Clinical Bacteriology - Laboratory PDF

CLINICAL BACTERIOLOGY - LABORATORY MIDTERMS TOPIC 5. Mycobacteria Lecturer/s: Mr. Jason “Ab” Chua, RMT, MSMT FULL TRANSES MASTERLIST: https://bit.ly/masterli_st INTRODUCTION The...

CLINICAL BACTERIOLOGY - LABORATORY MIDTERMS TOPIC 5. Mycobacteria Lecturer/s: Mr. Jason “Ab” Chua, RMT, MSMT FULL TRANSES MASTERLIST: https://bit.ly/masterli_st INTRODUCTION The genera are closely related to Mycobacterium, it includes Every year, 10,000,000 people fall ill with Nocardia spp., Rhodococcus spp., Tsukamurella spp., and Tuberculosis or the so-called ‘TB,’ despite being a Gordonia spp. preventable and curable disease CELL WALL OF Mycobacteria 1,500,000 people die from TB each year, making it Cell wall contains N-glycolylneuramic acid instead of the world’s top infectious killer N-acetylmuramic acid TB is the leading cause of death for people with HIV, The cell wall has extremely high lipid (Mycolic Acid) and also a major contributor for antimicrobial content resistance ○ resist decolorization with acid-alcohol Most of the people who fall ill with TB live in low and ○ Creates a hydrophobic permeability barrier middle income countries—but TB is present all over ➔ Because of the cell wall structure, Mycobacteria are difficult the world, of all people TB be found in countries such to stain with commonly-used basic aniline dyes such as those as Bangladesh, China, India, Indonesia, Nigeria, used with Gram-staining Pakistan, Philippines, and South Africa Although these organisms cannot be readily Gram-stained, About one-quarters of the population is estimated to they are generally considered GRAM-POSITIVE be infected by TB bacteria Only 5-15% of these people will fall ill with active TB ACID FASTNESS disease; the rest have TB infection, but are not ill, and - however, they resist decolorization with Acidified cannot transmit the disease alcohol or 3% Hydrochloric Acid after prolonged Both TB infection and disease are curable with application of the Basic Fuchsin Dye or the heating antibiotics of this dye after its application. GENERAL CHARACTERISTICS - This important property of Mycobacteria, which derives from their cell wall structure is referred to as ACID-FASTNESS - This characteristic distinguishes Mycobacteria from other genera Acid fast bacilli (AFB): basic distinguishing feature The organisms that belong to the genus Mycobacterium are AEROBIC, although some may grow in reduced oxygen concentrations They are non-spore-forming except for Mycobacterium marinum Most pathogenic species requires 2-6 weeks of Non-motile incubation Very thin Most are slow growers except Mycobacterium Slightly curved or straight rods fortuitum-chelonei ○ Usually 0.2-0.6 or 1-10 mm in size ○ 3 DAYS on Modified MacConkey Agar ○ Some species may display a branching Mycobacterium leprae – FAILS to grow in vitro (using morphology agarose medium) Mycobacterium is the only genus in the Mycobacteriaceae Mycobacterium tuberculosis family other name: Koch’s bacillus/ Tubercle bacilli order Actinomycetales First described by ROBERT KOCH in 1882 class Actinomycetes ➔ The bacteria usually attacks the lungs; but TB bacteria can attack any part of the body, such as the kidneys, spine, and brain Made by Miguel Astronomo & Daniel Budo 1  Not everyone infected with TB bacteria becomes TUBERCULOSIS sick; as a result 2 TB conditions exist 1. Latent TB infection (LTBI) 2. TB Disease – can be fatal Agent of Tuberculosis A chronic infection (host carries the organism chronically) 20th century - 2nd leading cause of death VIRULENCE FACTORS of Mycobacterium tuberculosis A disease of the respiratory tract (Pulmonary TB [PTB]) Acquired by person with active disease by means of: 1. Coughing 2. Sneezing or talking TB bacteria can spread to the air from person-to-person by 