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Jour nal of Pharmacy Research Vol.4.I ssue 7. Jul y 2011

Ni lam A.Gadhave et al. / Jour nal of Pharmacy Research 2011,4(7),2107-2119

2107-2119

Review ArticleISSN: 0974-6943

Available online throughwww.jpronline.info

*Corresponding author.Nilam A. GadhaveSmt. Kashibai Navale College of Pharmacy,Saswad Kondhwa Road,S. No 40/4, near Octroi post,

Kondhawa (Bk),Pune 411048, Maharashtra, India.Tel.: + 91-9960237674

E-mail:[email protected]

INTRODUCTION

Tuberculosis: A dreaded or curable disease- A ReviewNilam A. Gadhave*, Kedar S. Lade 1 ,Meera C. Singh2, Sanjay D. Sawant3

Smt. Kashibai Navale College Of Pharmacy, Saswad Kondhwa Road, S. No 40/4, near Octroi post, Kondhawa (Bk), Pune 411048, Maharashtra, India.

Received on: 12-04-2011; Revised on: 18-05-2011; Accepted on:21-06-2011

ABSTRACTTuberculosis is a communicable chronic granulomatous disease caused by Mycobacterium tuberculosis is usually involves the lungs but may affect any organ ortissue in the body. Tuberculosis granulomas undergoes cereous necrosis among medically and economically deprived persons through out the world. TB continues to

be the leading killer disease for Indian adults amongst all infectious diseases. More than 80% of TB patients have pulmonary TB. The WHO recommended DOTSstrategy is the most effective tool for TB control. All treatment regimens is RNTCP have two phases; an intensive phase & continuation phase. MDR- TB is caused

by strain of the TB bacteria resistant to at least the two most effective Anti-TB drugs; namely Isoniazid & Rifampicin. To prevent MDR-TB is to ensure the provisionof effective regimens of short-course treatment with first-line drugs. Also laboratory test use for diagnosis of TB. It is estimated that one third of world population isinfected with TB. Development of drug resistance in the population has increased concern that TB may once again become an incurable disease (MDR-TB) .Theinsufficient knowledge of the mechanism of pathogenesis and protection from it, creates a need to establish long-term and effective tuberculosis treatment and control

programmes

Key words:Mycobacterium tuberculosis, MDR- TB, Middlebrooks medium, Lowenstein-Jensen medium.

Tuberculosis is an infectious disease caused by Mycobacterium tuberculosis.Pulmonary tuberculosis is the most common form of TB (more than 85% ofall TB cases.), While extra-pulmonary tuberculosis can affect almost anyorgan in the body. Transmission occurs by airborne spread of infectious dropletsand droplet nuclei containing the tubercle bacilli. The source of infection is a

person with sputum smear-posit ive pulmonary TB. Transmission often occursindoors, where droplets and droplet nuclei can stay in the air for a long time.

There were globally an estimated 8.8 million new cases of TB, of which 3.9million were sputum smear positive, and 80% were in 22 high burden countries.One fifth of the global TB incidence is in India, with 1.8 million new casesoccurring every year and 0.8 million of these being infectious smear-positivecases. In India, an estimated 4 lakh deaths occur from TB every year.Every Day, in India.More than 5000 develop TB disease.More than 1000 people die of TB ( 2 deaths every 3 minutes)

TB is a serious public health problem in India causing immense morbidity,mortality and distress to individuals, famili es and communities. TB kills moreadults in India than any other infectious disease. The disease incidence peaksin people belonging to the most economically productive age group of 15-60years. The link between TB and HIV is quite significant with WHO estimatingthat 5% of TB patients are also co infected with HIV.

Tuberculosis is a communicable chronic granulomatous disease caused byMycobacterium tuberculosis is usually involves the lung but may affectany organ or tissue in the body. Tuberculosis granulomas undergo cereousnecrosis among medically and economically deprived persons through out theworld.

TB remains a leading cause of death. Since time immemorial Tuberculosis(TB) has been a scourge to mankind. Fossil bones dating back to 8000 BC

provide the earl iest evidence of TB in man and animal . TB was respons ible for20% of deaths in London in 1651.

Benjamin Morton made the first credible speculation on the infection nature

of TB in 1722. In early 19th

Century, TB probably accounted for a third of alldeaths in Paris. In 1890, Koch announced that culture filtrate of tuberclebacilli tubercle bacil li cured the disease . Those filtrates, later partia lly purif ied,become the principal means to establish infection the tubercul in skin test .

During late 19 Century, pasteurization of cow, milk was started to reduce thepossibi lity of infection of Mycobacterium bovi s being a cause of human TB. In1908, Albert Calmette and Camille Guerin, seeking to overcome the problemof bacillary clumping associated with Mycobacteria, grew bovine tubercle bacilliin dispersed culture that contained ox bile.

The varian 231st passage was first used to immunize a child whose died of TBafter childbirth. This vaccine, BC G (bacilli Calmette Guerin) is still the mostwidely used vaccine in the world. The discovery of p -amino salicylic acid byBernhein in 1941 was the first step in the development of effective anti- TBtherapeutic agents. The introduction antibiotics, streptomycin in 1947 werethe first successful antibiotics in chemotherapeutics intervention of TB. Butits full therapeutic potential could be utilized only after 1952 when isoniazid(INH) was used simultaneously with streptomycin.

Though isoniazid was first synthesized in 1912, its effectiveness against TBwas not known for long time until its successful use in 1952. Even today,

isoniazid is the most potent anti-TB drug both singly and in combinations(including FDCs i.e. fixed dose combinations) it is the most important, effectiveand potent first line anti-TB drug susceptible strains of tubercular mycobacteriain man. The major problem of INH is the most commonly used anti-TB drug

because of it s better therapeutic efficacy and safety over other available anti-TB drugs.

Tuberculosis is estimated to affect 1.7 billion individual worldwide, with 8 to10 million new cases and 1.7 million deaths each year. After HIV, tuberculosisis the leading infection cause of death in the world. Infection with HIV makes

people susceptible to rapidly progressive tuberculosis; over 50 mill ion peopleare infected with both HIV and M. tuberculosis. From 1985 to 1992, thenumber of tuberculosis cases in the United States increased by 20% because ofincrease in the disease among people with HIV, among immigrants, and amongthose in jail or homeless shelters. Because of increased public health efforts,the number of cases of tuberculosis has declined since 1993. Currently, thereare about 16, 00 new cases of active tuberculosis in the united states annually,and about 45% of theses are in immigrants.

Tuberculosis flourishes wherever there is poverty, crowding, and chronicdebilitating illness. In the United States, tuberculosis is mainly a disease of theelderly, the urban poor, and people with AIDS. Certain disease states alsoincrease the risk; diabetes mellitus, Hodgkins lymphoma, chronic lung disease(particulary silicosis), chronic renal failure, malnutrition, alcoholism, andimmunosuppression.

More than half (50%) of cases involving foreign-born individuals in 2005 werereported in persons from Mexico (25%), Philippines (11%), Vietnam (8%),India (7%), & China (5%). Foreign-born persons account for a steadilyincreasing proportion of all reported TB cases. An estimated 10-15 million

people in t he Unit ed- stat es have latent infect ion.


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It is important that infection with M. tuberculosis be differentiated fromdisease. Infection is the presence of organism, which may or may not causeclinically significant disease.

Typical signs of TB are:Chronic or persistent cough and sputum production. If the disease is at anadvanced stage the sputum will contain blood.Fatigue.Lack of appetite.Fever.Night sweats.

The general symptoms of TB disease including feeling sick or weak, weightloss, fever and chest pain. Other symptoms depend on the part of body that isaffected.

Most infections are acquired by person-to-person transmission of airbornedroplets of organisms from an active case to a susceptible host. In most

people , primary tuberculosis is asymptomatic, although it may cause fever andpleural effusion . Generally , the only evidence of infection , if any remains, i sa tiny, fibrocalcific nodule at the site of the infection; viable organisms mayremain dormant in such lesions for decades. When the persons immune defensesare lowered, the infection may reactivate to produce communicable and

potentia lly l ife- threatening di sease.

Infection with M. tuberculosis typically leads to the development of delayedhypersensitivity to M. tuberculosis antigens, which can be detected by thetuberculin (Montoux) test. About 2 to 4 weeks after infection, intracutaneousinjection of purified derivative of M. tuberculosis (PPD) induces visible and

palpable induration that peaks in 48 to 72 hours.

TB :THE STUDYAs per WHO 1.7 billion people infected worldwide. 3 millions deaths occur peryear and about 8 million new cases admit per year.

Tuberculosis is an infectious disease, that usually affect the lungs but that canattack other parts of the body. There are two forms of tuberculosis-TB infection& TB disease. Most people with TB have TB infection.

