Tuberculose multirresistente no Brasil : histórico e medidas de controle Multiresistant tuberculosis in Brazil : history and control

The article aimed at assessing multidrug-resistant tuberculosis control in Brazil, based on the experiences of reference institutions, and the most relevant studies carried out to determine local and national resistance rates. Control measures and the current situation of treatment and diagnoses after the implementation of the national guidelines, which were revised in 2004, are considered. The fi rst national survey on resistance to anti-tuberculosis drugs was performed in the middle of last decade. From its outcomes, a regimen to treat all cases of multidrug-resistant tuberculosis was validated and adopted. Government measures enabled the implementation of a surveillance system, whose outcomes are also commented.


INTRODUCTION
The last decades were marked by huge social differences in developing countries such as Brazil.These inequalities are translated into an increase in poverty, lack of access to services, population growth, and urban concentration.This has a negative repercussion in endemic diseases, such as tuberculosis (TB), and the pandemic of infection by Aids virus.In this scenery, worsening of TB control programs resulted in great challenges to fi ght infectious diseases.They are: expressive increase in TB treatment drug resistance, emergence of multidrug resistant forms (MDRTB), and more recently, in 2006, extensively drug resistant tuberculosis (XDRTB). 6,16,23 Brazil, several facts followed the development of the I Inquérito Nacional de Resistência (1st National Survey on Resistance), whose outcomes were published in 1998, 2 they were: creating a protocol to validate standard therapeutical regimen to treat MDRTB cases; 9 recommending the performance of culture and sensitivity test in all cases of disease retreatment, according to the national guidelines for TB, designing several research works together with national institutions; implementing a system of epidemiologic surveillance of resistant cases, which implies reporting all cases and follow them up during and after treatment.
The object of the present article was to assess multiresistant TB control in Brazil, based on the experiences of reference institutions and on the main studies performed to determine local and national resistance rates.

BACKGROUND
Clinical form of TB resistant to rifampicin and to isoniazid was conceptually called multidrug-resistant tuberculosis in the United States in the end of the 1980's.Between 1982 and 1986, there was an expressive increase in the multiresistant forms, leading to recognition of the problem as a public health one, and revealing the impact of AIDS epidemics and the disorganization of health services including those of New York.In the beginning of the 1990's, outbreaks of nosocomial transmission of multidrug resistant TB were reported by New York hospitals, they were all characterized by late diagnoses, unsuitable therapy, high mortality, and high transmission rate, especially because patients were in hospital and were immunocompromised due to Aids. 3,15Other authors have already reported similar experiences in patients with AIDS severely immunocompromised, leading to mortality rates up to 60% in a 16-week period.
In Brazil, there is a standard regimen for retreatment cases whose fi rst treatment was unsuccessful, which, until 2004, it was recommended to apply a combination of rifampicin, isoniazid, and pyrazinamide, regardless of the performance of sensitivity test to drugs.*We have considered as multiresistant for reporting and treatment purposes, those cases that present in vitro resistant to rifampicin and to isoniazid, and to a third drug on the standard treatments.
Case concept of MDRTB in Brazil is the following: • Confi rmed -TB case with diagnoses confi rmed by culture and identifi cation of Mycobacterium tuberculosis and by sensitivity test, with rifampicin + isoniazid resistance and to another drug; • Probable -failure to retreatment regimen (phase III) with sensitivity to rifampicin or isoniazid.

