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RUSVM Bacteriology Gram Negatives > Brucella > Flashcards

Flashcards in Brucella Deck (115):
1

Brucella Gram____? Shape?

Gram negative, small, coccobacilli (more coccoid) Pairs or clusters

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Brucella Motile/Non-motile?

Non-Motile

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Brucella Capsulated?

Non-encapsulated

4

Brucella Flagellated?

No flagella

5

Brucella Spore-forming?

Non-spore forming

6

Brucella Name the dominant surface antigens in LPS in rough versus smooth colonies

Rough: "R" antigen Smooth: "A" abortus or "M" melitensis antigens

7

Brucella What is significant about the cell wall?

Peptidoglycan layer = thicker than other Gram negative (modified acid fast stains for diagnosis)

8

Brucella Distribution?

Worlwide

9

Brucella Stable in environment?

No, obligate pathogen Survives 4 months in milk, urine, water, damp soil but NEEDS MAMMALIAN HOST

10

Brucella Often chronic or acute infections?

Mainly chronic infections

11

Brucella Target site

Reticuloendothelial system & genital tract Abortion & epididymitis/orchitis

12

Brucella Pathogenic species

*B. melitensis, B. abortus, B. suis* B. canis, B neotomae, B. ovis B. cetaceae, B. pinnipediae

13

Brucella Reportable?

YES REPORTABLE

14

Brucella Virulence factors

Cell Wall Erythritol Outer membrane proteins Superoxide dismutase & catalase T4SS

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Brucella Erythritol role

4 carbon sugar alcohol *Allantoic fluid factors* (stimulates growth) Preferential location for reproduction & mammary glands

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Brucella Omp (Outer membrane protein)

Stimulate delayed type hypersensitivity

17

Brucella Superoxide dismutase and catalase function

Defense against oxidative killing

18

Brucella T4SS function

Intramacrophage survival

19

Brucella Resistance factors

Survives freezing & thawing Survives in milk & other fluids Pasteurization kills it

20

Brucella Diversity

Genome very stable Different species vary in host preference & degree in virulence

21

Brucella What type of colonies are more virulent? Smooth or rough?

Smooth more virulent than rough Depends on polysaccharide side chain in LPS

22

What factors change Brucella host preference and virulence?

CO2 requirements H2S production Urease production Susceptible to different concentrations of certain dyes Susceptibility to bacteriophage (Tbilisi phage)

23

How does Brucella abortus differ from other strains?

B. abortus is susceptible to Tbilisi phage, others are resistant

24

Brucella Zoologic & Geographic reservoirs

Obligate parasite Host preference (some broad range) Survival time in environment depends on temperature & moisture

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B. abortus

Cattle Smooth colonies, virulent Bison, camels, yaks

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B. suis

Swine Smooth colonies, virulent Hares, reindeer

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B. melitensis

Goat & Sheep Smooth colonies, virulent Camelids

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B. neotomae

Wood Rat Smooth colonies, virulent

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B. ovis

Sheep

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B. canis

Dogs

31

Brucella Modes of transmission

Ingest cow/goat milk In utero (direct) Venereal Urine (dogs) Insects (maybe, minor)

32

Describe Brucella's venereal transmission

Male sex gland infections Dissemination occurs without lesions Common in swine, sheep, dogs

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Brucella Pathogenesis

Penetrates intact mucosa--> Macrophage--> survives --> Regional lymph node --> hematogenous dissemination & localization in reticulo-endothelial system & repro tract --> Abortions

34

How do Brucella species survive inside macrophages?

T4SS, Superoxide dismutase, catalase

35

How do Brucella species cause abortions?

Interference with fetal circulation / placentitis Endotoxin Fetal stress / Inflammation

36

What type of inflammation is shown in the picture?

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Granulomatous inflammation!

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37

What can cause Brucella infections to move extra-genitally? (Disseminate)

Prolonged bacteremia

38

Brucella Pathology

Grossly visible lesions in placenta Intercotyledonary thickening with yellow gelatinous fluid Necrotic cotyledon with thick brown exudate Aborted edematous fetus Male genitalia

39

Describe Brucellosis in males

Palpable enlargement of epididymus Increase neutrophils in semen (acute cases) Orchitis Scrotal dermatitis in dogs Results: decrease fertility, sometimes sterility

40

Describe orchitis from Brucellosis in males

Necrosis in testicular parenchyma with pus, prostatitis, fibrinopurulent, seminal vasculitis

41

What is shown in the picture? What species caused it?