Common symptoms of TB loss of strength, weight loss, rise in temperaturemostly in evening time, cough, pain in chest, blood in the sputum. And also acommon cause of TB Alcohol, smoking, mental stress & strain, unhealthyliving style, loss of sleep, exposure to cold, hence it is estimated that one thirdof world population is infected with TB, so it is required to development of

drugs resistance in population has increased concern that TB may once againbecome an incurable disease (MDR-TB) . Also there is insuffic ient knowledgeof the mechanisms of it s pathogenesis and protection from it, so it is necessaryto establish long-term and effective tuberculosis treatment and control

programmed.

CAUSATIVE ORGANISM - MYCOBACTERUM TUBERCULOSIS .1

Fi g1. M ycobacteri um tubercul osis. Acid-fast stain. CDC.

Mycobacteria are transition from between bacteria and fungi. These genusmycobacteria belong to the order actinomycetaes and family mycobacteriaseae.

Non-motil e, non- sporu lating rods that resis t decolorisation with acidifiedorganic solvents. For this reason, they are called acid-fast bacteria. Mycobacteriaare slender bacilli and sometimes exhibit filamentous forms resembling fungalmycelium.

They are difficult to stain by ordinary stains because of the presence of waxymaterials in their cell walls. Four very closely related species are responsiblefor mammalian tuberculosis, M. tuberculosis (human tubercle bacillus), and M.

bovine tubercle bacillus) M. microti (vole tubercle bacillus) and M. afr icanum(intermediate in form between human and bovine types) Tubercle bacilli arethin straight rods, measuring about 0.4-3. Mycobacteria are strict aerobesand derive energy from the oxidation of many simple carbon compounds.Increased CO2 tension enhances growth.

CULTURETubercle bacilli are aerobes, grow slowly (generation time 14-15hrs.) optimumtemperature 37C, PH-6.4-7.0 They grow only in specially enriched mediacontaining egg, asparagines, and potatoes, serum and meat extracts. Coloniesappear in 2-6 weeks. Mycobacteria are be killed by exposure to direct sunlightfor 2hrs. However bacilli may remain alive for 20-30hrs.In sputum cultureviable for 6-8 months at room temperature and may be stored for two yearsat-20C.

Reaction to physical and chemical agents-Mycobacteria tend to be more resistant to chemical agents than other bacteriaof the hydrophobic nature of cell surface and their clumped growth.Mycobacteria are relatively to chemical disinfectants surviving exposure.They are better stained by hot carbol fuchsin and once stained, they resistdecolourisation by dilute mineral acid and are therefore referred to as acid fast

bacilli is called as gram-positive bacteria.

CULTURE MEDIUM

A) LIQUID MEDIUMLiquid media are used for drug sensitivity, biochemical tests and preparation ofantigen and vaccine. The bacilli grow either on surface as pellicle or as flocculesthroughout the medium due to hydrophobic nature of their cell wall (lipid)diffuse bacterial growth can be obtained by addition of a detergent tween-80.Dubos medium containing casein hydrolysate, bovine serum albumin, aspargineand certain salts with Tween 80 is widely used liquid medium.

B) SOLID MEDIUMMost solid medium contain egg, blood, serum or potato. Lowenstein-Jensen(Lj)

Medium is the most widely used solid medium. It contains hens egg, aspargine,buffer solut ion and malachite green. Malachi te green inhibits contaminants.Glycerol is replaced by pyruvic acid. In Lj medium, visible colonies appearafter 2-6 week, incubation for 6-8 week before being discarded. The coloniesof M.tuberculosis appear dry rough, yellowish or buff colored and those ofM.bovise are moist, smmoth, flat and white.

Two media are used to grow M.TB.Middlebrooks medium which is an agarbased medium and Lowenstein-Jensen medium which is an egg basedmedium. M.TB. colonies are small and buff colored when grown on eithermedium. Both types of media contain inhibitor s to keep contaminants fromout-growing M.TB. It takes 4-6 weeks to get visual colonies on either type ofmedia.

Fi g 2. Colonies ofMycobacteri um tuberculosison Lowenstein-Jensen medium. CDC


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BIOCHEMICAL REACTIONThe following biochemical tests are helpful in distinguishing di fferent speciesof Mycobacteria.

1. NIACIN TESTAlthough all Mycobacteria produce niacin, a nicotinic acid, during growth, butonly M.Tuberculosis produces niacin, a sufficient quantities in egg medium togive a positive test. The reagent contains 10%cyanogens bromide & 40%anilinein ethanol. When a freshly prepared reagent is added to a suspension of

bacterial cultu re, a canary yellow color shows a posit ive react ion. Posit iveniacin test is also seen with culture of M.Chelonee and M. Simiae.

2. ARY SULPHATASE TESTThe test becomes positive in culture at atypical mycobacteria as they fromthe enzyme arylsulphatase. The bacteria are grown in media containingtripotassium phenolphthalein disulphate (0.001M) for 2-4 weeks. To theculture medium when 0.2 N NaOH is added drop by drop, a pink color developsin positive reaction due to liberation off f ree phenolphthalein.

3. CATALASE TESTMost atypical mycobacteria are strongly catalase positive and peroxide negative.The tubercle bacilli (M.Tuberculosis, M. bovis) are peroxides, positive andweakly. Catalase positive and weakly catalase positive. Tubercle bacilli loose

peroxides and ca talase ac tivi ty when they become INH- resis tant. The reagentis prepared by mixing equal volume of 30% volume H2O2 and 0.27% catecholin distilled water. This reagent is added to a 5ml test culture in phosphate

buffer (PH-7) at 68C in a water bath and left for few minutes for the reactionto occur catalase production is indicated by and effervescence and peroxidaseactivity by browning of colonies.

4. NEUTRAL RED TESTThe virulent, strains of tubercle bacilli can bind neutral red in alkaline buffersolution whereas avirulent strains fail to do so. Positive result are with theM.tuberculosis, M.bovis, M.avium, M.ulcerrans.

5. AMIDASE TESTAtypical mycobacteria are differentiated by their ability to split amides. Theamides commonly used for the test are acetamide, benzamide, carbamide,nicotinamide and pyrazinamide M.tuberculosulsis nicotinamidase and

pyrazinamidase.

6. NITRATE REDUCTION TESTM.tuberculosis can reduced nitrate but in M.bovis cannot. The test organism isSuspended in a buffer solution containing nitrate and incubated at 57 C for3hrs. Then sulphanilamides and N-Napthylethlene diamine dihydrochloridesolution is added. Positive reaction is indicated by development of pink or redcolor.

7. SUSCEPTIVILITY TO PYRAZINAMIDEM.tuberculosis is sensitive to 50g/ml pyraziamide while either mycobacteriaincluding M.bovis are resistant.

8. SUSCEPTABILITY TO THIOPHEN-2-CARBOXYLIC ACIDHYDRZIDE (TCH)The growth of M.tuberculosis is usually not inhibited by addition of 10 g/mlTCH. However south Indian strains of M.tuberculosis are susceptible M.bovisis usually susceptible to the chemical.

MODE OF TRANSMISSIONHuman being acquires infection with tubercle bacilli by one of the following

routes- Inhalation of organism present on free cough droplet or in dried sputumfrom an open case of pulmonary tuberculosis.Ingestion of the organism leads to development of tonsillar or intestinaltuberculosis. This mode of infection of human tubercle bacilli is from self-swallowing of infected sputum of an open case of pulmonary tuberculosis, oringestion of bovine tubercle bacilli from milk of diseased cows. Inoculation of the organism into the skin may rarely occur from infected

postmortem t issue. Tranplacental route result in development of congenital tuberculosis infoetus from infected mother& is a rare mode of transmission.

SPREAD OF TUBERCULOSISThe disease spreads in the body by various routes-

1. Local spreadThis takes place by macrophages carrying the bacilli i t to surrounding tissues.

2.Lymphatic spreadTuberculosis is primarily an infection of lymphoid tissues. The bacilli may

pass into lymphoid folli cles of pharynx, bronchi, i ntest ine or regional lymphnodes resulting in regional tuberculous lymphadenities which is typical ofchildren infections. Primary complex is primary focus with lymphangitis&lymphadenitis.

3. Haematogenous spreadThis occurs either as a result of tuberculous bacillaemia because of the drainageof lymphatics into the venous system or due to caseous material escapingthrough ulcerated wall of a vein. This produces millet seed-sized lesions indifferent organs of the body like lungs, liver, kidneys, bones and other tissuesand is known as milltary tuberculosis.

4. by the natural passagesa. Lung lesion into pleura (tuberculous pleurisy )

b. Transbronchia l spread into the adjacent lung segments .c. Tuberculous salpingitis into peritoneal cavity.(tuberculous peritonitis)d. Infected sputum into larynx (Tuberculous laryngitis)e. Swallowing of infected sputum (ileocaecal tuberculosis)f. Renal lesions into ureter &down to trigone of bladder.