Bases of multiple chemotherapy
Together with the introduction of TB chemotherapy, is a progressive knowledge on the pharmacodynamics of its etiological agent, M. tuberculosis.Among pathogenic mycobacterias, that from TB is a severe challenge to public health, leading to complex articulation between early diagnoses and effi cient chemotherapy to stop its transmission chain.Thus, chemotherapy is the most powerful sanitary resource to treat TB, because it is the only thing capable of directly interfering in smear positive cases, the "main bacilli reservoir" defi ned by Canetti, in 1959. 3,11olated use of only one medication shows the high proportion of resistant mutants and explains the phe-nomena of primary, natural, and acquired resistance, and the consequent need for drug association in the regimen.Based on in vitro and in vivo models, currently the singularity of differentiated multiplication of M. tuberculosis is well understood, according to the oxygen supply.That is, the different growth speed in intra and extra cellular environments, in closed caseous lesions, and in the wall of cavitary lesions.This differentiates populations that present rapidly growing (they are more sensitive to medication) from persistent populations (slow or intermittent multiplication), that require prolonged treatment for elimination.These are the bases for regimens to neutralize naturally resistant bacilli and for the long treatment period to eliminate persistent bacilli present in resistant forms. 3,8,11,26servation of the direct proportion of persistent bacilli populations and the disease morbidity with the total bacilli population led to the development of a twophase treatment.This presents a phase called intensive phase, aiming at quick reduction of bacterial load, and the continuation phase aiming at preventing disease reactivation or relapse, by sterilizing resistant ones.This principle applies especially to susceptible forms of TB, however, in a rational fashion.It also applies to resistant forms, despite prolonged treatment, as observed in studies published for MDRTB treatment where there is the presence of a maximum number of associated drugs in the fi rst 12 months of treatment. 10,20 ional use of anti-TB drugs in designing regimens is the following: 3 • Group 1 -fi rst line drugs, oral: isoniazid, rifampicin, ethambutol, pyrazinamide.
It has been known for a while that previous contact with drugs is the greatest clinical indicator of development of multi-drug resistance. 10,11,19Thus, multiresistant is considered a biological iatrogenic phenomena, due to the inadequate use of short-course regimens -especially those made by the combination of rifampicin, isoniazid, pyrazinamide and ethambutol.The strategy of prevention to multiresistance is the correct application of short-course fi rst-line treatment, using the components of Directly Observed Treatment Short course (DOTS) strategy, or directly observed treatment.
Launched by the World Health Organization in the middle of the 1990's, DOTS strategy has the following measures: a) political commitment and fi nancial support by countries and regions; b) suitable and regular supply of medications, with quality control of drugs; c) early detection of cases with good quality bacteriology; d) standardized treatment categories applied under direct observation of the patients, aiming at regular intake of medications; and e) monitor and assessment of the information system and the impact of the measures adopted. 26Up to the beginning of that decade, in different introduction levels, the DOTS had been adopted in 155 countries, including Brazil.The Ministry of Health determined that these measures would be adopted as a priority over fi ve years ago.Currently in Brazil there are acknowledged effective experiences in implementation with encouraging outcomes, in the Mid-West region and in Parana, in areas of Rio de Janeiro, such as the community of Rocinha, and other areas in São Paulo.
In 2006, the offi cial mission of the Stop TB partnership came to Brazil bringing new contributions to the goals of TB control, such as: increase the quality of TS/ DOTS; reinforce the combat to TB/HIV co-infection, MDRTB and other vulnerable groups; strengthen the health system of the country; foster greater involvement of health professionals, empowering the civil society; and develop operational research.In this sense, creation of a Forum Stop TB Partnership by the Ministry of Health was a milestone for TB control in the country, once it offi cially recognized the important role of social mobilization.This decision and the political commitment were determinant for the country to be able to, in 2006, receive a US$ 27 million investment of the Global Fund to fi ght Aids, Tuberculosis and Malaria for a fi ve-year-period.

