Brucella

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Brucella

• The genus Brucella consists of six species, four
  of which cause human brucellosis ???? Brucella
  melitensis ????, Brucella suis ????, Brucella
  abortus ????, and Brucella canis ????
• Are all intracellular organisms
• B. neotomae B. ovis

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• Brucella are small (0.40.8 0.51.5µm),
  non-motile, non-capasulate, gram-negative
  coccobacilli.
• The organism is aerobic, and their nutritional
  requirements are complex.
• All strains grow best in a medium enrich with
  animal serum and glucose
• 5-10 carbon dioxide

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Antigenic Structure and classification

• Two main antigen A and M
• The three main Brucella differ from one another
  in the amount or the two main antigen they have
  in common
• B.abortus AM201
• B.melitensis AM120
• B.suis AM21

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B. abortus

• Bacteria is excreted in genital secretions
  (including semen), milk, colostrum.
• Survival time
• Cheese at 4oC 180 days !!!
• Water at 25oC 50 days
• Meat and salted meat 65 days
• Manure at 12oC 250 days !!!!
• Widespread Cattle, Bison, Elk, Deer, Moose,
  Horse, Sheep, Goat, Swine, Donkey, Dogs, Birds,
  Hares, Fox, Rats, mice, Camels and Human.

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Spread of Brucella in the body
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Incubation period

• Acute or subacute disease follows an incubation
  period which can vary from 1 week to 6 or more
  months.
• In most patients for whom the time of exposure
  can be identified, the incubation period is
  between 2 and 6 weeks
• The length of the incubation period may be
  influenced by many factors
• virulence of the infecting strain
• size of the inoculum
• route of infection
• resistance of the host

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Portals of entry

• Oral entry - most common route
• Ingestion of contaminated animal products (often
  raw milk or its derivatives)
• contact with contaminated fingers
• Aerosols
• Inhalation of bacteria
• Contamination of the conjunctivae
• Percutaneous infection through skin abrasions or
  by accidental inoculation

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Clinical Manifestations

• The presentation of brucellosis is
  characteristically variable
• The onset may be insidious or abrupt
• Influenza-like with fever reaching 38 to 40oC
• Limb and back pains are unusually severe, night
  sweating and fatigue are marked.
• Anorexia, weakness, severe fatigue and loss of
  weight, depression
• Headache
• The leukocyte count tends to be normal or
  reduced, with a relative lymphocytosis
• Relative leukopenia
• On physical examination, splenomegaly may be the
  only finding.

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COURSE OF BRUCELLOSIS

• If the disease is not treated, the symptoms may
  continue for 2 to 4 weeks
• Many patients will then recover spontaneously
• Others may suffer a series of exacerbations
• May produce an undulant fever in which the
  intensity of fever and symptoms recur and recede
  at about 10 day intervals.

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Brucellosis

• Cyprus fever/Gibraltar fever/Malta fever/Rock
  fever/Undulant fever

• Most affected persons recover entirely within 3
  to 12 months
• Some will develop complications
• involvement of various organs,
• a few may enter an ill-defined chronic syndrome.

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COMPLICATIONS

• Arthritis, often sacroiliitis, and spondylitis
  (in about 10 percent of cases)
• central nervous system involvement including
  meningitis (in about 5)
• Uveitis, epididymo-orchitis
• Endocarditis very rare
• In contrast to animals, abortion is not a feature
  of brucellosis in pregnant women.

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LARGE JOINTS
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SPONDYLITIS???
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SACROILIITIS ?????
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Chronic Brucellosis- Depression
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Population risk

• The main source of infection for the general
  population is dairy produce prepared from
  infected milk.
• B. melitensis presents the greatest hazard.
• The milk of infected sheep and goats may contain
  large numbers of viable organisms, which become
  concentrated in products such as soft cheeses.
• Indeed, soft cheese has been recognized as a
  major vehicle of infection in the Mediterranean
  region, the Middle East and Latin America

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Occupational hazard

• Infection arises from occupational or domestic
  contact with infected animals or with an
  environment contaminated by their discharges
• Farmers and their families, abattoir workers,
  butchers and veterinarians are particularly at
  risk
• Infected animals that have recently aborted or
  given birth present the greatest hazard

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Extending spectrum of zoonosis

• The recent isolation of distinctive Brucella
  strains, tentatively named Brucella maris, from
  marine animals in the United Kingdom and the
  United States extends the ecologic range of the
  genus and, potentially, its scope as a zoonosis
• seals, sea otters, dolphins and porpoises
• An incident of laboratory-acquired infection
  suggests that this type is pathogenic for humans
• Infection could result from occupational contact
  with infected seals or cetaceans.

