MYCOBACTERIA

• Mycobacteria


  Non-motile, non-spore forming slender, slightly curved or straight, rod-shaped organisms that have tendency to clump

  
• Mycobacteria


  • The cell wall has extremely high lipid content; thus, mycobacterial cells resist staining with commonly used basic aniline dyes, such as those used in the Gram stain, at room temperature
  • Take up dye with increased staining time or application of heat but resist decolorization with acid-ethanol
  • This characteristic is referred to as acid fastness—hence, the term AFB
  • Strictly aerobic, but increased carbon dioxide (CO2) will enhance the growth of some species

  
• Mycobacterium tuberculosis Complex


  • M. tuberculosis
  • M. bovis (including the vaccination strain Bacillus Calmette-Guérin)
  • M. africanum
  • M. canettii
  • M. microti

  
• Mycobacterium tuberculosis


  • TB is usually a disease of the respiratory tract
  • Tubercle bacilli are acquired from persons with active disease who are excreting viable bacilli by coughing, sneezing or talking
  • Hard tubercle or granuloma may be formed
  • The granuloma is an organization of lymphocytes, macrophages, fibroblasts, and capillaries
  • With granuloma formation, healing occurs, as well as fibrosis, encapsulation, and calcification, with scar formation as a reminder of the past infection
  • In infected individuals, there is a potential for reactivation of TB

  
• Primary TB


  • Exudative lesion
  • Granulomatous lesion
• Secondary TB


  Reactivation or reinfection
• Disseminated or Extrapulmonary TB


  Seeding of many organs outside the pulmonary tree with AFB through hematogenous spread
• Almost any organ of the body can be infected by M. tuberculosis
• Pleurisy, an unexplained pleural effusion with mononuclear pleurocytosis, manifests as cough, fever, and chest pain, resembling the presentation of bacterial pneumonia
• Lymphadenitis, Genitourinary TB, Skeletal TB of the spine (Pott disease), Meningitis
• Cerebrospinal fluid (CSF) examination usually reveals an elevated protein level, decreased glucose level, and a predominance of lymphocytes
• Mycobacterium tuberculosis Identification


  • Colonies are typically raised, with a dry, rough appearance
  • The colonies are nonpigmented and classically described as being buff-colored
  • Elaboration of cord factor can result in characteristic cord formation
  • Optimal growth occurs at 35° C to 37° C
  • Positive for niacin accumulation
  • Reduction of nitrate to nitrite
  • Production of catalase
  • Grows on thiophene-2-carboxylic acid hydrazide (T2H)

  
• Mycobacterium bovis


  • TB primarily in cattle but also in other ruminants, as well as in dogs, cats, swine, parrots, and humans
  • The disease in humans closely resembles that caused by M. tuberculosis and is treated similarly
  • 21 days of incubation at 37° C
  • Niacin-negative
  • Do not reduce nitrate
  • Do not grow in the presence of (T2H) thiophene-2-carboxylic acid hydrazide

  
• Tuberculosis Treatment (Quadruple Therapy)

  • Isoniazid (isonicotinic acid hydrazide, INH)
  • Rifampicin
  • Pyrazinamide
  • Ethambutol
  • Streptomycin
• DOTS (Direct Observed Treatment Shortcourse)


  Patient's compliance to drug regimen
• Tuberculosis Treatment


  • 9-month course of therapy
  • Isoniazid and rifampin
  • Other drugs used if resistant (Kanamycin, ciprofloxacin)

  
• Treatment failures


  • Patient noncompliance
  • Drug-resistant mycobacteria
• MDR-TB


  Resistant to at least two of the most powerful first line anti-TB drugs, INH and RMP
• XDR-TB


  Resistant to INH and RMP plus any fluoroquinolones and at least injectable second-line drugs (eg. amikacin, kanamycin, or capreomycin)
• Non-tuberculous Mycobacteria


  • Atypical mycobacteria or mycobacteria other than tuberculosis (MOTT)
  • Found in the environment that colonize the skin, respiratory tract and GI of healthy individuals
  • Chronic pulmonary disease resembling TB
  • Non transmissible from person to person
  • AIDS contributed to incidence to NTM disease

