Campylobacter: Introduction, Classification, Morphology, Pathogenesis, Laboratory Diagnosis

Posted on Author Binod G C

Introduction

  • Slender, spirally curved, non spore forming, gram negative rods
  • Cells may form coccoid bodies when exposed to oxygen or upon prolonged culture
  • Motile with a characteristic corkscrew- motion by means of a single polar unseatheted flagellum at one or both ends (except gracilis and C. hominis )
  • Microaerophilic to anaerobic atmospheric requirements
  • Grow at 37oC but not at 15oC
  • Hydrogen may be required by some species for microaerobic growth
  • Oxidase positive (except gracilis )
  • Nonsaccharolytic , energy is obtained from amino acids or TCA intermediates
  • G + C content of DNA- 29-46 mol%

Historical Perspective of Campylobacter

Campylobacter (Greek, curved rods) was named by Sebald and Veron (1963) the group of bacteria formerly known as the microareophilic vibrios

  • Theodar Escherich(1886) –described small vibrios in the large intestinal mucus of infants who died of cholera infantum. His drawings suggests that he was looking at Campylobacter spp but could not isolate
  • First isolated form uterine exudate of aborting sheep in 1906 – McFadyean and Stockman (1913)
  • A few years later Smith isolated similar organism from fetus of aborting cow and named it Vibrio fetus
  • In 1927, another microaerophilic vibrio was isolated from the jenunum of calves with diarrhea and later named Vibrio jejuni
  • In 1944, Doyle isolated similar organism from pigs with dysentery, he named it Vibrio coli
  • These later became Campylobacter jenuni and Campylobacter coli

Classification of Campylobacter

  • rRNA superfamily VI (Vandamme and De Ley 1991)
  • Campylobacteraceae family
  • 18 species
  • Established human pathogens
    • Campylobacter jejuni
    • Campylobacter coli
    • Campylobacter fetus

Resorvoir

  • Widely spread in nature
  • jejuni & C. coli -wide variety of wild and domestic animals especially birds
  • Domestic poultry are major source of human infection
  • Pigs are the main hosts for coli
  • C. consisus, C. sputorum,C.rectus, C. curvus, C. showae and C. gracilis – natural gingival flora of humans

Morphology of Campylobacter

  • Most striking feature – spiral conformation
  • Slender, spirally curved, non spore forming, gram negative rods “Gull-winged” appearance (0.2-0.5um X 0.5x5um)
  • Rapid darting motility in cork screw fashion due to single unsheathed polar flagellum at one or both ends
    • Exceptions- gracilis & C. hominis -no flagella, C. showae – multiple polar flagella)
  • May undergo coccal transformation in old cultures or when exposed to air(C. jejuni & lari)

Growth Requirements

  • Strictly microaerophilic (do not grow in air on primary isolation)
  • 5-10% O2 required for growth
  • Key to successful cultivation of Campylobacter is addition of supplement that consists of ferrous sulfate, sodium metabisulfite and sodium pyruvate (FBP)
  • Most species require CO2 (1-10%) for growth
  • Under ideal conditions, C. fetus and most other produce visible colonies after 24 h at 37oC, but colonies are not well formed until 48 h
  • jejuni, C. coli, C. lari – optimally grow at 42-43oC (thermophilic group)
    • Body temperature of natural avian reservoir

Susceptibility to Physical and Chemical Agents

  • Heat and cold – exposure to temp. of 500C or below 4oC causes sublethal injury
  • Drying – Heavy saline suspension – survive for 2-10 hrs   at 37oC, Skimmed milk suspension – survive for several weeks at 4oC
  • Disinfectants- Hypochlorite, phenols, iodophores & quaternary ammonium compounds effectively kills within 1 min at standard working dilution
  • pH – generally grows at  pH 5.5 – 8. pH <5 and >9 – detrimental to organism
  • Salt – most do not grow in media containing more than 1.5 % NaCl.
    • jejuni can survive for 3-5 days in 4.5 % NaCl at room temp. whereas it can survive for 3 weeks in 6.5% NaCl at 4oC
  • Ascorbic acid – bactericidal to jejuni at 5mmol/l concentration present in wide range of foods

Pathogenecity of Campylobacter

  • Gastroenteritis
    • jejuni ,C. coli – mostly associated with diarrhea in humans
  • Systemic disease
    • fetus – uncommon cause of bacteremia or septecemia in patients with  immune defeciency or serous underlying disease
    • jejuni and C. coli –may also cause septecemia in immunocompromised
  • Reproductive disease
    • fetus subsp. fetus – abortion in cattle
    • fetus subsp. venerealis – infertility in cattle
  • Periodontal disease
    • sputorum, C. concisus, C. rectus, C. curvus, C. gracilis – associated with periodontal disease.