1. Inhalation of droplet nuclei containing the organism a. Infectious aerosols, usually 1-5 micrometers are produced when people with Pulmonary Tuberculosis cough or sneeze, 1. Mycoside speak a. Mycolic acid bound to a carbohydrate b. May also be produced by 2. Cord Factor manipulation of lesions, a. is a mycoside formed by the union of two (2) processing of clinical specimens in mycolic acid with a disaccharide (trehalose). the laboratory b. Inhibits neutrophil migration and damages Droplets are so small that air currents keep them mitochondria. airborne for long periods once inhaled c. Release Tumor Necrosis Factor resulting in They are small enough to reach the lungs → alveoli, rapid weight loss. so when a person breathes a TB Bacteria, the i. If there are persons infected with bacteria can settle in the lungs and begin to grow. these bacteria causing TB infection, ○ They can move through the blood-through they appear thin the other parts of the body (spine, kidney, 3. Sulfatides etc.) a. are mycosides that resemble cord factor with sulfates attached to the disaccharide. TUBERCULOSIS IN THE LUNGS AND THROAT b. Inhibit the phagosome from fusing with the can be INFECTIOUS lysosome that contain bacteriocidal the bacteria can spread to other people enzymes. 4. Wax D TUBERCULOSIS IN THE KIDNEYS AND OTHER PARTS a. is a complicated mycoside that acts as an usually NOT INFECTIOUS adjuvant (enhances antibody formation to an antigen) and activates the protective DIAGNOSIS OF PULMONARY TUBERCULOSIS cellular immune system. CHEST RADIOGRAPH is useful for diagnosis of TB disease Chest abnormalities suggest pulmonary TB disease Made by Miguel Astronomo & Daniel Budo 2  Posterior and anterior radiograph of the chest is the BCG (bacille Calmette-Guerin) replicates Primary TB standard view used for the detection of TB-related sufficiently to protect from disseminated TB and chest abnormalities meningitis but not from post-primary TB In some cases, especially for children, a lateral view Maintenance and protection depends from may be helpful immunocompetent persons In pulmonary TB disease, radiographic abnormalities Lesions of primary TB can recur whenever are often seen in the apical and posterior segments immunosupperssion reduces systemic T of the upper lobe, or in the SUPERIOR SEGMENTS of cell-mediated immunity the LOWER LOBE Lesions may appear anywhere in the lungs and may PATHOGENESIS OF PRIMARY TUBERCULOSIS differ in size, shape, density, and cavitation, especially 1. M. tuberculosis cells are phagocytized by alveolar in HIV-infected, and other immunosuppressed macrophages and are capable of intracellular results multiplication. RADIOGRAPHIC ABNORMALITIES in children tend 2. In a person with adequate cellular immunity, T cells to be minimal with a greater likelihood of arrive within 4 to 6 weeks owing to LYMPHADENOPATHY more easily diagnosed on the macrophage-activating polypeptides termed lateral film– lymphokines. 3. This enables the macrophage in the area of Mixed nodular and periodic lesions may contain infection to destroy the intracellular mycobacteria. slowly-multiplying tubercle bacilli and have the 4. There is then a regression and healing of the primary potential for progression to TB Disease laesion and any disseminated foci PRIMARY TUBERCULOSIS In many exposed individuals, the immune system does not eliminate the bacteria. If there is little antigen and a strong hypersensitivity reaction, a hard tubercle or granuloma may be formed an organization of lymphocytes, macrophages, fibroblasts, and capillaries. ○ Formed in the lungs ○ Includes giant cell formation or cellular fusion displaying multiple nuclei, if the Mycobacterium antigen concentration is high, the hypersensitivity reaction may result in TISSUE NECROSIS caused by enzymes released by the macrophages