TUBERCULOSIS

Presenter Poonam Bsc MLT 3RD YEAR. Moderator- Dr.Neelam kaistha

Definition

Tuberculosis is an air-borne disease caused by Mycobacterium tuberculosis

It usually affects the lungs but can be present anywhere in the body.

History

In 1882, Robert Koch discovered TB.

Soon he found out that it was caused by a microorganism Mycobacterium tuberculosis. After discovering that this disease was infectious, he started to consider treatments. Many treatments were tried but none were discovered until the year of 1943 were the activity of Streptomycin was discovered.

Mode of transmission
The mode of infection is by

Inhalation of aerosol bacilli Coughing Sneezing

talking

Epidemiology

Tuberculosis is an ancient disease.

It has been called the ` white plague. More than 40% of the population are infected.

Each year 2 billion people or one third the earths population dies from tuberculosis.

Pathogenesis
Tuberculosis is divided into three clinically important categories: Primary TB Secondary Reactivated TB Disseminated TB

Primary Tuberculosis

Primary TB is the initial infection caused by inhalation of tubercle bacilli. Inhaled tubercle bacilli are engulfed by alveolar macrophages in which they replicate to form the initial lesion or Ghon focus. Ghon focus together with lymph nodes forms the

primary complex.
This occurs about 3-8 weeks from the time of infection. The lesion heals spontaneously 2-6 months leaving behind a calcified nodule. Few bacilli survived in the healed lesion & remain

Secondary or reactivated tuberculosis

This is caused by reactivation of the primary lesion ,or by exogenous reinfection.

This reactivation typically happens to those who have weakened or suppressed immune system.

Disseminated Tuberculosis
The spread of the disease most often occur in the apex of lungs but may be widely disseminated in the lungs, kidneys, meninges, bones and other organs

Types Based on organ System Infected

M.tuberculosis can involves various systems

causing Pulmonary tuberculosis Renal tuberculosis Tubercular meningitis Bone and joint tuberculosis Miliary tuberculosis Intestinal tuberculosis Skin tuberculosis

Symptoms
The symptoms of tuberculosis are-: Fever

Chronic cough Haemoptysis

Weight loss
Loss of appetite

Morphological features

Acid-fast rods of size about 2-3 0.4m Straight or slightly curved with rounded ands Branching and filamentous form also seen Beaded, barred forms- seen specially in sputum Arranged singly or in clumps. Non motile Nonsporing noncapsulated

Lab diagnosis
Lab diagnosis based upon two types-:

Pulmonary tuberculosis Extra pulmonary tuberculosis

Pulmonary tuberculosis
Specimens -: Sputum Laryngeal swab Bronchial washings Bronchial alveolar lavage(BAL) Gastric lavage

Collection
Sputum

Early morning sample. Three successive specimen should be examined.

Processing

Microscopic examination Culture method Microscopy &culture is done after concentration of specimen

Concentration of specimen
They can be classified into two types-: Methods that kill bacteria Treatments with hypochlorite Treatment with antiformin Autoclave method- they kill bacilli so cannot be used for culture
Methods that do not kill bacteria Petroffs method Modified Petroffs method

Petroffs method

Sputum is incubated with an equal volume of 4% sodium hydroxide solution at 37C with frequent shaking till it becomes clear for 20 mins. Then centrifuged at 3000rpm for 20 mins. Then neutralize the sediment with N/10 HCL. Then sediment is used for smear preparation and culture.

Extra pulmonary tuberculosis

Specimens-: C.S.F

Urine (in renal T.B)

Others as the site is involved for e.g. bone marrow, liver biopsy, pleural effusion etc.

Collection of specimens
C.S.F

Through lumbar puncture,in aseptic conditions. Best site for puncture is the interspace between 3rd and 4th lumber vertebrae.

URINE

Early morning atleast 30-40 ml midstream urine sample should be collected in two 25 ml bottles. For 3 consecutive days.

Processing
CSF Centrifuge the fluid. use the deposit and make the smear& culture.
URINE Centrifuge the Both specimen as received at1500g for 30 mins. Remove the supernatant and pool the centrifuge deposit. Treat the pooled deposit by Petroffs method. Make smear and culture.

Methods for demonstration

Ziehl-Neelsen stain-:shows acid- fast bacilli against blue /green background. Fluorescent stain-: shows yellow luminous rods against dark background. Sensitivity of microscopy is 10,000baclli /ml of sputum

Ziehl-Neelsen procedure

Heat fix the smear Put carbol fuchsin over it Heat from the below until fume appears,heat being applied at intervals to keep stain hot Keep for 5 mins Wash with water Pour 20% sulphuric acid until yellowish brown colour appears Wash with water Malachite green/methylene blue 1min. counter stain.

minimum 300oil immersion fields of Z.N stained smears are examined and reported as:

00 AFB in 300 OIF - AFB not seen 1-2 AFB in 300 OIF - doubtful-repeat smear 1-9AFB in 100 OIF - 1+ 1-9AFB in 10 OIF - 2+ 1-9 AFB in 1 OIF - 3+ 10 or more AFB IN 1 OIF - 4+

Types of culture media

Solid media Containing eggLowenstein-Jenson media Dorset egg medium Petragnini medium Containing blood- Tarshis medium Containing potato- Pawloskys medium Containing serum- Loefflers Serum Slope liquid medium

Liquid medium Proskaurs medium Dubos medium Middle brooks medium

Lowenstein Jensen(LJ) medium

Most commonly used .

Constituents:Coagulated hens egg Mineral salt solution Asparagine Malachite green Glycerol

Cultural characteristics
Solid media-:colonies are

Dry Rough Tough Buff Raised and wrinkled surface.

Liquid media-:growth occurs as

Bottom creeps up the side Forms prominent surface pellicle, extends along the sides above the medium

Identification
To identify isolate, following observations are helpful Slow growth at 36c taking 2-6 weeks No pigment production Weakly catalase positive Niacin-positive Nitrate reduction positive

Nucleic Acid detection method

Nucleic acid detection methods are Polymerase chain reaction(PCR)

Ligase chain reaction(LCR)

Nucleic acid sequence base amplification(NASBA) DNA probes

Antibody detection method

Ab detection methods are-:

Enzyme linked immunosorbent assay (ELISA)

Radioimmunoassay(RIA) Latex agglutination

Tuberculin test
Also called Mantoux test. Principle-: It is a type 4 hypersensitivity reaction.

Procedure-: Purified protein derivative(PPD) is inoculated intradermally on forearm. Site observed at 72 hours for appearance of erythma & induration.

Interpretation
About Erythma & induration-: 10mm positive test Below 5mm indicate indicates negative Between 5&9mm indicates doubtful test.

False positive Atypical Mycobacterial infection

False negative Early tuberculosis Miliary tuberculosis Advanced tuberculosis Immunosuppresive individuals Severe malnutrition.

Treatment
Bacteriocidal and Bacteriostatic drugs are available for treating tuberculosis

Bacteriocidal-: Rifampicin,isoniazid,pyrazinide, streptomycin. Bacteriostatic-: Ethambutol,cycloserine,capreomycin,kanamycin, ciprofloxacin

Prevention
By vaccination of BCG i.e. Bacilli Calmette-Guerin.

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