  
• Slowly Growing Non-tuberculous Mycobacteria

  • Mycobacterium avium complex
  • Mycobacterium avium subsp. paratuberculosis
  • Mycobacterium genavense
  • Mycobacterium haemophilum
  • Mycobacterium kansasii
  • Mycobacterium malmoense
  • Mycobacterium scrofulaceum
  • Mycobacterium simiae
  • Mycobacterium ulcerans
  • Mycobacterium marinum
  • Mycobacterium xenopi

  
• Mycobacterium avium complex


  • Mycobacterium avium and M. intracellulare are part of the Mycobacterium avium complex (MAC)
  • Environmental saprophytes and have been recovered from soil, water, house dust, and other environmental sources
  • M. avium is a cause of disease in poultry and swine - Zoonotic
  • The cells are short, coccobacillary, and uniformly stained, without beading or banding
  • Production of a heat-stable catalase and the ability to grow on media containing 2 μg/mL of T2H
• Mycobacterium avium subsp. paratuberculosis

  • Causative agent of Johne disease, an intestinal infection occurring as a chronic diarrhea in cattle, sheep, goats, and other ruminants
  • Very slow growth rate (3 to 4 months)
  • Needs mycobactin-supplemented medium for primary isolation

  
• Mycobacterium genavense


  • Cause of disseminated infections in patients with AIDS
  • Enteritis and genital and soft tissue infections
  • Middlebrook 7H11 agar supplemented with mycobactin
  • Heat-stable catalase, pyrazinamidase, and urease - positive

  
• Mycobacterium haemophilum


  • Submandibular lymphadenitis, subcutaneous nodules, painful swellings, ulcers progressing to abscesses, and draining fistulas are often the clinical manifestations
  • Unique characteristic of this organism is its requirement for hemoglobin or hemin for growth
  • Optimal growth temperature is 28° C to 32° C; little or no growth occurs at 37° C
  • Cells are strongly acid-fast, short, occasionally curved bacilli without banding or beading, and arranged in tight clusters or cords

  
• Mycobacterium kansasii


  • Infections are not normally considered contagious from person to person
  • Susceptible to rifampin and ethambutol, partially resistant to isoniazid and streptomycin, and resistant to pyrazinamide
  • A multidrug regimen of isoniazid, rifampin, and ethambutol is currently recommended
  • Long rods with distinct crossbanding
  • Colonies appear smooth to rough, with characteristic wavy edges and dark centers when grown on Middlebrook 7H10 agar
  • Colonies are photochromogenic
  • With prolonged exposure to light, most strains form dark red crystals of β--carotene on the surface of and inside the colony
  • Strongly catalase-positive
  • Hydrolyze Tween 80 in 3 days
  • Strong nitrate reduction
  • Pyrazinamidase production

  
• Mycobacterium malmoense


  • Chronic pulmonary disease and cervical lymphadenitis
  • Resistant to isoniazid, streptomycin, p-aminosalicylic acid, and rifampin and susceptible to ethambutol and cycloserine
  • Short coccobacillus without cross bands on acid-fast–stained smears
  • Colonies are smooth glistening, and opaque, with dense centers

  
• Mycobacterium scrofulaceum


  • Cervical lymphadenitis in children
  • Resistant to isoniazid, streptomycin, ethambutol, and p-aminosalicylic acid when tested in vitro
  • The organism grows slowly (4 to 6 weeks) at incubation temperatures ranging from 25° to 37° C

  
• Mycobacterium simiae


  • Isolated from the lymph nodes of monkeys
  • Colonies on Middlebrook 7H10 agar are thin, transparent or tiny, and filamentous

  
• Mycobacterium ulcerans


  • Mycobacterium ulcerans is a rare cause of mycobacteriosis, also referred to as Buruli ulcer
  • Acid-fast cells are long, without beading or crossbanding