Infection that occurs in a person with insufficient ○ In this case, no granuloma forms– solid / immunity to localize and control MTP semisolid caseous material is left at the (Mycobacterium tuberculosis culi pili) and granulomas primary lesion site, ○ it can produce a spectrum of clinical disease ○ In some patients infected with primary states, ranging from disseminated TB in reactive tuberculosis, the disease will persons with AIDS, Meningitis, Miliary spread via the lymph system or tuberculosis, and extrapulmonary hematogenously leading to meningal, granulomas miliary or disseminated tuberculosis ○ Typically occurs in very young, ○ This most often occurs with patients in a immunologically naïve, very old, or repressed or ineffective cellular immunity immunologically-suppressed persons With granuloma formation, healing occurs, as well ○ The infection can disseminate via as fibrosis, encapsulation, and calcification, with scar lymphatics or the blood stream to lymph formation as a reminder of the past infection. nodes and diverse other organs ➔ Immunocompetent people – the infection is controlled in weeks and the lesions heal Primary TB produces SYSTEMIC IMMUNITY that effectively protects the entire body from the disseminated infection ○ This immunity has been extensively studied and has become the CENTRAL DOGMA OF PROTECTIVE IMMUNITY mediated by macrophages Granulomas in the production of IF-N gamma by CD4+ T cells Made by Miguel Astronomo & Daniel Budo 3  POTT’S DISEASE resistant-TB) Latent TB infection can be treated; will grow into disease if not treated TB diseases can be treated by a standard, 6-month course for antibiotics, sometimes 5 ○ Common drugs include RIPES (First-line) ○ CCCOKER → second line agents For MDR-TB, XDR-TB If the treatment is not properly completed, then the disease can become drug-resistant and spread In the case of TB infection, where a person is infected with TB bacteria but not ill, TB preventive treatment can be given to stop the onset of disease PREVENTION OF MTB INFECTION "Tuberculosis Spondylitis" or Skeletal TB of the Spine invasion into the spinal vertebrae (graveform) Bacillus Calmette-Guérin (BCG) vaccine is Most dangerous form of mucoskeletal tuberculosis prepared from a strain of the attenuated because it can cause bone destruction, deformity, (virulence-reduced) live bovine tuberculosis and paraplegia. bacillus, Mycobacterium bovis, that has lost its Most commonly involves the thoracic and ability to cause disease in humans. lumbosacral spine Pott’s Disease and Miliary Tuberculosis are diseases Not widely used in the United when M. tuberculosis spreads to other parts of the States body Given to infants and children in MILIARY TUBERCULOSIS other countries where TB is common seeding of many organs outside the pulmonary tree BCG does not always protect people from TB (Extrapulmonary Tuberculosis) THIS VACCINATION SHOULD ONLY BE common sites: spleen, liver, lungs, BM, kidney, CONSIDERED TO CHILDREN WHO TESTED adrenal gland & eyes NEGATIVE TB TEST AND ARE CONTINUALLY Potentially fatal form of the disseminated disease EXPOSED AND CANNOT BE SEPARATED FROM due to the hematogenous spread of tubercle bacilli ADULTS WHO ARE UNTREATED FOR TB DISEASE into the lungs and other organs ○ The child cannot be given long-term ○ Formation of Millet-seed sized, usually primary preventive treatment for TB 1-2mm in size of tuberculus cocci Infection TREATMENT ○ or have Isoniazid and Rifampicin resistant strains of the TB disease First-line TB Rifampicin Drugs LABORATORY DIAGNOSIS - 2 TESTS Isoniazid Tuberculin Test Pyrazinamide TB blood test Ethambutol #1 - TUBERCULIN TEST/ TB SKIN TEST/ MANTOUX Steptomycin TUBERCULIN SKIN TEST Second-line Capreomycin Agents Ciprofloxacin Cycloserine Ofloxacin Kanamycin Ethionamide Skin test which