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Necrotic cotyledons in aborted material. B. melitensis

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42

Brucella Disease Pattern Female reproductive tract

Abortion (most commonly) Retained placenta Mastitis (goats)

43

Brucella When will it cause cattle to abort?

~5month gestation

44

Brucella When will it cause swine to abort?

At any time

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Brucella When will it cause dogs to abort?

~50 days gestation

46

Brucella Repeat abortions or lasting immunity?

Females usually abort once Acquired immunity

47

What does this picture show?

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Brucella melitensis Stillbirth fetus with necrotic placenta

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48

What do these pictures show?

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Orquitis - Swelling & irregular conformation of tail of epididymis of ram infected with Brucella ovis.

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49

Brucella Extragenital manifestations in Swine

Arthritis Lumbar spondylitis Tissue necrosis Posterior paralysis

50

Brucella Extragenital manifestations in Dogs

Meningoencephalitis Osteomyelitis Discospondylitis Anterior uveitis Ocular manifestations

51

Brucella Extragenital manifestations in Cattle

Hygromas - Chronic (false bursa, on joint sacs)

May be secondary to abortion

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52

Brucella Extragenital manifestation in Horses

Poll evil or fistulous withers

53

Brucella Extragenital manifestation in Humans

Undulant Fever Malta Fever

54

What body system does undulant fever affect?

Reticulo-endothelial system

55

Common signs of Undulant Fever

Mild lymphadenopathy, splenomegaly, hepatomegaly 2-3 weeks post-exposure Fever, chills, night sweats, fatigue, muscle & joint pain, backaches, depression, insomnia, arthritis, osteomyelitis, endocarditis

56

How do humans get Undulant fever?

Handling infected tissues Often: aborted fetuses, placentas, post-abortion uterine fluieds, milk, milk products Human-human rare

57

What human professions are at risk for Brucellosis?

Veterinarians, ranchers, slaughterhouse workers

58

What Brucella species can affect humans?

B. melitensis** B. suis B. abortus B. canis

59

Which Brucella species do not affect humans, generally?

B. ovis B. neotomae (always questionable)

60

What predisposes an animal to a Brucellosis infection?

Younger more resistant Sexually mature animals MORE susceptible, (infected for life possible) Higher herd size & density

61

What helps SPREAD Brucellosis in swine herds?

New boars Confine breeding swine in common pens Feral swine

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What helps CONTROL Brucellosis in swine herds?

Separate the breeding pairs Quarantine boars Limit exposure to feral population Eliminate infected boars Minimize exposure to aborted tissue

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How is Brucellosis often spread in sheep herds?

Dissemination during breeding season Older rams more likely infected Homosexual rams spread it

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What are common differentials to Brucellosis in older rams?

Actinobacillus seminis Histophilus somni

65

How is Brucellosis spread in dogs?

Close confinement/ Kennels Urine!

66

Brucella Immune mechanisms in pathogenesis

Antibodies = protective & detrimental Phagocytes disseminate the organism

67

Describe the role of antibodies in a Brucella infection

Low levels= complement High levels = opsonization IgA autoantibodies in dogs

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What class of immunoglobulin are used in low level antibody production?

Mainly initially IgM and low IgG (Cause complement-mediated lysis)

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What class of immunoglobulin are used in high-level antibody production?

High levels of IgG Survive inside phagocytes Interfere with complement and opsonize bacteria

70

Brucella Immunologic aspects Mechanisms of Resistance & recovery

Effective immunity = **cell mediated** Macrophages & NK cells

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What cytokines are released in cell mediated immunity against Brucella infections?

IL-2, -12, -18 IFN-g TNF

72

When will an animal develop the most effective immune response to Brucella?

**Better response if infected prior to sexual maturity**

73

Brucella Vaccinations in cattle

B. abortus 45/20 (killed, 2 doses) B. abortus 19 & RB51 (live-attenuated, 1 dose, 4-8mo old)

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What can happen if you vaccinate a full grown bull for B. abortus?

Orchitis

75

What are some problems with the B. abortus vaccines in cows?

Abortions Serological false positives

76

Brucella vaccines for Goats and Sheep

B. melitensis Rev 1 (Live attenuated) B. melitensis H38 (killed) -Sometimes protects against B. ovis

77

Are there vaccines for B. suis or B. canis?

No

78

Can vaccines for B. melitensis be used in eradicated countries?

Cannot be used in countries free of B. melitensis because of serological test interference

79

What is important about submitting suspected Brucella specimens?

Biosafety level 3 practices (prevent aerosolization) Submit to reference labs!

80

What samples should be collected for Brucella lab diagnosis?

Abscess material, semen, vaginal fluids, milk samples, blood (dogs, bacteremia of B. canis) Necropsy & Abortion material

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What necropsy material is collected for Brucella lab diagnosis?