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CLINICAL DIAGNOSIS
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Sanitary

• Pasteurization of dairy products and use of
  protective clothing prevent human infection. More
  importantly, systematic identification and
  elimination of infected animals and vaccination
  of animals reduces the reservoir.

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Prevention

• Eradication of brucellosis in cattle can be
  attempted by test and slaughter,active
  immunization of heifers with avirulent live
  strain 19,combined testing,segregation, and
  immunization.Cattle are examined by means of
  agglutination tests
• Active immunization of humans against brucella
  infection is experimental.Control rests on
  limitation of spread and possible eradication of
  animal infection,pasteurization of milk and milk
  products, and reduction of occupational hazards
  wherever possible.

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Treatment

• Brucella may be susceptible to tetracyclines or
  ampicillin.
• Symptomatic relief may occur within a few days
  after treatment with these drugs is begun.
• However ,because of their intracellular
  location,the organisms are not readily eradicated
  completely from the host.
• For best results,treatment must be
  prolonged.Combined treatment with streptomycin
  and a tetracycline may be considered

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YERSINIA
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Genus Yersinia
Enterobacteriaceae
Genus Escherichia

• Y. pestis
• Y. enterocolitica
• Y. pseudotuberculosis

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Biological Features

• Small, 0.5-0.8 µm in width
• 1.0-2.0 µm in length.
• Gram-negative rods.
• Sometimes appearing as
• cocco bacilli.
• Bipolar StainingRetaining stain at the ends of
  cells.

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Biological Features

• Cultural Features
• Facultative anaerobes.
• Optimal growth temperature range form 28C to
  30C.
• Optimal growth pH 6.97.2.
• Growth is more rapid in media containing blood or
  tissue fluids.
• Nonmotile when grow at temperatures above 30 C.

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Pathogenicity

• Transmission

Flea
Bite
Respiratory Tract
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Antigenic Structure

• F1 Antigen
• V,W Antigen
• Yersinia Outer membrane Protein (Yop)
• Murine Toxin (MT)
• 0.3-0.4 formaldehyde Toxoid
• Endotoxin (LPS)

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Y. Pestis Virulence factors schematic diagram
Plasmid
Plasmid
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Pathogenesis
Lymph Nodes In Groin and Axilla
Invade
Phagocyte
Y. pestis
Enter Respiratory System
Pneumonic Plague
Bubonic Plague
Invade Blood Stream
Septicemic Plague
meningitis
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Pathogenicity

• Clinical Forms
• Bubonic Plague High fever, Swelling, Bleeding,
  Necrosis of lymph nodes
• Pneumonic Plague chills, cough, respiratory
  failure, circulatory collapse Black Death
• Septicemic Plague Fever (39-40 C) , Shock , DIC

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Y.Enterocolitica Y.Pseudotuberculosis

• Gram negative, No capsule, No spore, Facultative
  anaerobes
• V-W antigen
• More than 50 serotypes of Y.Enterocolitica
• 6 serotypes of Y.Pseudotuberculosis
• Diseases
• Gastroenteritis
• terminal ileitis, appendicitis, mesenteric
  lymphadenitis,
• dermatitis contusiformia, arthritis
• Septicemia
• Sanitary precautions, Antibiotic

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Epidemiology

• Plague
• Probably originated in Asia or central Africa.
• One of the earliest record pandemics occurred in
  542 B.C.
• Three pandemics in the history.
• 1989-19985440 cases, 681 dead.

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Immunity
Cellular Immunity
Humoral Immunity
Antibody To
1) F1 Ag
2) V,W Ag
Phagocytose
Promote phagocytose , agglutinate and kill
bacteria
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Diagnosis

• A. Specimens
• Aspirates of lymph nodes
• Cerebrospinal fluid
• Blood
• Sputum

• B. Smears
• Giemsas stain
• immunofluorescent stain

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Diagnosis

• C. Culture
• All materials Cultured on blood agar and
  MacConkeys agar and in infusion broth
• Positive in 24 hours
• Tentatively identified by biochemical reations
  Definite identified by immunofluorescence

CAUTION All cultures are highly infectious and
must be handled with extreme caution
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Diagnosis

• D. Serology
• In patients who have not been previously
  vaccinated, a convalescent serum antibody titer
  of 116 or greater is presumptive evidence of
  Y.pestis infection.A titer rise in two sequential
  specimens confirms the serologic diagnosis.