identifies tuberculosis infection, Rifabutin recent or past, with or without disease. Persons infected with tubercle bacilli develop TREATMENT FOR DRUG-RESISTANT TB (More complex) hypersensitivity to the proteins ○ Injecting a small amount of fluid called MDR-TB Resistant to RIFAMPICIN and tuberculin onto the skin and the lower part (Multidrug ISONIAZID of the arm resistant- TB) ○ Must return within 48-72 hours to check XDR-TB MDR-TB + resistant to 2nd-line TB for reactions (Extensively drugs and Fluoroquinolone ○ Result depends on the size of the raised or drug the hard area/ swelling of the arm Preferred tests for children under the age of 5 Made by Miguel Astronomo & Daniel Budo 4  positive results tells a person has been infected with TB bacteria Sputum and bronchial aspirates are the most ○ Does NOT tell whether the person has common | 2 sputum samples (early-morning latent TB infection or has progressed to TB specimens that are collected 1 hr apart or atleast disease within three consecutive days) ○ Other tests such as test x ray or sample of ○ If sputum is not available, check Box 26-2 sputum are needed to see whether the DECONTAMINATION AND DIGESTION person has TB disease METHOD OF ADMINISTRATION Mantoux test: intracutaneous injection 1. Sodium Hydroxide (NaOH): The standard dose 0.1 ml of Purified Protein a. usual concentration 2-4%; serves as a Derivative (PPD) or 5TU (tuberculin units) digestant (mucolytic) and decontaminating (+) erythema & hardening of the site of injection agent (10mm) 2. N-Acetyl-L-Cysteine (dithiothreitol) as mucolytic agent with 4% NaOH (decontaminating agent): ➔ Positive skin test means that the person’s body was recommended technique to decontaminate and infected with TB bacteria and additional tests are needed to digest sputum determine if the person has latent TB infection/disease 3. Zephiran (benzalkonium chloride) + trisodium ➔ negative skin test means the person’s body did not react to phosphate (Z-TSP) the test a. TSP liquefies sputum rapidly but requires a long exposure time to decontaminate You can also perform b. Zephiran shortens the exposure time ○ Von pirquet test ZIEHL-NEELSEN AND KINYOUN STAINS ○ Vollmer patch test ○ Moro percutaneous tests ○ Tuberculin time SPECIMENS BOTH uses: carbolfuchsin as the primary stain acid-alcohol as a decolorizing agent methylene blue counterstain Ziehl-Neelsen: hot stain | heat as mordant ○ ➔ For better penetration of the stain into the Mycobacterial cell wall, hence it is known as the Hot Stain procedure ○ Appear red; Mycobacteria appear blue Kinyoun: cold stain | turgitol as mordant ○ Acid-fast bacilli appear purple to red;slightly curved, short/long rods 2-8 mm in size ○ May appear beaded or banded (Mycobacterium kansasii) ➔ For some non-tubercule species, such as Mycobacterium avium complex; they appear pleomorphic and they are usually cocoid Made by Miguel Astronomo & Daniel Budo 5  DIRECT SPUTUM SMEAR MICROSCOPY EGG BASE Smear size: 2x3 cm With egg yolk (lipid source) and malachite green to inhibit the Read at 300 fields under OIO growth of contaminants ○ Count those different AFB on your AFB smear Gram stain qualifies specimen if acceptable Bartlett’s criteria – sputum specimens should have… ○ 25 PMNs NATIONAL STANDARD REPORTING SCALE REPORT # OF ORGANISM SEEN 0 No AFB seen in 300 fields +n 1-9 AFB/ 100 fields PETRAGNANI 1+ 10-99 AFB/ 100 fields ○ Contains twice the concentration of malachite than the LowensteinJensen 2+ 1-10 AFB/ in at least 50 medium – inhibitor of contaminating fields organisms 3+ More than 10 AFB/ in at ○ Improves recovery from heavily least 20 fields contaminated specimens Lowenstein-Jensen: Most commonly used American Thoracic Society (ATS) medium CULTURE MEDIA AGAR-BASED (SERUM-ALBUMIN AGAR MEDIA) CULTURE Middlebrook 7H11: Colonies of this