  
• Mycobacterium marinum


  • Mycobacterium marinum has been implicated in diseases of fish and isolated from aquariums
  • Cutaneous infections in humans occur when traumatized skin comes into contact with salt water or inadequately chlorinated fresh water containing the organism
  • Tender red or blue-red subcutaneous nodule, or swimming pool granuloma, usually occurs on the elbow, knee, toe, or finger
  • Susceptible to rifampin and ethambutol, resistant to isoniazid and pyrazinamide, and partially resistant or intermediate to streptomycin
  • Cells of M. marinum are moderately long to long rods with cross barring
  • M. marinum is photochromogenic; young colonies
  • Colonies grown in or exposed to light develop a deep yellow color
  • Growth is optimum at incubation temperatures of 28° C to 32° C
  • None reduces nitrate or produces heat-stable catalase
  • The organisms hydrolyze Tween 80 and produce urease and pyrazinamidase

  
• Mycobacterium xenopi


  • Recovered from hot and cold-water taps (including water storage tanks of hospitals)
  • Susceptible to the quinolones (ciprofloxacin, ofloxacin); some isolates are susceptible to vancomycin, erythromycin, or cefuroxime
  • Colonies on Middlebrook 7H10 agar are small, with dense centers and filamentous edges
  • Cornmeal-glycerol agar reveals distinctive round colonies with branching and filamentous extensions; aerial hyphae are usually seen in rough colonies
  • Young colonies grown on cornmeal agar have a bird's nest appearance, with characteristic stick-like projections

  
• Rapidly Growing Non-tuberculous Mycobacteria

  • Mycobacterium chelonae–Mycobacterium abscessus Group
  • Mycobacterium fortuitum Group
  • Mycobacterium smegmatis Group

  
• Mycobacterium chelonae–Mycobacterium abscessus Group

  • M. abscessus subsp. abscessus (formerly M. abscessus), M. chelonae, and M. fortuitum
  • M. chelonae have been associated with a variety of infections of the skin, lungs, bone, central nervous system, and prosthetic heart valves
  • M. abscessus seen in patients with cystic fibrosis (CF)
  • Tap water is an important reservoir
  • Positive 3-day arylsulfatase test, no reduction of nitrate, and growth on MacConkey agar without crystal violet

  
• Mycobacterium fortuitum Group


  • M. fortuitum, M. peregrinum, and an unnamed third species
  • Isolated from water, soil, and dust
  • Associated with localized cutaneous infections
  • Middlebrook 7H11 agars after 1 to 2 days of incubation colonies with branching filamentous extensions and rough colonies with short aerial hyphae
  • Pleomorphic, ranging from long and tapered to short, thick rods partially acid-fast
  • Positive 3-day arylsulfatase test and reduction of nitrate

  
• Mycobacterium smegmatis Group


  • The M. smegmatis group contains two species, M. smegmatis and M. goodie
  • M. smegmatis has been implicated in rare cases of pulmonary, skin, soft tissue, and bone infections
  • Cells are long and tapered or short rods with irregular acid fastness. Occasionally rods are curved with branching or Y-shaped forms; swollen, with deeper staining, beaded, or ovoid forms are sometimes seen
  • Colonies appearing on egg medium after 2 to 4 days are usually rough, wrinkled, or coarsely folded; smooth, glistening, butyrous colonies may also be seen
  • Negative arylsulfatase reaction, positive iron uptake, ability to reduce nitrate, and growth in the presence of 5% NaCl and on MacConkey agar without crystal violet

  
• Mycobacterium leprae


  • Causative agent of Hansen disease (leprosy), an infection of the skin, mucous membranes, and peripheral nerves
  • The two major forms of the disease are tuberculoid leprosy and lepromatous leprosy
  • Symptoms of tuberculoid leprosy include skin lesions and nerve involvement that can produce areas with loss of sensation
  • Patients with lepromatous leprosy if untreated, life-threatening
  • Characterized by skin lesions and progressive, symmetric nerve damage
  • Lesions of the mucous membranes of the nose may lead to destruction of the cartilaginous septum, resulting in nasal and facial deformities
  • Combination of diaminodiphenylsulfone (dapsone), clofazimine, and rifampin
  • M. leprae has not been grown on artificial media
  • Mouse footpad
  • The length of the bacillus is at least five times the width of the bacillus
• The most commonly recommended container for specimen collection is a sterile, wide-mouth cup with a tightly fitted lid
• An early-morning sputum specimen should be collected on three consecutive days, although recent studies have suggested that the addition of a third specimen does not significantly increase the sensitivity of detecting mycobacteria

  
• If at least two of the first three sputum direct smears are positive, then three specimens are sufficient