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Treatment

• Streptomycin
• Tetracycline
• alternative drug
• combination with streptomycin
• essential for control early in disease
• Sulfonamides

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Summary of Yersinia infections
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Bacillus
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Spore-Forming Gram-Positive Bacilli
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Bacillus Species

• At least 48 species are known but only
• B. anthracis and B. cereus cause defined diseases
  in humans.
• B. anthracis is responsible for the disease
  anthrax.
• This is a disease primarily of animals
  (zoonosis) but humans can acquire via handling,
  inhaling or ingesting contaminated animal
  products.
• B. cereus is predominantly responsible for food
  poisoning in humans.
• Bacitracin and polymyxin are two well-known
  antibiotics obtained from Bacillus species.
• Spores of many Bacillus species are resistant to
  heat, radiation, disinfectants and desiccation

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• It was from studies on anthrax that Koch
  established his famous postulates in 1876
• Pasteur (1881) developed a vaccine against
  anthrax

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B. anthracis Gram staindemonstrating spores
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B. anthracis, Colony on SBA
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STICKY Consistency ofB. anthracis Colony on
SBA
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Anthrax infections are classified by route of
entry

• Cutaneous
• Gastrointestinal
• Respiratory

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Cutaneous Anthrax

• gt 95 of naturally occurring cases
• Spores enter breaks in skin after contact with
  contaminated animal products
• Papule??- Vesicle??- Ulcer - Eschar??
• Up to 20 case fatality rate if untreated
• Mortality with treatment lt 1

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• After a 2- to 3-day incubation period, a small
  pimple or papule appears at the inoculation site.
  
• A surrounding ring of vesicles develops
• Over the next few days, the central papule
  ulcerates, dries, and blackens to form the eschar
  

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Vesicles Black Eschar
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Painless Edema

• The lesion is painless and is surrounded by
  marked edema that may extend for some distance
• Pus and pain appear only if the lesion becomes
  infected by a pyogenic organism
• Similarly, marked lymphangitis????and fever
  usually point to a secondary infection.

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Evolution of an anthrax eschar in a 4-year-old
boyDAY 6
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DAY 10 - 15
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Evolution of an anthrax eschar in a 4-year-old
boy. (AB) the lesion when first seen (day
0).Note the arm swollen from the characteristic
edema. (C) Day 6. (D) Day 10. (E) Day 15.
Although penicillin treatment was begun
immediately and the lesion was sterile by about
24 hours, it continued to evolve and resolve as
seen.
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Cutaneous anthrax Differential diagnosis
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Cutaneous anthrax

• For cutaneous and gastrointestinal anthrax,
  low-level germination occurs at the primary site,
  leading to local edema and necrosis

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Inhalation

• Bacillus spores are inhaled and ingested by
  alveolar macrophages??????
• These cells carry the bacteria to the regional
  lymph nodes, causing necrotic hemorrhaging which
  leads to death

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Gastrointestinal

• Ingestion of contaminated meat produces systemic
  symptoms which can lead to death
• Mortality by gastrointestinal anthrax may be 50

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Gastrointestinal and pulmonary anthrax are both
more dangerous than the cutaneous form because
they are usually identified too late for
treatment to be effective
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PATHOGENESIS

• Anthrax infections result only if the bacteria
  produce a
• i) capsule (poly-y-D-glutamic acid polypeptide)
• ii) exotoxins
• both encoded on plasmids
• three proteins
• protective antigen (PA) (82. 7 kDa)
• lethal factor (LF) (90.2 kDa)
• edema factor (EF) (88.9 kDa)

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ANTHRAX TOXINS
20 kDa
PA
LF
EF
PA
Host Protease
The complex (PALF or PAEF) is internalized by
endocytosis acidification of the endosome the LF
or EF cross the membrane into the cytosol via
PA-mediated ion-conductive channels
PA
HOST CELL
LF
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Effects of anthrax exotoxins on macrophages

• Edema toxin is a calmodulin?????-dependent
  adenylate cyclase that increases intracellular
  levels of cyclic AMP (cAMP) on entry into most
  types of cell
• This is believed to alter water homeostasis
• resulting in massive edema