Complexity of multiresistant tuberculosis
Mycobacterium tuberculosis is a slow-growing aerobe pathogen with a high proportion of resistant mutants. 19- 21Of the four known mechanisms leading to bacterial resistance (conjugation, transformation, transduction, and mutation), M. tuberculosis acquires resistance to drugs only through mutation.Resistance to isoniazid is due to a mutation frequently observed in katG gene and to rifampicin, in the rpoB gene.Recent studies using molecular biology techniques have been relevant to understand the transmission mechanisms and the virulence of the bacilli, and also to open new ways for diagnoses and therapy. 8,24,25,27 Irazil, despite the clear worsening of several local programs to control TB, one of the reasons that resistance cases could be prevented was the regular use of isoniazid associated with rifampicin in the same tablet, since the introduction of short course regimen. 14,17pes of resistance to M. tuberculosis may be summarized in: • Natural -due to spontaneous mutation, regardless of previous exposure to drugs, and directly proportional to the number of bacilli.
• Initial -observed when patients start treatment, with resistance to one or more drugs.It includes patients with primary or acquired resistance, with no known history of prior treatments.
• Primary -observed in patients who have not been previously treated, infected by a resistant form.
• Acquired or secondary -resulting from the previous inadequate use of medication. 20oss-resistance and adherence are known as major problems in TB control, especially regarding managing retreatment. 19,20Cross-resistance is a well known phenomena well described in the literature, occurring among the most important drugs used in treatment.Among ansamycin drugs, a TB case resultant to rifampicin presents a higher than 80% chance to be also resistant to rifabutin.A similar phenomenon is expected to rifapentine, an ansamycin derivative recently tested for sensitive TB cases.As its half life is fi ve times higher than that of rifampicin, rifapentine presented promising results regarding the possibility of intermittent use in treatment regimen, and up to once a week in chemoprophylaxis regimens.Cross resistance may also occur among aminoglycosides, in the sequence: streptomycin, kanamycin, amikacin, and the polypeptide capreomycin; among quinolone derivatives, as recently confi rmed by pharmacokinetic studies; and among isoniazid and ethambutol, with a current question on the common role of the presence of the katG gene in both. 8,27ere is great diffi culty to comparatively assess studies on MDRTB published in the literature according to the criteria based on evidences, either because of methodological limitations, differences in experiences reported, or regimens used.These studies do not contemplate explanatory or pragmatic clinical essays because of ethical aspects, being limited to describing observed outcomes and the adverse effects of treatment.For these reasons, the "best evidences" are those from the action of each drug available, the drug association used, and the treatment regimen in the different public experiences. 3,7,9,22veral authors suggest that for the effective treatment of MDRTB the association of at least four medications in several Latin American countries because they act in bacillary population with intermittent multiplication and where almost all patients are treated with rifampicin monotherapy.Besides, combined administration of medications in sensitive patients using fi xed dose combination (FDC) or combined therapy, ensures the best serum peaks and facilitates surveillance.In resistant patients, it is important to observe that all medications are taken at the same time.

Background of resistance control in Brazil
Historical experiences were reported in the Brazilian literature in previous decades by authors such as Poppe de Figueiredo, Jayme dos Santos Neves, Hélio Fraga, Germano Gerhardt, José Rosemberg and others, showing the national concern with resistance.In the years 1958-59, 66.6% of patients in treatment in the former state of Guanabara (current city of Rio de Janeiro) had become chronic resistant to two or three drugs in the standard regimen.Several corrective actions have been introduced by the then called Serviço Nacional de Tuberculose (National Tuberculosis Service), for the whole country as of 1960: a regimen was standardized, made of streptomycin, para-aminosalicylic acid and isoniazid for new cases; regimens with "reserve drugs" for surgical cases; changes on admission criteria of patients, among other measures.In Rio Grande do Sul, for example, outcomes were modest, since in 1966, cure rate was still low (36.9%),although it had increase three fold. 17,22 study that compared the evolvement of resistance rates at Instituto Clemente Ferreira, in São Paulo, showed a decrease from the 1960's to the 1980's, and less marked from the 1970's to the 1980's (9.6% to 7.4%). 13This decrease is likely to have occurred due to a better organization of the TB program in the 1970's and to the introduction of more powerful drugs in the treatment, particularly to the combined administration of rifampicin and isoniazid, in the 1980's. 14 epidemiological surveillance system of MDRTB is currently running, and it should evolve to monitor all retreatment cases nationwide.The most relevant facts with proved in vitro sensitivity is necessary.Among these, two may not have been used in the past and one must be injectable, such as aminoglycosides or polypeptides; together with oral quinolone, for at least 24 months, or at least 18 months after negative culture. 3,14,20Applying this methodological recommendation to the methodology of published studies, a direct observation of the number of drugs not previously used with the fi nding of best results.

