Mycoplasma And Ureaplasma

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Mycoplasma known as human pathogens

Back 1

• Mycoplasma genitalium
• Mycoplasma hominis
• Mycoplasma pneumoniae
  • Ureaplasma urealyticum

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General Characteristics of Mycoplasma and Ureaplasma

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• Do not process cell walls; resistant to cell wall active antibiotics
• Slow growing
• Small (0.3-.08um)
• Fastidious; require cholesterol and fatty acids for growth
  • Difficult to detect; contaminant of cell culture

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General Characteristics of Mycoplasma

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• Known as pleuropneumonia-like organisms (PPLO)
• Colonies grow with center imbedded below agar surface; fried eggs
  • First isolated from bovines as pleuropneumonia; Eaton agent

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Location of infection

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• Epithelium of mucosal surfaces in respiratory and urogenital tract
• Adhere tightly to epithelial cells
• Oropharynx: Common of M. pneumoniae not M. hominis
  • Urogential tract: M. hominis (10-50%), normal flora in women, M. genitalium (15-20% non-gonococcal urethritis)

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Clinical Infections of Mycoplasma pneumoniae

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• Bronchitis
• Pharyngitis
• Walking pneumonia (atypical pneumonia)
• Mostly asymptomatic >90%
  • Isolation always indicated pathogenicity
• 20% pneumonia in general populations
  • 50% military settings

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Spread of Mycoplasma pneumoniae

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• Spread via close contact: schools, prison, military personnel, aerosol droplets
  • Incubation: 2-3 weeks: headache, fever, malaise, anorexia, sore throat, dry cough, earaches

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Mycoplasma hominis

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• Infections of the urogenital tract
• Can be found in normal flora
• Opportunistic pathogen
• Salpingitis: inflammation of the fallopian tubes
• Pyelonephritis: infection of the kidney and ducts
  • PID

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Mycoplasma genitalium

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• 15-20% of PID and non-GC urethritis
• Vertical transmission from mother to fetus
  • Very difficult to take culture takes 2-3 months: interferes with serology for other mycoplasma

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Mycoplasma fermentans

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AIDS-related mycoplasma

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Infection of the Urogenital tract: Ureaplasma urealyticum

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• Infections of the urogenital tract: normal flora in women
• Still cause UTI
• Can infect fetus: chorioamionitis, congenital pneumonia, meningitis of newborns with negative culture
  • 10% of cases of non-GC urethritis in men

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Specimen collection

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• Any body fluid (CSF, tissue, blood, throat, etc.)
• Blood culture bottle should be collected without anticoagulant- Mycoplasma inhibited by SPS
• Extremely sensitive to drying;lack of cell wall
• Swab; keep moist (Dacron)
  • Plate immediately if not freeze -70C

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Culturing Mycoplasma and Ureaplasma

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• M. hominis and U. urealyticum
• Culture: initially in liquid media and watch for pH change
  • Plate enriched culture and check for characteristic colonies

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Culturing Mycoplasma pneumoniae

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• Generally not cultured: takes too long and sensitivity is low
  • Use serology

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Culturing Mycoplasma hominis

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• Arginine broth (enrichment) 7-14 days: turns from a salmon to pink color
• Plate to agar: look for characteristic fried egg colonies of a variety of shapes and sizes
  • Diene's stain: light blue "egg white", Dark blue "yolk"

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Culturing Ureaplasma

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• U-9B broth (enrichment) 3-5 days: yellow to pink (release of NH4)
• Plate to A-7B agar plate: look for irregularly shaped colony, add MgCl2- rust brown color
  • Examine under low power on stereoscope: T-strain Mycoplasma

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Mycoplasma/Ureaplasma: Culture Media

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• Several media have been developed for recovery of Mycoplasma
• Ureaplasma is still most problematic: May require added things, Shepards A-7B agar, SP4broth, A8 broth
• Complex media
• Grows slowly 3 wks
  • Require sterols for growth. mem. synthesis

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SP4 Media

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• Beef heart infusion
• Pepton supplemented with yeast extract
• Glucose arginine, urea can be added; helps to differentiate
• Amphotericin B nad penicllin added to inhibit fast growers
  • Phenol red is added to broth media as a pH indicator

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SP4 media use

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1. Plate: liquid inoculum or swab; incoluate serially up to 10^-3; diluting minimizes inhibitors
2. Apply tape or similar sealant over agar plates to restrict dehydration
3. Incubate plates in 5-10% at 35C for up to 30days
4. Examine with stereomicroscopically
   1. Observe "fried-egg" colonies. M. hominis range from 20-300mm in diameter

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Direct detection methods

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• Currently, non are FDA approved
• Immunobloting
• Indirect immunofluorescence
• Gram stain: rule out other organisms
  • Acridine Orange: non-specific

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M. pneumoniae: Laboratory Diagnosis

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• Culture rarely used
• Serology: Complement fixation: 4 fold rise in titer indicative of a recent infection: need acute and convalescent serum; high false positive rate
  • ELISA

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ELISA

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• Most frequently used
• Immunocard: IgM antibody directed
  • Remel EIA: detects both IgM and IgG

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Cold Agglutinins

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• Nonspecific reaction to outer membrane glycolipids
• Ab to M. pneumonia after 1 wk of infection and peak at 3-6 weeks
• Cold agglutinins are IgM Ab that agglutinate human "o" RBC at 4C but not 37C (in 65%0
  • False pos. Epstein-Barr virus, cytomegalovirus, adenovirus

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Molecular Diagnostic

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• PCR-methods: 16rDNA
• Patient signs symptoms correlate with infectious agent
  • Inconsistent results between: PCR/Culture/serology
    • Multiplex PCR assay for C. penumoniae, L. penumophila, and M. penumoniae has been developed

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Treatment

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• M. hominis: lincomycin
• U. urealyticum: erythromycin
• M. pneumoniae: erythromycin, tetracycline

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Mycoplasma and L-Forms

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• Variant that can replicate in the form of small filterable protoplasmic elements with defective or absent cell walls
• Cell wall synthesis is impaired by antibiotic treatment or high salt concentration
• Contain a rigid cell wall at one stage of their life cycle; contain no sterols in the cytoplasmic membrane
  • Require a sterol and serum proteins for growth