifiable in some states. Reporting depends on the regulations of the state vet.

B. Canis Testing Rationale

• The is generally tested;
• Pre-breeding screen for both sires and bitches
• Testing for clinical disease
• Diskospondylitis is the most common manifestation
• Orchitis
• Abortions
• Lymphadenopathy
• Uveitis

Diagnostic approach to B. Canis

• Serological testing is very sensitive for screening of breeding dogs and ruling-out testing of symptomatic dogs

Serological Tests

• Cornell multiplex antibody test (Luminex instrument)
• Rapid slide agglutination test (RSAT)
• ELISA (CDA starting to use – probably repurposed B. ovis ELISA)
• Tube agglutination test (TAT)
• Indirect fluorescent antibody test (IFA)
• Lateral flow test
• Confirmatory tests are more specific, like AGID (Agar gel immunodiffusion)
• Blood culture is important in dogs with clinical signs
• Rule-in some other causes of diskospondylitis, uveitis
• It is the most specific test but only about 50% B. canis infections are bacteremic at any given time
• Negatives are likely true negatives but false positives are common and must be confirmed with a different test

Agglutination tests (RSAT, TAT)

• Should employ 2-mercaptoethanol step
• IgM has low affinity and high avidity
• IgG has high affinity and low avidity
• 2-ME abolishes the effect of non-specific IgM or may help distinguish an early immune response
• Fewer false positives are seen

Management of B. Canis

• Recommended euthanasia is typically recommended
• Spay or neuter animal if not already done
• Dual antimicrobial therapy of; Doxycycline or Enrofloxacin combined with Aminoglycoside (e.g. Amikacin)
• Animals can never be assuredly non-carriers
• Owners must be educated of risk
• Children are the most at risk historically
• Many dogs with diskospondylitis have pain which does not resolve and is unmanageable.