HYPERSENSITIVITY AND IMMUNITY IN TUBERCULOSIS 3

Hypersensitivity or allergy, and immunity or residence, play a major role inthe development of lesions in tuberculosis. Tubercle bacilli as such do not

produce any toxins. Tissue changes seen in tuberculosis instead the result ofhost response to the organism which is in the form of development of cell-mediated hypersensitivity (or type IV hypersensitivity) and immunity. Boththese host responses develop as a consequence of several lipids present in themicroorganism which include the following.1. Mycosides such as cord factor which are essential for growth and virulenceof the organism in the animals; and2. Gycolipids present in the mycobacterial cell wall like Wax-D; which actsas an adjuvant acting along with tuberculoprotein.

It has been known since the time of Robert Koch that the tissue reaction totubercle bacilli is different in healthy animal that is previously ingested (primaryinfection) from an animal that is previously infected (secondary infection)

Fi gure No-3. The natural hi story and spectrum of tu berculosis.

1. In the primary infection, intradermal injection of tubercle bacilli into theskin of a healthy guinea pig evokes on visible reaction for 10-14 days. Afterthis period a nodule develops at the inoculation site, which subsequentlyulcerated and heals poorly as the guinea pig unlike human begins, does not

possess any natural resis tance . The regional lymph nodes also develop type ofhypersensitivity and are comparable to primary tuberculosis in children althoughhealing invariably occurs in children.

2. In the secondary infection, the sequence of changes is different. The tuberclebacil li are injected into the skin of the guinea pig that has been infected withtuberculosis 4-6 weeks earlier. In 1-2 days, the site of inoculation is induratewhich heals quickly and the regional lymph nodes are indicative ofhypersensitivity and immunity in the host.

Similar type of changes can be produced if injection of live tubercle bacilli isreplaced with old tuberculin (OT).

Hypersensitivity and immunity are closely and are initiated through Tlyumphocytes sensitized against specific antigen in tuberculin. As a result of


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this sensitization, lymphokines are released from T cells which induced increasedmicrobicidal activity of the macrophages.

PATHOGENENESIS OF TUBERCULOSIS-Stages of disease:

Disease progression depends on:Strain of M.TB. Prior exposure.Vaccination.Infectious dose.Immune status of the host.

Stage 1Droplet nuclei are inhaled. One droplet nuclei contains no more than 3 bacilliDroplet nuclei are so small that they can remain air-born for extended of timethe most effective ( infect ive) droplet nuclei tend to have a diameter of 5 m.Droplet nuclei are generated by during talking, coughing & sneezing. Coughinggenerates about 3000 droplet nuclei. Talking for 5 minutes generates. 3000droplet nuclei in one minute. Sneezing generates the most droplet nuclei byfar, which can spread to individuals up to 10 feet away.

Spread of droplet nuclei from one individual to another. CDC. After dropletnuclei are inhaled, the bacteria are nonspecifically taken up by alveolar mac-rophages. However, the macrophages are not activated and are unable todestroy the intracellular organism.

Tuberculosis begins when droplet nuclei reach the alveoli. When a personinhales air that contains droplets most of the larger droplets become lodged inthe upper respiratory tract (the nose and throat), where infection is unlikelyto develop. However, the smallerdroplet nuclei may reach the small air sacsof the lung (the alveoli), where infection begins.

Spread of droplet nuclei from one individual to another. CDC after dropletnuclei are inhaled, the bacteria are nonspecifically taken up by alveolarmacrophages. However the macrophages are not activated and are unable todestroy the intracellular organisms.

Tuberculosis begins when droplet nuclei reach the alveoli. When a personinhales air that contains droplet most of the larger droplets becomes lodged in

the upper respiratory tract. (The nose and throat), where infection is unlikelyto develop. However, the smaller droplet nuclei may reach the small air sacsof the lung (the alveoli), where infection begins.

Stage 2 Begins 7-21 days after initial infection. M.TB. multiplies virtually unrestrictedwithin inactivated macrophages until the macrophages are burst. Othermacrophages begin to extravassate from peripheral blood. These macrophagesalso phagocytes M.TB but they are also inactivated and hence destroy M.TB.

Stage 3 At these stages lymphocytes begins to infiltrate. The lymphocytes specifi callyT-cells recognize processed and presented M.TB. Antigen in context of MHCmolecules. This result in T- cell activation and the liberation of cytokinesincluding gamma interferon (IFN) The liberation of IFN causes in theactivation of macrophages. These activated macrophages are now capable ofdestroying M.TB.

It is at this stage that the individual becomes tuberculin-positive. This positivetuberculin reaction the result of the host developing a vigorous cell mediatedimmune (CMI) response. A CMI response must be mounted to control anM.T.B. infection. An antibody mediated immune (AMI) will not aid in thecontrol of a M.T.B infection because M.T.B is intracellular and if extra cellular,it is resistant to complement killing due to the high lipid concentrated in itscell wall.

Stage 4Although a CMI response is necessary to control an M.TB. Infection, it is alsoresponsible for much of the pathology associated with tuberculosis. Activatedmacrophages may release lytic enzymes and reactive intermediates thatfacilitate the development of immune pathology. Activated macrophages andT-cells also secrete cytokines that may also play a role in the development ofimmune pathology, including interleukin I (IL-I), tumor necrosis factor(TNF), and gamma IFN.

It is also at this stage that tubercle formation begins. The center of thetubercle is characterized by caseation necrosis meaning semi-solid orcheesy consistency M.T.B. cannot multiply within these tubercle becauseof the low PH and anoxic environment M.T.B. can however, persisted withinthese tubercle for extended periods.

Although many activated macrophages can be ford surrounding the tubercles,many other macrophages to replicate and hence the tubercle grows.The growingtubercle may invade a bronchus if this happens; M.T.B infection can spread toother parts of the lung. Similarly the tubercle may invade an artery or other

blood supply line. The hematogenous spread of M.T.B. may resul t in extrapulmonary tuberculosis otherwise known as milliary tuberculosis. The namemilliary is derived from the fact that metastasizing tubercles are about thesame size as a millet seed, a grain commonly grown in Africa.The secondarylesions caused by milliary TB can occur at almost any anatomical location,

but usually involve the genitou rinary system, bones, joints, lymph nodes, andperi toneum. These lesions are o f two types-

1.Exudative lesionsResult from the accumulation of PMNs around M.TB. Here the bacteriareplicate with virtually no resistance. This situation gives rise to the formationof a soft tubercle.

Fi gure No- 4 The sequence of events in Pri mary Pulmonary Tubercul osis.

1.Productive or granuliomatous lesions-Occur when the becomes hypersensitive to tuberculoprotein. This situationgives rise to the formation of a hard tubercle

For unknown reason the caseous centers of the tubercles liquefy. This liquid isvery conductive to M. TB. Growth and hence the organism begins to rapidlymultiply intracellular. After time, the large antigen load causes the walls ofnearby bronchi to becomes necrotic and rupture. This results in cavityformation. This also allows M.TB to spill into other airways and rapidlyspread to other parts of the lung.

As stated previously, only a very small percent of M.TB. Infection result indisease and even a smaller percentage of M.TB. Infections progress to anadvanced stage. Usually the host will begin to control the infection at some

point . When the primary l esion heals, it becomes fibrous and calcifies. When


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this happens the lesions is referred to as the Ghon complex. Depending on thesize and severity, the Ghon complex may never subside. Typically the Ghoncomplex is readily visible upon chest X-ray.Small metastatic foci containinglow numbers of M.TB may also calcify. However, in many cases these foci willcontain viable organisms. These foci are referred to Simon foci. The Simon

foci are also visible upon chest X- ray and are often the site of diseasereactivations.

TYPE OF TUBERCULOSIS 5 :Lung is the main organ affected in tuberculosis. Depending upon the type oftissue response and age, the infection with tubercle bacilli is of 2 main types.I.Primary tuberculosis &ll. Secondary tuberculosis.

I. Primary tuberculosis 5:The infection of an individual who has not been previously infected orimmunized is called primary tuberculosis orGhons complex or childhoodtuberculosis.

Fi gure No- 5 Primar y Plumonary Tuberculosis

The complex or Ghons complex is the lesion produced at the portal of entrywith foci in the draining lymphatic vessels and lymph nodes. The mostcommonly involved tissues for primary complex are lungs and hilar lymphnodes. The other tissues, which may show primary complex, are tonsils andcervical lymph nodes. And in the case of ingested bacilli the lesions may befound in small intestine and mesenteric lymph nodes.

The incidence of disseminated from of progressive primary tuberculosis isparticular ly high in immunocompromised host e.g. in pati ents of AIDS.Primary complex or Gnons complex in lungs consists of 3 components.

1. Pulmonary component lesions in the lung are the primary focus or Ghonsfocus. It is 1-2 cm solitary area of tuberculosis pneumonia located under the

pleura. In the pleura. In the lower part of upper lobe.

Fi gure No-6 Primary complex or Gnons complex

2. Lymphatic vessel component the lymphatic draining the lung lesion containphagocytes conta ining bacil li and may develop beaded. Mili tary tubercles

along the

The primary complex composed of three components: Ghons locus, draining

lymphatics and Hilar lymph nodes.