Spleen, liver, udder, lymph nodes, epididymis, testicle, accessory male sex organs

82

What abortion material is collected for Brucella lab diagnosis

Abomasal fluids Lungs of aborted fetus Placenta

83

What direct exam methods are used for Brucella Lab diagnosis?

Gram Stains Modified Gram Stains Modified Ziehl-Neelsen Macchiavello stains

84

Brucella Lab Dx Describe Gram Stains (fetal contents vs. semen)

Fetal contents reveal high # gram negative cocci Few numbers of bacteria in semen

85

Brucella Lab Dx Modified Gram Stain

Use carbol fuschin instead of safranin

86

Brucella Lab Dx Modified Ziehl-Neelsen (with good differentials)

0.5% Acetic acid to destain Stain Red DDx:: Coxiella burnetii and Chlamydophila abortus

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Brucella Lab Dx Macchiavello stains

Mainly from fetal stomach content & placenta Stain red

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Brucella Lab Dx Microscopy

Gram stain & immunofluorescence

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89

Brucella Lab Dx Isolation - samples used?

Tissues Fluids Milk (cream & sediment)

90

Brucella Lab Dx Isolation - Agars used?

Serum dextrose Tryptose Columbia agar w/ 5% serum Brucella agars

91

Brucella Lab Dx Isolation - Mediums used?

Farrell's medium Thayer-martin modified medium

92

Brucella Lab Dx Isolation - Mediums - Conditions for culture

Aerobic environment, 37C, 10% CO2, 10-21 days Bluish color under obliquely transmitted light

93

What is the most sensitive method for Brucella laboratory diagnosis?

Animal inoculation

94

Describe animal inoculation in Brucella lab diagnosis

Necessary when low # are present Guinea pigs are most sensitive Test 3-6 weeks post inoculation

95

Describe the colonies on blood agar

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Brucella Gram negative, coccobacilli, non-hemolytic Catalase & oxidase positive Urease positive (except B. ovis) No fermentation of glucose or lactose

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96

Brucella species and biovar determinations?

Urease production CO2 requirements H2S production Oxidation of metabolic substrates Agglutination in mono-specific antisera Growth in presence of varying concentration of thionin Basic fuschin Phage typing

97

Molecular techniques in Brucella Lab diagnosis

PCR, real-time PCR, RLFP PCR Strain 19 - deletion in ery locus Strain RB51 - Insertion in wbo A gene

98

What makes strain 19 different from other Brucella strains.

Deletion in the ery locus

99

What makes strain RB51 different?

Insertion sequence in wbo A gene

100

What does the Brucellin Test test?

Type 4 Hypersensitivity

101

What tests for B. ovis best?

Agar gel immunodiffusion

102

Brucella Immuno Dx Rose-Bengal plate test

Antigen suspension, pH=3.6, allows IgG1 agglutination Positive results need confirmation

103

Brucella Immuno Dx Brucella milk ring test

Purple ring develops in top with cream (picture)

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104

Brucella Immuno Dx Confirmatory tests

CFT - complement fixation test Indirect ELISA

105

Brucella Immuno Dx For canine brucellosis

Rapid slide agglutination test

106

Brucella Immuno Dx Which species share antigens?

Antigens shared with Y. enterocolitica O:9 and E. coli O:157

107

Brucella Control & Prevention

Depends on animal species Brucella Species Husbandry Availability/Efficacy of vaccines

108

Brucella Treatment Livestock

No treatment in livestock High failure Cost Problem maintaining infected during eradication program

109

Brucella Treatment Rams with B. ovis

Prior to palpable epididymal lesions, abscesses or fibrosis: Tetracycline or dihydrostreptomycin After these lesions - difficult to treat

110

Brucella Treatment Dogs

Neuter affected animals Antibiotics (tetracycline & dihydrostreptomycin) Cull males in breeding program Don't treat breeding colonies

111

Brucella Prevention Immunization alone

Reduces # abortions No eradication

112

Brucella Prevention Bovine Brucellosis Immunization THEN test & slaughter (Tests in dairy vs. beef)

Vaccinate young cattle, test @ sexual maturity Seropositives culled & slaughtered -Ring test in dairy -Blood test in beef cattle

113

Brucella Prevention Testing & removal without immunization

Depopulation of entire swine, sheep, goat population Re-stock

114

How often Rams palpated?

Rams tested twice per year by palpation

115

Brucella Prevention Dogs (# tests needed for "free" status)

Dogs tested before breeding Need 3+ seronegative test results for "free" status