slowly growing species are typically raised, ○ Composed of: 0.1% casein hydrolysate with a dry, rough appearance. The colonies are nonpigmented ○ Purpose: improves recovery of and classically described as being buff-colored. isoniazid-resistant MTB On Lowenstein-Jenson (LJ) slants, M. tb typically has a Mitchison’s selective 7H11 crumbly, "bread-crumb" appearance whereas the M. avium complex has a bright yellow, LIQUID MEDIA smooth, characteristic colony (so-called "condom Mycobacterium spp. grow more rapidly in liquid medium, and it colony"). can be used for both primary isolation and subculturing These morphologic features are useful in making a Bactec 128 medium presumptive speciation, which is usually confirmed ○ Used in MGIT 960 system by DNA probe. ○ The Polymyxin B, amphotericin B, nalidixic acid, trimethoprim and azlocillin (PANTA) is BIOCHEMICAL TESTS added before incubation and the C-labeled 1. NIACIN ACCUMULATION palmitic acid is metabolized to produce CO2 which is detected by the machine Middlebrook 7H9 broth ○ for aseptically collected specimens from normally sterile sites Middlebrook 7H13 ○ for larger volumes of bone marrow or blood specimens Made by Miguel Astronomo & Daniel Budo 6 Once categorized into a preliminary subgroup based on its 3. CATALASE TEST growth characteristics, an organism must be identified to species or complex level. One of which is NIACIN or NICOTINIC ACID Plays an important role in the oxidation-reduction reactions that occur during Mycobacterium metabolism. Although all species produce nicotinic acid, M. tuberculosis accumulates the largest amount M. simiae and some strains of M. chelonae also produce niacin Most species of Mycobacteria, except for some strains of Most mycobacteria possess the enzyme that converts free Mycobacterium tuberculosis complex, some isoniazid resistant niacin to niacin ribonucleotide. strains and Mycobacterium gastri produce the intracellular However, 95% of M. tuberculosis isolates produce free enzyme catalase which splits hydrogen peroxide into water niacin (nicotinic acid) because this species lacks the and oxygen. niacin connecting enzyme. Mycobacteria are catalase positive. However, not all strains produce a positive reaction after the culture PRINCIPLE: is heated to 68°C for 20 minutes (heat-stable NIACIN + Cyanogen bromide (reagent) + AMINE = catalase). Yellow-pigmented compound Most M. tuberculosis complex organisms do not produce (+) = YELLOW PIGMENTATION heat-stable catalase. (-) = COLORLESS REACTION It can be assessed using the semi-quantitative catalase test or 2. NITRATE REDUCTION the heat stable catalase test. Semi-quantitation of catalase production [shown above] – based on the relative activity of the enzyme. ○ As is determined by the height of a column of bubbles of oxygen formed by the action of untreated enzymes produced by the organism. Based on the semiquantitative catalase test, Mycobacteria are divided into two group: ○ produce less than 45 mm of bubbles ○ produce more than 45 mm of bubbles. The control tube does not have any effervescence or bubble formation. 1+ reaction indicates light pink, 3+ is pink, and 5+ reaction is dark ○ Tween 80 (a detergent) + 30% H2O2 + pink. The control tube remains clear, which has no color change at 2-week old culture grown in an agar deep all. The reaction is read after 5 It is valuable for identifying: minutes Mycobacterium tuberculosis, The column size is recorded as Mycobacterium kansasii, greater than or less than 45 mm Mycobacterium szulgai, Mycobacterium fortuitum 4. TWEEN 80 HYDROLYSIS The ability of acid-fast bacilli to reduce nitrate is influenced by the (1) age of colonies, (2) temperature, (3) pH, and (4) enzyme inhibitors. Rapid growers can be tested within two weeks and slow growers should be tested after three to four