3. Lymph node component. This consists of enlarged hi lar and tracheobronahil

lymph nodes in the area drained. The affected lymph nodes are matted and

show caseation necrosis.

In the case of primary tuberculosis of alimentary tract due to ingestion of

tubercle bacilli, a small primary focus is seen in the intestine with enlargedmesenteric lymph nodes producing tabs mesenterica. The enlarged and caseousmesenteric lymph nodes may rupture into peritoneal cavity and causetuberculous peritonitis.

FATE OF PRIMARY TUBERCULOSIS 7Primary complex may have one of the followed sequences:1.The lesions of primary tuberculosis of lung commonly do not progress butinstead heal by fibrous, and in time undergo calcification and even ossification.2. In some cases, the primary focus in the lung continues to grow and thecaseous material is disseminated through bronchi to the other parts of thesame lung or the opposite lung. This is called progressive primary tuberculosis.3. At times, bacilli may enter the circulation through erosion in a blood vesseland spread to various tissues and organs. This is called primary militarytuberculosis and the lesions are seen in organs like liver, spleen, kidney, brainand bone marrow.

Fi gure No-7 Fate of Primary Tuberculosis

4. In certain circumstances like in lowered resistance and increasedhypersensitivity of the host, the healed l esion of primary tuberculosis may getreactivated. The bacilli lying dormant in a cellular caseous material are activatedand cause progressive secondary tuberculosis. It affects children morecommonly but adult may also develop this kind of progression.

II.SECONDARY TUBERCULOSIS 8 -The infection an aceullar who has been previously infected or sensitized iscalled secondary, or post previously or reinfection, or chronic tuberculosis.

The infection may be acquired from,1.Endogenous source such as reactivation or dormant primary complex.2.Exogenous source such as fresh dose of reinfection by the tubercle bacilli.

Secondary tuberculosis occurs most commonly in lungs in the resions of apex.Other sites and tissue, which can be involved, are tonsils. Pharynx, larynx,small intestine and skin. Secondary tuberculosis of other organs and tissues isdescribed in relevant chapters later while that of lungs is discussed here.

Fi gure No-8 Progressive Secondary Tuberculosis


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The risk of reactivation increases if the immune system is weakened for anyreason.On average, a normal person who has been infected with tuberculosis hasabout a 10% chance of developing a reactivation of the disease over the courseof their lifetime. In people with HIV, however, they have a risk of about 7%per year.In the past, it was thought that almost all adult cases of tuberculosis were dueto reactivation. However, newer testing methods have revealed that a sizeablenumber of adult cases may actually be due to newly acquired infections, especiallyin areas where there are a large number of people with tuberculosis.People from certain parts of the world, such as the Philippines, china, SoutheastAsia, Haiti, and India have a much higher risk of having resistant tuberculosis.Resistant tuberculosis occurs when the organism is not sensitive to the usualanti-tuberculosis medicines.

LABORATORY DIAGNOSIS OF TB:

A) SKIN TESTS:REAGENTS:1.OLD TUBERCULIN (OT):Kochs old tuberculin is a crude preparation of a 6-8 week culture filtrate oftubercle bacilli grown in 5% glycerol broth concentrated by evaporation on aheated water bath.

2.PURIFIED PROTEIN DERIVATIVE (PPD):

A purified preparation of active tuberculoprotein has prepared by Sichert(1941) from old culture by precipitating with 50% ammonium sulphate. Thisstandardized and stable antigen as purified protein derivative (ppd) is nowgenerally employed for allergic skin tests instead of OT.

.METHOD:The tuberculin tests are based on reaction to intradermal inoculation oftuberculins.1. This is the method of choice and performed by intracutaneous injection of0.1 ml of PPD into flexor aspect of forearm.

B) HEAF TEST:It makes use of multiple puncture apparatus; which pricks the skin through afilm of tuberculin.

RESULT:Result is read within 48-72 hrs after intradermal injections of 0.1 ml PPD.1.POSITIVE TEST:In a positive reaction there is a local oedema (indurations) of 10mm diametersurrounded by erythema at the site of injection. Positive test confirm pastinfection by tubercle bacilli but does not indicate present active stage of thedisease.

2.FALSE NEGATIVE RESULT:The test may become negative in tuberculosis patients with military tuberculosis.

C) BACTERIOLOGICAL DIAGNOSIS:SPECIMEN-Specimen is collected from the suspected site of lesion.Pulmonary tuberculosis-sputum.Meningitis-CSF.Bone and joints- aspirated fluid.Renal- three consecutive morning sample of urine.

DIRECT MICROSCOPY:Smears made from the specimen in new glass slides are stained by Ziehl-Neelsen method.The acid-fast bacilli appear as pink brightened rods. Smears stained byfluorescent stains such as auramine phenol or aura mine rhodamine are examinedunder UN light the bacilli appear as bright rods against dark background.

CONCENTRATION OF SPECIMENS:The bacteria are decontaminated and concentrated in a small volume withoutinactivation. Such concentrate is mostly used for culture and animal inoculationtest and sometime for smear examination.

METHODS IN USE:1.PETROFFS METHOD:Sputum mixed with equal volume of 4% sodium hydroxide is i nfected at 37cfor about 30 min. the mixture is frequently shaken till it gets liquefied andbecome clear. Sodium hydroxide also kills the contaminating bacteria. Aftercentrifugation at 3000 r. p. m. for 30 mins. It is neutralized with 8% HCL inpresence pf a drop of phenol red indicator . The depos it is used for smear, aculture and animal inoculation.

2.HOMOGENISATION METHOD:The specimen is treated with diluted acids as 8% sulphuric acid.3% hydrochloricacid or 5% oxalic acid then clearing the acid by repeated washing with sterilenormal saline.

3.TRISODIUM PHOSPHATE METHOD:Trisodium phosphate is lethal for many contaminating bacteria but tuberclebacilli remain unaffected.

SENSITIVITY TESTING:The isolated tubercle bacilli is tested for drug sensitivity in Dubos mediumor L Jmedium after incorporating different concentrations of anti-tuberculardrugs in the media before inoculation.

PREVENTION AND TREATMENT-

AIM OF TREATMENT:To cure the patient of TB.To prevent death from active TB or its late effects.To prevent relapse of TBTo decrease transmission of TB to others.To prevent of development of acquired drug resistance.It is vital to achieve these aims while preventing the selection of resistantbac ill i in infect ion pat ien ts. The the rap y of tub erc ulo sis has und erg oesremarkable chance during last two decades. The conventional 12-18 monthstreatment has been largely replaced by more effective and less toxic 6 monthstreatment, which also yield higher completion rates, i.e. better compliance.

VACCINES:Anti-tubercular vaccines were developed much before the development ofanti-tubercular TB drugs to prevent incidence or to build immunity against TBbut such vaccines have s ignifi cant failures too . E.g. BCG vaccine.

a. STRAINS OF BCG VACCINE:Albert Calmettte and Calmette Guerin (1921) prepared an attenuated strain ofM.bovis by growing it to potato medium for 13 years (1908-1921).When stain proved incapable of producing TB in susceptible guinea pig. It wasnamed Bacilli Calmette Guerin (BCG).

b.DOSE AND ADMINISTRATION:It is available in liquid and lyophilized from.The lyophilized vaccine is reconstituted by sterile physiological saline solutionto make final concentrating of 0.1mg in 0.1 ml of vaccine.Vaccine once reconstituted should be utilized within 3-6 hrs. Administrationby oral route. It i s now given intradermally .

c.AGE OF VACCINATION :It should be administered soon after birth or any time during the first year oflife.

d.OBSERVATION :A popular develops at the site of vaccination after 2-3 weeks after intradermalinjection, which increase slowly and in about five weeks it break into shallowulcer, which heals up leaving permanent round scar.Such individual becomes tuberculin positive after 4-6 weeks.

e.PROTECTIVE EFFICACY:Major Field trial of BCG vaccine was undertaken in different countries before1970 and result varied greatly ranging from 0 to 80%.

REGION AGE RANGE PROTECTION % YEAR OF(YEARS) COMMENCEMENT.

North American Ind ian 0-20 80 1935

Great Britain 14-15 78 1950Chicago, USA. Neonates 75 1937Puerto, Rico. 1-13 31 1949South India (Bang lore) All ages 30 1950Georgia & Alabama, USA. >5 14 1997Georigia, USA. 6-17 0 1950Illinois , USA Young adult 0 1947South India (Chinglepur) All age 0 1968

The difference in protection appears to be related to exposure of individual toenvironmental mycobacteria that had already occurred before BCG vaccination.The variation in protection may also be contributed by some other factorssuch as virulence of organism in different communities and potency of vaccines.

PRINCIPLE ROLE OF BCG:Vaccination does not give absolute protection against tuberculosis.Vaccines prevent serious forms of primary tuberculosis such as meningitis,skeletal TB and military spread of disease.