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Effects of anthrax exotoxins on macrophages

• Lethal toxin is a zinc metallo-protease that
  causes a hyperinflammatory condition in
  macrophages
• activating the oxidative burst pathway
• release of reactive oxygen intermediates
• production of proinflammatory cytokines
• responsible for shock and death.
• MAPKK denotes mitogen-activated protein kinase
  kinase

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Once they have been released from the
macrophages, there is no evidence that an immune
response is initiated against vegetative bacilli
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Protective immunity

• Antibodies against protective antigen
• Both the noncellular human vaccines and
  live-spore animal vaccines confer protection by
  eliciting antibodies to protective antigen
• The poly-g-D-glutamic acid capsule of B anthracis
  is poorly immunogenic, and antibodies to the
  polysaccharide and other components of the cell
  wall are not protective.

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Species differences

• Anthrax has been documented in a wide variety of
  warm-blooded animals
• Some species, such as rats, chickens, and dogs,
  are quite resistant to the disease
• Others (notably herbivores such as cattle, sheep,
  and horses) are very susceptible
• Humans have intermediate susceptibility.

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reservoir of B anthracis is contaminated soil

• Spores remain viable for long periods
• Herbivores, the primary hosts, become infected
  when foraging in a contaminated region
• Because the organism does not depend on an animal
  reservoir, it cannot readily be eradicated from a
  region
• anthrax remains endemic in many countries
• Humans become infected almost exclusively through
  contact with infected animals or animal products

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Cycle of infection in nature

• As a susceptible animal with anthrax approaches
  death, its blood contains as many as 109
  bacilli/ml
• Necrosis of the walls of small blood vessels
  during the acute phase of the illness leads to
  hemorrhages and to characteristic bloody
  exudations from the mouth, nose, and anus, a
  highly diagnostic sign
• These exudates carry vast numbers of the bacilli
• sporulate on exposure to air
• produce a heavily contaminated environmental site
  
• potentially capable of infecting other animals
  for many years

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Handling of carcasses????

• Sporulation of B anthracis requires oxygen
• therefore does not occur inside a closed carcass
• regulations in most countries forbid postmortem
  examination of animals when anthrax is suspected
• The vegetative cells in the carcass are killed in
  a few days by the process of putrefaction.

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• In endemic areas, animals that die suddenly
  should be handled cautiously
• Livestock should be vaccinated annually.

Do I look that I am going to die?
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Non-Industrial vs Industrial Anthrax

• Nonindustrial anthrax
• usually affects people who work with animals or
  animal carcasses
• farmers, veterinarians, butchers
• almost always cutaneous
• Industrial anthrax
• acquired from handling contaminated hair, hides,
  wool, bone meal, or other animal products
• higher chance of being pulmonary as a result of
  the inhalation of spore-laden dust

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Bacillus Cereus??????

• B. cereus food poisoning results from the
  ingestion of preformed enterotoxins, producing
  predominantly vomiting and diarrhea.
• The vomiting form is most often associated with
  ingestion of a heat stable toxin from
  contaminated rice, while the diarrheal form is
  most often associated with ingestion of a heat
  labile toxin from contaminated meat or vegetables

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B cereus virulence factors

• A 38 to 46-kDa protein complex has been shown in
  animal models
• to cause necrosis of the skin or intestinal
  mucosa
• to induce fluid accumulation in the intestine
• a lethal toxin
• Responsible for the necrotic and toxemic nature
  of severe B cereus infections and for the
  diarrheal form of food poisoning

Bacillus cereus also produces two hemolysins
Phospholipases produced by B cereus may act as
exacerbating factors by degrading host cell
membranes following exposure of their
phospholipid substrates in wounds or other
infections
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Bacillus Food Poisoning Two Distinct Types

• Diarrheal type
• diarrhea and abdominal pain
• 8 to 16 hours after consumption of the
  contaminated food
• Associated with a variety of foods, including
  meat and vegetable dishes, sauces, pastas,
  desserts, and dairy products
• Emetic?? disease
• nausea and vomiting begin 1 to 5 hours after the
  contaminated food is eaten
• Boiled rice that is held for prolonged periods at
  ambient temperature and then quick-fried before
  serving is the usual offender, although dairy
  products or other foods are occasionally
  responsible.