weeks of luxuriant growth. The production of nitroreductase, which catalyzes the reduction of nitrate to nitrite, is relatively uncommon among Mycobacterium spp., but a POSITIVE RESULT may be seen in M. kansasii, M. szulgai, M. fortuitum, and M. tuberculosis. The commonly non-pathogenic slow growing scotochromogens and non-photochromogens produce a LIPASE that can hydrolyze Tween 80 Made by Miguel Astronomo & Daniel Budo 7  It is a detergent of polyoxyethylene sorbitan MYCOBACTERIUM SPP. monooleate into oleic acid and polyoxyethylated Mycobacterium bovis sorbitol. Pathogenic species do not produce that product. Tween 80 hydrolysis is useful for differentiating species of photochromogens, non-chromogens, and scotochromogens. Some Mycobacteria possess a lipase that can split the detergent Tween 80 into oleic acid and polyoxyethylated sorbitol. The pH indicator, neutral red, is initially bound to Tween 80 and has an amber color. After hydrolysis of Tween 80, neutral red can no longer bind, and it is released, causing a pink color to form. Another organism that can cause TB disease in people that can 5. ARYLSULFATASE TEST affect lungs, lymph nodes, etc. The enzyme arylsulfatase is present in most Mycobacteria. However, as with M. tuberculosis, NOT EVERYONE Test conditions can be varied to distinguish different infected M. bovis becomes sick. forms of the enzyme. Then, the rate at which the enzyme breaks down Most commonly found in CATTLE and other animals, such as Phenolphthalein Disulfate into Phenolphthalein bison, elk, and deer forms a red color in the presence of SODIUM People are most commonly infected with M. bovis by BICARBONATE eating or drinking contaminated unpasteurized dairy Other salts help to differentiate certain strains of products Mycobacteria. ○ The pasteurization process destroys disease The three-day test is useful for identifying the causing organisms in milk. Rapid heating and potentially pathogenic rapid growers such as M. cooling the milk eliminates M. bovis fortuitum or M. chelonei or M. fortuitumchelonei Infection can also occur from direct contact with a complex. wound, such as what might occur during slaughter or Slow growing M. marinum and M. szulgai are positive hunting, or by inhaling the bacteria in the air exhaled in Arylsulfatase test using the 14- day test. by animals infected with M. bovis Negative organisms are M. evium complex and M. ○ Direct transmission through the air – rare tuberculosis. ○ Person-to-person transmission – when Principle: detects Rapid Growers Tripotassium people with a lung disease cough or sneeze Phenolphthalein Disulfide/sulfate → Free Phenolphthalein (END PRODUCT) Mycobacterium bovis produces TB primarily in cattle but also in other ruminants, as well as in dogs, cats, swine, parrots, and Positive result = Red/ Pink humans Strongly (+): M. fortuitum-chelonei Negative: MAC, M. tuberculosis Most strains of M. bovis are NIACIN-NEGATIVE Mycobacterium leprae BIOCHEM + - TESTS Niacin M. tuberculosis M. bovis (no color Accumulation (yellow) change) Nitrate M. tuberculosis, M. Reduction kansasii, M. szulgai, No Color Change M. fortuitum (red) Catalase Test Most Mycobacteria M. tuberculosis (vigorous bubbling, (no bubbles, 45 mm height of mm height of bubbles) bubbles) Non tuberculosis M. leprae Tween 80 Non-photochromog Hydrolysis ens and M. tuberculosis Close relative of M. tuberculosis Scotochromogens (no color change) (pink) Leprosy – chronic disease of the skin, mucous membranes and the nerve tissue. Arylsulfatase M. Remains a world-wide public health concern as a MAC, M. Test fortuitum-chelonae result of the development of drug-resistant isolates. tuberculosis (red/pink) Made by Miguel Astronomo & Daniel Budo 8  Is the causative agent of Hansen disease (leprosy), an Mycobacterium leprae has not yet