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B) CHEMOTHERAPUTIC AGENTS:M.Tuberculosis can cause chronic and fatal condition in humans known astuberculosis. No effective chemotherapeutic agent was available for treating.TB as late as1940.The discovery of p-amino salicylic acid by Bernhein in

Drugs used to treat TB disease. From left to right isoniazid, rifampin,pyrazinamide,and ethambutol. Streptomycin (not shown) is given byinjection. CDC.

Since the successful introduction of Rifampicin in the early 1970s as the Anti-TB drugs, TB control and treatment were largely successful during 1970s asthe combined use of available drugs.

This led to negligible of TB research due to less interest and less market. Nonew drugs were developed during the last thirty years. Though there werecontinuously increased since 1980s report of incidence of TB and Multi-drugsresistance TB (MDR-TB) and even co-infection with AIDS. The world healthorganization declared emergency in TB on 1993. Increasing report of MDR-TB since 1990s have posed a serious threat to mankind again as there will beno remedy or treatment of TB in man again in the face of war or challenge ofmycobacteria of MDR-TB.

PROFILE OF CURRENTLY AVAILABLE DRUGS FOR TB:Many drugs are selectively active against tubercle bacilli (TB) while someinhibit other microbes as well, though their use is primarily limited to TB.

FIRST LINE:These drugs have high antitubercular efficacy as well as low toxicity

SECOND LINE:These drugs have low antitubercular efficacy or high toxicity or both.

1. FIRST LINE AGENTS:

Drug Structure Mechanism of action

Isoniazid (IHD) a Mycolic acid synthesis is inhibited resulting in loss of some areas of outer membrane and thinness of the cellwall.

Streptomycin (Sm) b Inhibition of enzymatic polymerization of amino acids i.e.bactericidal.

Ethambutol ( Emb) c Prevent synthesis of protein and DNA and reduces RNAsynthesis

Rifampin (Rmp) d Inhibit DNA-directed RNA synthesis.Pyazinamide (Pzm) e Active against drug resistant strains of M. tuberculosis.

N

CONHNH2

isonicotinohydrazideO O

OH

OH

NH

OH

HN

CH O

H OO

OH

N H C H 3HO

HO H2 C

HN

NH 2

NH

H 2 N

H 3 C

1941 was the first step in the developmentof effective anti-TB therapeutic agents.

Remarkable progress has been made in thelast 45 years.

In 1943, anti-TB research resulted in thediscovery of active anti0TB antibiotic,Streptomycin.A number of clinically usefuldrugs have been discovered since 1940s,including isoniazid (1952), pyrazinamide(1952), cycloserine (1952) ethionamide(1950), rifampin (1971), ethambutol(1962).

NH

HN

OH

HO

O

O

O

N

O

OO

O

O

O

O

O

O

N

N

N

H

HH

H

H

H

H

H

HH

HHH

HH

H

H

H

H

H

H

HHH

HH

HH

H

H

H H

H

H

HH

H

H

H

H

H

H

H

H

H

H

H H

H

H H

H

H

HH

H

H

H

N

N

NH2

O

py r az in e- 2 - ca r bo xa m id e

a

b

c

d

e

Drug Adverse effects Route of Resistanceadministration

Isoniazid He pati ti s, Per ip her al Orally, Resistance is by mu tation of th e( INH) Neuropathy, Parent rally. of catalase perox idase gene so

Hypersensitivity. that the bacilli generate the

active metabolite of isoniazid.Streptomycin Ototoxicity, I nt ra mu scu lar ly , Many d rug s d ev elo p rap id( SM) Nephrotoxicity. intradermally. resistance to streptom ycin either

by one step mutation or byacquition of plasmid, whichcodes for inactivating enzymes.

Ethambutol Optic neuritis Orally. Resistance to ethambutol( Emb) and hypersensitive. develops slowly, in many cases

due to alteration in the drugtarget gene.

Rifampin Hepatitis, fever, Orally. Rifampin resistance is nearly(Rmp) thrombocytopenia. always due to mutation in the

repo B gene (the target ofrifampin action) reducing itsaffinity for the drug.

Pyrizinamide Hepatotoxicity, Orally. Resistance develops rapidly if it(pzm) Hyperuricemia. is used alone, & is due to

mutation in the gene whichencodes for the enzymesgenerating the active metaboliteof pyrizinamide.

II.Second line agents:

NH 2

OH

OHO

4-amino-2-hydroxybenzoic acid

a

O

NH2O

OOH

NH2

HO

HUH2

C

HO

H2N

HO

HO

H2NH2C

OH

bC 2H5

SNH2

3-ethylbenzothioamide

cN H

O

O

H2

N

4-a mi no i soxa z o l i d i n -3 -one

O

O

O

O

O

O

O

N

N

N

N

N

NN

N

N

N

N

N

N

N

H

H H

H

H

H

H

HH

H

H

H

H

H

H

H

HH

H

H

H

H

HH

HH

H

HH

HH

H

H

H

H

H

H

H

H

H

H

HH

H

d

e

Drug Adverse effects Route of administration.

P-amino salicylic GI in to le rance, Hepatotoxic ity, Ora lly.acid ( PAS) Hypersensitivity, Fluid retention.Kanamycin Ototoxici ty , Nephrotoxici ty , Int ramuscular ly .Ethionamide Oto tox icit y, Ne ph ro tox icity , O ral ly .

Hypersensitivity.Cycoloserin Hepatotoxicity, Hyperuricemain. Orally.Caprepmycin Ototoxici ty , Nephrotoxici ty . Int ramuscular ly .

Drug Structure Mecanism of action

p-amino salicylic acid (PAS) a Inhibit respiration of M. tuberculosis.Kanamycin b Aminoglycosi des like kanamyc in irreversi bly

bind to specific 30S-subunit proteins and 16SrRNA. Specifically Kanamycin binds to fournucleotides of 16S rRNA and a single amino acidof protein S12. This interferes with decoding site inthe vicinity of nucleotide 1400 in 16S rRNA of 30Ssubunit.

Ethionamide c Inhib it peptide synthesi zing M.TB by b locking theincorporation of amino acids containing sulphur(cysteine, Methionin.)

Cycoloserin d Active against drug resistant strains ofM.tuberculosis.

Caprepmycin e Inhibition of protein synthesis.


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Journal of Pharmacy Research Vol.4.I ssue 7. July 2011

Ni lam A .Gadhave et al. / Journal of Pharmacy Research 2011,4(7),2107-2119

2107-2119

Sputum smear microscopy provides definitive diagnosis. It is easy to perfo rm,is replicable and cost effective. If good diagnosis practices are followed. It isexpected that more than 50% of the new pulmonary TB patients will be smear

posit ive. Chest X-ray examination, though sens itive, is not speci fic and has awide inter-observer variability. The tuberculin test gives only evidence of

infection and does not differentiate between infection and disease. ESR is alsonot specific and is unreliable as a tool for diagnosis or evaluation. Culture ofsputum is highly sensitive and specific, but it i s costly, not freely available andtakes a long time in reporting.

Other new investigation are not reliable, are expensive and not useful intackling a public health problem like TB. The high false positivity (non- TB

patients ge tting diagnosed as TB) and low replicabili ty (repeated tests givingdifferent results.) are the factors which make newer tests inferior to sputumsmear examination. Besides, the high specificity of sputum microscopy (onlyactual TB cases getting diagnosed as positive) almost eliminates the chance ofa non-TB patients getting treated for TB.

Domiciliary chemotherapy has been proved to be as effective as sanatoriatreatment. Studies in the India have shown that TB patients do not get anyadditional benefit by bed rest or special diet. The risk of infection amongcontacts of patients does not increase with domiciliary treatment and there isno need for hospitalization for most patients. The economic burden on societyis also lowered by domiciliary treatment.

TYPE OF TB BACILLI EFFECTIVE DRUGS

Extra-cellular rapidly multiplying Rifampicin, Isoniazid, Streptomycin,

Ethambutol.Extra-cellular intermittently multiplying/ semi-dormant Rifampicin.

Intra- and extra-cellular acidic environments intermittently Pyrazinamide.multiplying/semi-dormant.

Dormant. No drug.

The phenomenon of Persisters expla ins to some extent why all bacilli are notkilled during treatment. Relapse with drug-susceptible organisms after the endof treatment or endogenous reactivation may be bacilli that have persisted inresidual lesion for a long time in a semi-dormant/ dormant state.

Fi gure No-14 DOT S Theropy

DOTS THEROPY14:

DOTS (Directly observed treatment, short- course chemotherapy.)The essential principles of DOTS were first demonstrated in India. Thedetermine that tuberculosis patients need not be hospitalized and the necessitytreatment was first proven at the tuberculosis research center in Chennai inthe 1950s and 1960s.

The key to the success & DOTS strategy is that is place s the responsibility forcuring TB patients on the health workers not the patient on the healthworkers not the patients. This strategy proven successful through out the than550 million people with cure rate of more than 90%.