Virulence Factors

  • Cellular components
    • Flagellum: Motility
    • Adhesins: Mediate attachment to mucosa
    • Invasins
    • S-layer protein “microcapsule” in fetus- mediates resistance to serum mediated killing and phagocytosis
  • Extracellular components:
    • Enterotoxins
    • Cytolethal toxins

Pathogenesis of Campylobacter

  • Infection is acquired by oral route from food, drink, or contact with infected animals or animal products
  • Infective dose – 104 organisms (as few as 500)
  • Organisms multiply in the small intestine, invade the epithelium, and produce inflammation that results in the appearance of red and white blood cells in the stools

Post Infection sequalae

  • Non infective reactive arthritis (1% of patients recovering from campylobacter eanteritis)
    • Increased risk among people with HLA – B27 allele
  • Guillan-Barre Syndrome (<1 in 1000)
    • Associated with penner O:19 in japan and USA  and O:41 in south Africa
    • Specific glycopeptide moities in Campylobacter lipopolysachharide  closely resemble normal human ganglioside  GM1, present in the peripheral nerve myelin
    • Enteric infection triggers production of cross reactive antibodies which then attack normal structure
    • resulting in acute paralysis (GBS)

Laboratory Diagnosis of Campylobacter infection

  • Samples
    • Stool (rectal swabs also acceptable for culture)
    • Blood
  • Transport media (if delay anticipated)
    • Cary – Blair transport media
    • Campy thio – thioglycolate broth base with 0.16% agar, vancomycin(10 mg/L), trimethoprim( 5 mg/L), cephalothin(15mg/L), ploymyxin B(2500U/L)and amphotericin B (2mg/L)

Direct Diagnosis

  • Microscopy- direct smear
    • Gram stain- small, curved or seagull –winged, gram negative rods
      • Counterstain with carbol fuchsin or basic fuchsin as it does not stain well with safranin
    • Motility – characteristic darting type of motility
  • Commercial kits – direct detection from feces by latex agglutination test
  • PCR

Stool Culture

  • Selective medium
    • Skirrow medium
    • Campy BAP (vancomycin, polymixin B, trimethoprim
  • Filtration method
    • Filtration method can be used in conjunction with non selective medium to recover Campylobacter spp
    • A filter (0.65 mm pore size) is placed on the agar surface and a drop of stool is placed on the filter
    • The plate is incubated upright for an hour at 37oC, the filter is removed and plate is reincubated in microaerobic atmosphere
  • Incubation at
    • 42oC  in atmosphere of 5% O2, 10% CO2 and 85% N2
  • After 48-72 hrs grey to pinkish or yellowish colonies
  • Some colonies may exhibit a tailing effect on the streaked line
  • Suspicious looking colonies grown on selective medium incubated at 42oC may presumptively identified as Campylobacter spp

Blood Culture

  • Septicemia
  • May require as long as 2 weeks for growth to be detected
  • Turbidity is not often visible in blood culture media, so blind subcultures or microscopic examination using acridine orange stain may be necessary
  • Subculture from broths must be incubated in microaerobic atmosphere or the organisms will not multiply

Prevention

  • No vaccines are available
  • Avoid consumption of properly uncooked meat and meat products, unpasteurized milk, untreated water.
  • Wash hands after handling raw meat to prevent cross contamination from raw poultry meat to other food items

Susceptibility to Antimicrobial Agents

  • Normally suceptible to macrolides, aminoglycosides, chloramphenicol and tetracycline
  • Highly resistant to trimethoprim
  • fetus and several Campylobacter are sensitive to β-lactams but C. jejuni, C. coli and C. lari are resistant to β-lactams including cephalosporins
  • jejuni and C. coli readily acquire resistance to fluroquinolones (limits their use in treatment).

Binod G C

I'm Binod G C (MSc), a PhD candidate in cell and molecular biology who works as a biology educator and enjoys scientific blogging. My proclivity for blogging is intended to make notes and study materials more accessible to students.