been cultivated in infection of the skin, mucous membranes, and vitro. It can be cultivated in the Armadillo and peripheral nerves. footpads of mice. ○ Hansen disease is not considered highly Molecular biologic techniques have provided contagious information about this organism’s genomic structure, its various genes, and its products. TWO MAJOR FORMS Polymerase Chain Reaction (PCR) assays have been used to Tuberculoid leprosy Symptoms of tuberculoid leprosy detect and identify M. leprae in infective tissues. include skin lesions and nerve involvement (loss of sensation). Non-culturable on artificial medium Patients eventually exhibit an effective cell-mediated immune (CMI) response. Lepromatous Characterized by skin lesions and leprosy progressive, symmetric nerve damage and DO NOT PRODUCE an effective CMI response. TUBERCULOID LEPROSY VS. LEPROMATOUS LEPROSY Tuberculoid Leprosy Lepromatous Leprosy Can be grown experimentally only in: Footpad of mice: success is in the low temperature Tuberculoid leprosy has a Lepromatous leprosy is (30°C) of footpads restricted pathogenic growth extensively spread all over the Armadillos: animals with low body temperature body Runyon’s Classification Has a relatively high cell Has a poor cell mediated Only those nontuberculous Mycobacterium are classified under mediated immunity immunity Runyon’s classification. A less severe case of The most severe stage leprosy Doesn’t involve a higher Involves a relatively higher amount of bacterial food bacterial load LABORATORY DIAGNOSIS Lepromin Test Photochromogens (Group I) – pigmented only when exposed to light ○ The NTM colonies that develop pigment on exposure to light after being grown in the dark and it will take longer than 7 days to appear in a solid media. ○ M. kansasii: “yellow bacillus” ○ M. marinum: means “of the sea”, agent of swimming pool granuloma Skin test consisting of a heat killed suspension of M. leprae prepared from lepromatous nodules. This test however lacks specificity. TYPES OF REACTION Early Fernandez reaction: induration appears in 24- 48 hours Late Mitsuda reaction: indurated nodule develops after 3-4 weeks Scotochromogens (Group II) – pigmented both in CULTIVATION darkness and light and it would take 7 days to appear in a solid media Made by Miguel Astronomo & Daniel Budo 9  ○ M. xenopi: branching colonies with aerial hyphae on cornmeal agar described as “bird’s nest” ○ M. gordonae: “tap water” bacillus Nonphotochromogenic (Group III) – nonpigmented both in light and darkness and it will take 7 days to appear in a solid media ○ M. tuberculosis belongs to this group. ○ M. avium – M. intracellulare (Battey bacillus): complex opportunistic pathogens in patients with AIDS and other immune dysfunctions ○ M. terrae: “radish bacillus” Rapid growers (Group IV) – produce visible growth within less than 7 days after subculture to Lowenstein-Jensen medium—may partially or completely lose this characteristic as a result of their growth characteristics. ○ By definition, it requires more than seven days to produce colonies on solid media, and then the variation in generation times among the Mycobacteria results in the formation of visible colonies in 2-60 days at an optimum temperature. M. fortuitum-chelonae complex: grows in Mac w/o CV (Mod. Mac) PHOTOREACTIVITY OF CLINICALLY IMPORTANT MYCOBACTERIA NON CHROMOGENES (Slow growers) 1. M. tuberculosis 2. M. avium intracellulare 3. M. bovis 4. M. celatum 5. M. gastri 6. M. genavense 7. M. hemophilum 8. M. malmoense 9. M. terrae complex 10. M.ulcerans RAPID GROWERS 1. M. chelonae 2. M. fortuitum group PHOTOCHROMOGENS (Slow growers) 1. M. asiaticum 2. M. kansasii 3. M. marinum 4. M. simiae SCOTOCHROMOGENS (Slow growers) 1. M. gordonae 2. M. szulgai 3. M. scrofulaceum 4. M. xenop Made by Miguel Astronomo & Daniel Budo 10