COMPONENTS OF DOTS :A DOT is a systematic strategy having 5 components: Political and administrative commitment. Good quality diagnoses, primarily by sputum smear microscopy. Uninterrupted supply of good quality drugs. Directly observed treatment (DOT). Systematic monitoring and accountability.

Political and administrative commitment:TB is the leading infectious cause of death among adults. As TB can be cured,and the epidemic reversed, it warrants topmost priority. TB control has beengiven this priority by the Government of India as well as the state, district andlocal governments and administration.

Good quality diagnosis:Diagnosis of TB is made primarily by sputum smear microscopy among chestsymptomatic patients attending health facilities. This policy allows effectivediagnosis in all setting and appropriate prioritization of efforts. Sputummicroscopy is a reliable, simple and cost effective test.

Good quality drugs:An uninterrupted supply of good quality anti-TB drugs must be available. Inthe revised national TB control programme (RNTCP).A box of medicationcontaining the entire course of treatment is earmarked for each patient,ensuring the availability of drugs for the full course of treatment. Hence inRNTCP, the treatment never fails on account of non-availability of medicines.Short- course chemotherapy given in a programme of direct observation:

The RNTCP uses the anti-TB drugs available. But, unless these drugs areconsumed properly by the patient, the treatment will fail. This is why theheart of the DOTS strategy is Directly Observed Treatment (DOTS) inwhich a health worker or a trained person watches which the patient swallowsthe medicines. This simple principle ensure adherence, helps prevent drugresistance to anti-TB medicines and achieves high cure rates.

Systematic monitoring and accountability :Quality of treatment is monitored by1) Follow-up sputum examinations during and at the treatment, and2) By using a robust and comprehensive recording and reporting system thatevaluates and monitors the outcomes the outcomes of every patient ontreatment. This is carried out by trained supervisory staff in the programme.Sputum smear conversion rate, cure rate and other key indicators are monitoredat different levels. If a defined area is not achieving 90% sputum smearconversion rate at the end of three months and 85% cure rate, supervision isintensified.

In RNTCP, the patient is the VIP . RNTCP has shifted the responsibility forcure from the patient to the health system.

SCIENTIFIC BASIS OF DOTS :DOTS is primarily based on sputum microscopy, domiliary treatment, shortcourse chemotherapy, and directly observed treatment.

Fully intermittent chemotherapy has been found to be as effective as dailychemotherapy. Studies in animal models demonstrated that for Isoniazid,Rifampicin, And Pyrazinamide, intermittent dosing actually increased theefficacy of treatment. In vitro experiments demonstrated that, after a cultureof M. tuberculosis is exposed to certain drugs for some time, it takes severaldays before new growth occurs ( Thelag phase ) all the commonly usedanti-TB drugs induce lag phases ranging from two days to forty days. Thereforethese drugs can be given intermittently; intermitten t regimens make treatmentobservation more feasible and convenient for DOT relapse rates. As the

quality of drugs consumed is less, adverse reactions and costs are allowingorganisms to re-enter the active metabolite phase in which the bactericidaldrugs are more effective.

Direct observation of treatment (DOT):Ensures the best possible results in treatment of TB. Here an observer watchesand assists the patient in swallowing the tablet, thereby ensuring that the

pati ent rece ives the medication. Many patients who do not rece ive directlyobserved treatment stop taking drugs after two months because they feel

better. Studies in India and many other countries consistent ly shows that atleast one third of patients do not take medicines regularly. It is neither possibleto predict who these patients will be nor to reliably prevents during the entirecourse of treatment health education.

Studies have shown that there will be poor treatment outcomes and high deathrates in the absence of direct observation, even when regular supply of drugs isensured. Hence, by observing the patients during the entire course of treatment,one ensures that they right drugs, in the right doses, at the right intervals andfore the right duration.

The bacillary sub-population in TB is of four kinds and different drugs act ondifferent kind of bacteria. These are tabled below:

Several studies in India by reputed researchers have shown that six months ofchemotherapy gives favourable results when compared with longer terms oftreatment for new sputum positive TB patients. It has also been proven thateight months of treatment is adequate for patient undergoing retreatment.These durations of treatment are adequate to prevent emergence of drugresistance. Short-coursechemotherapy (SCC) is therefore more convenientand economical than convectional treatment with longer duration. In additionthe shorter period of time makes direct observation more feasible and improves

pat ien t ad herence to tr eatmen t.


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Currently recommended treatment regimens aim to:Cure the patient.Prevent death from active disease or form its late effects.Prevent the emergence and spread of drug-resistant organisms. Minimum relapse.Protect the community from continued of infection.All treatment regimens consist of two phases:

INTENSIVE PHASE (IP):Aims for a rapid killing of bacilli. This means a shorter duration of infectiousness(d2 weeks), usually with rapid smear conversion (80%-90%) after 2 to 3months of treatment. Each dose in this phase should be directly observed toensure adherence and involves 3 to 5 drugs depending on the category intowhich the patient has been placed.

CONTINUATION PHASE (CP):Eliminates most residual bacilli and reduces failures and relapses. At the startof the CP, there are low numbers of bacilli and less chance of drug resistancemutants. Therefore fewer drugs are needed during this phase.

NUMERICAL FACTOR:The number of tubercle bacilli in a patient varies widely with the type of lesionthe patient has. Data shows that the number of bacilli in a medium- sizedcavity communicating with the bronchi is about 10 (one hundred million)whereas, in an encapsulated nodular lesion of the same size with no bronchialcommunication. It can be as low as 10 (one hundred). The numbers are alsolow in extra-pulmonary lesions of the skin, lymph glands, meninges and

bones . The large r the bacteria l load, the higher is the probabil ity tha t resis tantmutant are present even before treatment has been initiated.

EXTRA- PULMONARY TUBERCULOSIS (EP TB):The management of extra- pulmonary TB differs forms that of TB mainlydue to the difficulty in making a conclusive diagnosis. This can lead to empiricaltreatment and thereby to over-diagnosis and unnecessary treatment of a largenumber of patients.

EP TB is usually paucibacillary (with relatively less number of bacilli). Anytreatment regimen of treatment, extra-pulmonary TB is classified into seriouslyill and not-seriously ill forms (detailed under the section on treatment). Thedifficulty in establishing a clear-cut end-point of treatment in EP TB oftenleads to treating these patients for longer durations then required. Studies haveclearly established the efficacy of short-course chemotherapy in children andadults. Intermittent regions in EP TB have also been proven to be as effectiveas daily regimens.

CATEGORIES OF TREATMENT:

CATEGORY OF TYPE OF PATIENT REGIMENTREATMENT

Category I New sputum smear-positive. 2H3RZ3E3+4H3R3.Seriously ill **new sputum smear-negative.Seriously ill** new extra-pulmonary.

Category II Sputum smear-positive relapse 2H3R3Z3E3S3+ 1H3R3Z3E3+5H3R3E3.Sputum smear-positive failureSputum smear-positive treatment after defaultOther***

Category III New sputum smear-negative, not seriously ill 2H3R3Z3+4H3R3.New extra-pulmonary, not seriously ill.

The subscript after the letters refers to the number of dosage per week. Thedosage strengths are as follows: H: Isoniazid (600mg), R: Rifampicin (450mg),Z: Pyrazinamide (1500mg), E: Ethambutol (1200mg), S: Streptomycin(750mg).

** Seriously ill also including, any patient, pulmonary or extra-pulmonary

who is HIV positive?*** In rare and exceptional cases, patients who are sputum smear-negative orwho have extra-pulmonary disease can have Relapse or Failure.

The codes and dosage strengths:

MEDICATION DOSE(THRICE A WEEK) NUMBER OF PILL IN COMBIPAKE.

ISONIAZID 600 mg 2RIFAMPICIN 450 mg 1PYRAZINAMIDE 1500 mg 2ETHAMBUTOL 1200 mg 2STREPTOMYCIN 0.75 mg _

CATEGORY I:This is used for new patients who are sputum smear-positive or seriously illsputum smear-negative or seriously ill EP TB patients. Treatment is given intwo phases intensive phase (IP) and continua tion phases (CP). IP consists of

Isoniazid, Rifampicin, Pyrazinamide and Ethambutol, given for two months(8 weeks: 24 doses). At the time of the 22 dose, the patients is referred forfollow-up sputum examination so that the result are available by the end of IP( 24 dose), if follow-up sputum examination is negative, CP is started andconsists of 4 months of treatment ( 18 week; 54 doses) with Isoniazid and

Rifampicin, given three times a week is directly observed. Alternate days.During CP. at least the first dose two months into the start of and at the endof CP.

If the sputum smear is positive after 2 months of treatment, IP drugs areextended for another one month (12 doses from a prolongation pouch) beforestarting CP. A follow up sputum examination is repeated at the extended IP (3months). This helps in assessment of the progress of treatment and sputumconversion rate. Irrespective of the smear status, the patient is put on CPdrugs and next follow up sputum examination is done at 5 months.

If the sputum smear is positive after 5 months or more of started tr eatment,the treatment outcome of the patient is declared as failure and (s) he isstarted afresh on CAT II treatment. in the rare situation in which a CAT Ismear-positive patient has a negative sputum after 2 months of treatment anda positive sputum smear after 4 months of treatment , another sputum smearshould be examined at 5 months. If this is also positive, the patient is considereda treatment failure and is started afresh on CAT II treatment.

CATEGORY II:This is used to treat retreatment cases namely relapse failure treatmentafter default and others. Such patients are generally sputum smear-positive,the IP consists of two months (8 week: 24 doses) of Isoniazid, Rifampicin,Pyrazinamide, Ethambutol and streptomycin; all given under directs observationthree times a week on alternate days. This is followed by another one months(4 week; 12 doses) of Isoniazid, Rifampicin, Pyraziamide and Ethambutol, allof which are given under direct observation three a week on alternate days.This is immediately followed by CP, which consists of 5 months (22 weeks, 66doses) of Isoniazid, Rifampicin and Ethambutol given three a week on alternatedays. The first dose of every week being directly observed. If the sputumsmear is positive after 3 months of treatment, the four oral IP drugs arecontinued for another one month (4 week, 12 doses) before starting the 5-months CP.

Patients who have been previously treated are at an increased risk of havingdrug resistant bacilli. For this reason, such patients are given a more intensiveregimen, namely CAT II. Experience in India and elsewhere has shown thatCAT II treatment, if taken regularly by the patient, is effective and result in

curing in most patients.

Patients who relapse generally have better outcomes than those who arefailure or treatment after default cases, but even these latter types of patientsrespond well to treatment, provided them it regularly and are well supervised.

CATEGORY III:This is used for patients who are smear- negative PTB or EP TB, and are notseriously ill. The IP lasts for 2 months (8 weeks; 24 doses) with Isoniazid,Rifampicin, and And Pyrazinamide given under direct observation three timesa week on alternate days. This is followed by CP, which consists of 4 months(18 weeks: 54 doses) of first dose of every week being directly observed. If a

patients receiving CAT III regimen has a posi tive sputum smear at any stageof the treatment, (s) he should be declared a failure, re-registered, and treatedafresh with the CAT II regimen.

TREATMENT REGIMEN:

CATEGORY OF PHASE PERIOD & DRUG REGIMENTREATMENT TREATMENT

Category I 1) Intensive phase. 2 months Isoniazid- 2 tab of 300 mg rifampicin- 1 capof 450 mg pyrazinamide- 2 tab of 750 mgethambutol 2 tab of 600 mg.

2) Continuous phase. 4 months Isoniazid- 2 tab of 300 mg.Rifampicin- 1 cap of 450 mg.

Category II 1) Int ensive phase . 3 months Isoniaz id- 2 t ab of 300 mgRifampicin-1 cap of 450 mg.Ethambutol- 2 tab of 600 mg.Inj. Of streptomycin 0.75 g one inj alternateday only for 2 months.

2) Continuous phase. 5 months . Isoniazid- 2 tab of 300 mg.Rifampicin - 1 cap of 450 mg.Ethambutol- 2 tab of 600 mg

Category III 1)Intensive phase 2 mo nths. Ison iazid 2 tab of 300 mgRifampicin- 1 cap of 450 mg.Pyrazinamide 2 tab of 750 mg.

2)Cont inuous phase 4 months Isoniaz id- 2 tab of 300 mg.Rifampicin 1 cap of 450 mg.


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TREATMENT OF REGIMEN OF TB:THE RELATIVE ACTIVITY OF FIRST- LINE DRUGS IN ACHIEVING THESE GOALS IS:

GOALS HIGH MODERATE LOW

Early bactericidal ING Rmp, Emb Sm, Pzm.Sterilizing

Rmp,Pzm. ING Sm, Emb.Resistance prevention INH, Rmp Emb, Sm Pzm.

CONVENTIONAL REGIMEN:These consist of INH+Tzn (Thiacetazone) or Emb with or without Sm (forinitial 2 months) optimum result require 12-18 months. To improvescompliance and to reduce treatment cost, after the initial 2 months of dailytherapy as above drug (unhandy other) are given twice a week under supervision.

SHORT COURSE REGIMENS:Trail of combinations of drugs with high bactericidal, sterilizing and resistance

preventing activiti es have shown tha t durat ion of tr eatment can be reduced to6-9 months.

THE REGIMENS BEING USED ARE:Four drugs INH+ Rmp+ Pzn+ Emb or Sm daily 2 months, followed by 2 drugsINH+ Rmp daily for another 4 months, total 6 months.The above initialtreatment followed by INH+ Tzn or Emb daily for another 6 months, total 8months. This reduces cost of treatment.Thrice a week INH+R mp+ Pzm Sm

or Emb under supervision for 2 months with INH+Rmp+Rmp thrice a weekfor another 4 months in continuation phase, total 6 months.Recently it hasbeen shown that Sm/Emb can eliminate as a routine adjunct unless INH resistanceis suspected.

TREATMENT OF RESISTANT CASES:When sputum remains positive even after 6 months treatment with anyregimen, resistant organisms are most likely present.

THE CHOICE OF DRUGS DEPSNDS ON:Drug combination employed in the previous regimen.Dosage and regularly with which these drugs were taken.Presence of associated disease if any-leukemias, diabetes, ADIS.The NTP regimen for relapse & failure cases is INH+ Rmp+ Pzn+ Sm for 2months then 4 drugs(-sm) for another 1 months and 3 drugs ( INH + Rmp +Emb) for next 5 months.( total 8 months)If sputum is found positive at 3 months, the 4 drugs therapy is extended by 1month more.

EXTRAPULMONARY TUBERCULOSIS:Fresh patient of abdominal, genitourinary, lymphatic bone/ joint, disseminatedor meningeal tuberculosis are treated on the same lines with the same drugs onfor pulmonary tuberculosis.INH, Pzn, Emb, Cys (Cycloserine) have very goodCSF penetrability.In tubercle meningitis higher dose of INH (with pyridoxinecover) its used and Pzm is always included along with Rmp+ Emb or Sm. It isalso advised to extend the total duration of treatment to 9 months.

TUBERCULOSIS IN PREGNACY:INH, Rmp, Emb and possibly pzn are to the foetus. The standard 6 monthsINH+ Rmp (with pzn for initial 2 months) should be added during late but notearly in pregnancy.

TUBERCULOSIS IN ADIS PATIENTS:The standard short course regimen with INH+ Rmp (pzn + Emb for initial 2months)Be extended to 9 months.Another regimen of INH + Rmp+ Emb for 6 months supplemented with pzn inthe first 2 months has been found satisfactory.

PROBLEMS DUE TO IGNORANCE:Our ignorance of molecular of molecular basis of pathogenesis of TB andvirulence of causative and Mycobacteria is great. Mycobacteria are dauntingorganism to study. Inn contrast to the most commonly used organism formolecular biological studies, E. coli which produces a visible colon (10.7

bacilli) in about 8 hrs. M.tuberculosis requires 3 to 4 weeks to yield a comparablecolony. It has a formidable waxy coat composed of multiple complex lipidsand carbohydrates that renders it impermeable to many common drugs. The

bacil li tend to clump, which makes working with them and quant itation ofthem is difficult, Because of its ability to remain dormant working for a longtime after infection and sudden activation due to unknown reasons, researchersavoid working with Mycobacterium out of fear of being contaminated andinfected. Current bio safety regulations require that work with is expensiveand not widely available. Hence, research the pathogen is slow and demanding.

The problem is multiplied due to increasing emergence of multi-drug-resistancestrains of M. tuberculosis. Scince, 1980s and further complicated with TB inADIS patient with less immune capacity making them more susceptible toinfection.

The problem is aggravated further due to our insufficient knowledge about thedetail molecular mechanism of action of anti- TB drugs and on developmentof new effective better anti-TB drugs and on development of new effective

better anti- TB drugs during last three decades. Though the Mycobacteri a isgradually developing its own mechanism of living in presence of our arsenals(i.e the existing anti-TB drugs) by natural mutational changes.

CURRENT RESEARCH PROGRAMS ON TB:Due to improper and incomplete implementation of prescribed drugs- regimens,the resurgence of TB particularly with drug resistan t strains, the resurgence ofTB particularly with drug resistant strains, the TAACF southern Researchinstitute, P.O. Box 55305.Birmingham.AL. Is giving services to investigationfor screening their anti- TB drug samples.

Since the later of 1994, the TAACFhas actively pursued comp for screeningfrom academic, government agencies, and industry.The ongoing efforts of the TAACFcontinually reward the scientific communitywith valuable data on potential drug candidates with activity against tuberculosis.

Word Health Organization (WHO) has declared emergency research intuberculosis region and so it is motivating new researchers to work in this areaby providing fund and facilities.

Pharmaceutical companies like Lupine, Astra Zeneca, Haffkine.Wockhardt:Sadoz -novartis are seriously involved in research on TB andalso helping new researchers.

Many government funded research organizations and universities are alsoseriously involved in research on TB, its prevention, control and treatmentthrough various programs.

STRATEGIES FOR ANTI MYCOBACTERIA:1)Analogs and derivatives for existing and their SARImproved therapeutic indexActivity against drug sensitive strainsImproved bioavailabilityBetter tolerance and safely pharmacokinetics.2)Broad Screening

Nove l mode of action3) Target directedMycobacterial specific targetsE.g. Arabinomannan, liporabinogalactan, and Mycolic acid.

GOALS OF THE PROGRAMS:1)New anti- TB drugs preferentially for use against MDR TB.

New mechanism of action2) New anti TB drugs are more potent, selective and quicker actingDecrease over all treatment timeBactericidal activity3) New anti TB drugs that can be effective in a drugs cocktail4) Facilities early stage drug development5) Encourge the development of research ideas relating to anti MycobacteriadrugDevelopment New Molecules, Mechanism of action, Molecular biology, Newtargets,The Global Alliance for TB drug development is interested in forwarding rubeloses drug development projects throughout the R & D process, but is specifically

interested in drug discovery (preclinical)projects clinical development projects.

The Alliance recognizes three primary reasons why new TB drugs are neededspecifically. Shorter and more easily administered treatments improvedtreatment of patients with MDR TB More acceptable and effective treatmentof high risk patients with latent TB infection.

DRUGS UNDER DEVELOPMENT:It is recommended that the compounds listed below are worthy of further

pursuit by global R & D community.

A) DISCOVERY RESEARCH: Thiolactomycin analogues.

SWOT analysis.


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2107-2119

STRENGTHS WEAKNESSES

Synthetic routes to thiolactomycin and analogues have been No report of in vivo activi tyM.developed.The mechanism of action is known. tuberculosis.

Present molecule is chiral cost.Active against isoniazid- resistant strains. Implications if analogues also

chiral.Well absorbed orally.OPPORTUNITIES THREATS

Both pharmas and academia are pursuing inhibitors of FAS-II and Unknown at this stage.progre ss might be a ccelerat ed by col laborat ing with GAT B/TDR.

Mefloguine/halofantrine analogues.SWOT analysis.


More potent in vivo than ethambutol. Cytostatic action.Likely to have less toxicity than ethambutol. Parent molecule is chiral-cost

implications if analoguesalsochrial

OPPORTUNITIES THREATS

Progress might be accelerated by collaborating with GATB/TDR. Unknown at this stage.

B)PRECLINICAL DEVELOPMENT:

CALANOLIDES A AND B.SWOT analysis.


Possible dual role in treating TB and ADIS. For the TB indication, no Ivivo efficacy. ADME or

toxicity data

Synthetic route developed to calanolide A and analogues calanolide Breadily obtaibale B readily obtaibale from renewable natural sources.

No cross - resista nt with r ifampici n or str eptomy cin.OPPORTUNITIES THREATS

Collaboration with GATB / WHO could induce Sarawak to proceed Unless funds become available,

with the TB indication. Sarawak will concentrate onthe ADIS indication.

QUINOLONES PD 161148 AND CS- 940SWOT analysis;


Potent bactericidal activity. No in vivo TB efficacy, ADME or toxic ity dataava il able .

Active against both quinolone-sensitive and- From structural considerations, the molecules

resistant M. tuberculosis. do not appear to be too metabolically robust.

OPPORTUNITIES THREATSCollaboration with GATB / WHO might induce Numerou s other q uinolo nes are in develo pment.

pharmas topro gress th e compo undsra pidly f or TB

.

POLOXAMER 315.SWOT analysis;


Acts on microbes extra & intra cellularly. Non-sp ecific mechan ism of action .Synergises with other anti-TB drugs. Accumulation in tissues may be problematic with

long treatment regimensIND obtained.


Collaboration with GATB / WHO may None p ercei ved.encourage clinical progression for TB indication.

C) CLINICAL DEVELOPMENT:

MOXIFLOXACINSWOT analysis


Safe and well tolerated No clinical d ata for TB indi cation.

Excellent oral bioavailability & long 11/2

Already registered in Germany other approvals expected soon. OPPORTUNITIES THREATS

Collaboration with GATB/WHO might encourage Bayer to Many other quinolones either ingenerate clinical TB data. development or marked already.

SITAFLOXACINSWOT analysis;


Potent 4th generation fluroquinolones. No in vivo TB data.

Well absorbed and tolerated in man.


Collaboration with GATB/WHO might encourage The company has indicated a lake of

indication. enthusiasm for developing the drug for a TBDalichi to generate clinical TB data.

GEMIFLOXACIN

SWOT analysis;


Potent 4th generation fluoroquinolone. No in vivo TB data.Well absorbed and tolerated in man. Data from healthy volunteers suggest difficulty

in reaching a therapeutic dose for TB.OPPORTUNITIES THREATSCollaboration with GATB/WHO might encourage SKB Many other quinolones either in development oror LG chemicals to generate more definitive TB data. marketed already.

T-3811ME.SWOT analysis:


Potent, broad-spectrum quinolone lacking 6-F atom present in all No in vivo TB data.other 3rd /4th generation fluoroquinolones. No ADME data in public domain.Progressing into phase III trial so presumably well absorbed andtolerated in man.OPPORTUNITIES THREATSCollaboration with GATB/WHO might encourage Toyoma or BMS The company has indicated a lakechemicals to generate more TB data. of enthusiasm for developing the

drugs for a TB indication.

It was also recommended that the families of antifungal azoles, antibacterialnitroimidazole and oxazolidinones be further studied in the discovery research

phase in an iden tify molecules with grea ter anti -TB potenti al than currentcandidates which have been discussed specifically in this report. The use ofinhibitors of bacterial drug efflux mechanism and small molecules

immunomodulators were also felt to be areas worth investigation further inorder to identify agents as possible adjunts to existing TB drug regimen.

FUTURE DIRECTION IN TB RESEARCH:New approaches for discovering new, more selec tive anti - TB agent s themapping of M. tuberculosis genome, the delineation of the pathways inMycobacteria call wall biosynthesis (e.g. glycosylation, pathway, fatty acid

biosynthes is, diaminopimelic acid biosynthesi s ) , the discovery of genesinvolved in latency and virulence and the application of DNA micro-arraytechnology to M.tuberculosis.

Application of combination chemistry and high throughout screening to anti-TB drug discovery promises to greatly accelerate the drug discovery process.Training to increase the number and improved delivery methods will be integral

parts of a strategy to full control future outbreaks of TB, particular ly MDR-TB which has severely challenge the limited number of effective treatmentoptions.

To develop and review national TB control proramme plans based on the

DOTS strategy.To build capacity to implement, monitor and evaluate national TB control

programmers.To evaluate cost- effectiveness and impact of TB control interventions.To develop and implement interventions to combat the dual TB and HIV/ADIS epidemic (in close collaboration with ADIS control programmers.

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central book agency (P) Ltd, Calcutta: 17th Ed, 1999: 605-625.3. Ernest Jawetz, Joseph L. mellnick, Edward A. Adelberg, (Eds.),Review ofMedical Microbiology,

Large Medical Publication; Maruzen Company Limited, Japan; 7 th Ed; 1966; 194-202.4 . P. Chakra borty, a Textbook of Microbiology, New central book agency (P) limited Calcutta; 1stEd,

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8. Harsh Mohan, Textbook of pathology; Jaypee Publication, New Delhi India, 4

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Ed, 2002, 139-143.9. Wilson and Gisvolds ,Textbook of organic medicinal and PharmaceuticalChemistry;LippincottWilliams & Wilkins, Baltimore (USA), 11th Ed, 2004, 254-259.

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11. F. S. K. Barar;Essentials of Pharmacotherapeutics; S. Chand & Company Ltd.Ram nagar New Delhi, 3rd Ed, 434-441.

12. Literature Review of Central T.B. Division; Director General of health services; Ministry andfamily welfare.

13 .H. Rang, M. Dale, J. Ritter, P. Moore,Pharmacology,Churchill Livingstone an imprint of Elsevier,5th Ed, 2003, 649-951.

14. K. Park, Parks Preventive & Social Medicine,M/S Banarasides Bhanot Publishers Jabalpure, 15th

Ed, 1998.15. Dr. P.S. Ghadi ,Pathophysiology, Career Publication Nashik 1st Ed, 2000.16. J. Cappuccino, N. Sherman, Microbiology a laboratory manual, Replika press pvt.Ltd.Nerala

Delhi,4th Ed,1999,65-66.17. S.S Kadam, K. R. Mahadik K. G. Bothara,Principles of Medicinal Chemistry, Nirali Publication

Pune, 11th Ed, 1, 2003. 107-19.

Source of support: Nil, Conflict of interest: None Declared


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