Diagnoses of multiresistant tuberculosis
In almost all published studies, the criteria used to defi ne a case as multiresistant were the sensitivity test. 7,9,12,15Prevailing method is that of proportions, in solid Lowenstein Jensen medium, or in liquid medium, such as Bactec 460.MGIT 960 system or MB/Bact system are considered fast because they enable pathogen identifi cation in 15 days on average. 5e method of proportions, originally described by Canetti et al 4 in 1963, consists on putting the same amount of dilutions with approximated measure of the inoculum in control and in the medium with the drug tested.Resistance or sensitivity is estimated by comparing the number of colonies growing in the medium with the drug, and the number growing in the control medium expressed in percentage.The defi ning cut-off point considered is 1%. 5 Despite the technological progress reached, diagnoses of MDRTB still takes long, due to the time required by the methods of diagnoses, even in liquid mediums of culture, and by limitations and inconsistencies in sensitivity tests.Among the drugs regularly tested, the only ones which present proved 100% confi dence are rifampicin and isoniazide. 3A study recently published by Mitchison 20 reports the comparison among 16 reference laboratories in mycobacteriology worldwide and the differences in the outcomes found regarding resistance to fi rst and second line drugs.This author criticizes the confi dence values of sensitivity tests and suggests the defi nition of minimal individual inhibiting concentrations for each drug as a priority, through laboratory validation. 20erefore, to decide what regimen to be used in patients already treated for TB, it should be considered the therapy history of these patients together with the result of the sensitivity test. 22 addition to the important components to treat TB and MDRTB already mentioned, it is important to take into account the concept of early bacterial activity, to which isoniazid is the best medication.Rates of isoniazid resistance must be considered and they are expressive from the last ten years to the present are summarized as follows.
• 1995 -local experiences of institutions were acknowledged by the Coordenação Nacional de Pneumologia Sanitária (National Coordination of Sanitary Pneumology) of the Ministry of Health, such as: Hospital Sanatório Partenon (Porto Alegre), Instituto Clemente Ferreira (São Paulo), Hospital Raphael de Paula Souza (Rio de Janeiro). 5,9In the same year, the outpatient facility of the Centro de Referência Hélio Fraga together with Hospital Raphael de Paula Souza was created, at fi rst, as a state reference.
• 1995 to 1998 -the Multicentric Protocol was created to treat MDRTB cases, aiming at validating a standard regimen. 9 • 1996 and 1997 -the 1 st National Survey on Drug Resistance was carried out in Brazil with the fi rst outcomes published in 1999. 2 In the same year, all drugs were recorded in the Agência Nacional de Vigilância Sanitária (National Health Surveillance Agency -Anvisa), and the mechanisms for systematic purchase of the regimen were established.State laboratories started producing most drugs used, and only terizidone was bought in the international market.
• 2000 -the Ministry of Health considered the regimen made by the association of ofl oxacin, amikacin or streptomycin, ethambutol, terizidone and clofazimine as valid, starting epidemiological surveillance with reporting of cases undergoing treatment" in that year; at the same time, a database was set up for Epidemiological Surveillance of MDRTB.
• 2004 -the introduction of an epidemiological surveillance system of MDRTB was continued because of an agreement of technical cooperation between the Ministry of Health, Centro de Referência Hélio Fraga, and the Project Management Sciences of Health.In 2005 a control study of all fi rst and second line drugs used in TB control was published,* including MDRTB, presenting satisfactory outcomes regarding effi ciency.
In 2006, the 2 nd National Survey on TB drugs resistance started.The representative sample for the country should reveal the current resistance pattern of fi rst and second line drugs in outpatient and hospital cases.**

Current Situation of resistance in Brazil
A study performed in Brazil between 1986 and 1989on primary resistance to drugs, demonstrated resistance percentages to isoniazid from 6.1 to 6.8%, and to rifampicin from 0.4 to 0.6%; resistance to two or more drugs reached almost 3%.Total primary resistance, considering all patients resistant to at least one medi- cation, was 15.2% in 1986, and 11.5% in 1989.These data were similar to those obtained in the studies of difference series, 13,14 in the 1970's.A greater rate of primary resistance was observed in the north more than in the south and southwest region with high primary resistance rate to streptomycin (7.2%).This probably indicates infection previous to the 1980's, therefore, they were cases of endogenous reinfection, and the use of streptomycin in the treatment of new TB cases is not recommended, apart from special situations. 14 the World Survey on Resistance, coordinated by the World Health Organization and the International Union Against Tuberculosis, between 1995 and 1996, Brazil took part in the sample with 2,888 strains of patients seen in outpatient facilities, treated (N=793) and not previously treated (N=2.095).2,26 Difference between the global rate of multiresistance was observed (rifampicin + isoniazid and/or other drugs) in patients not previously treated (0.9%) and already treated (5.4%), reinforcing the importance of the previous use of medication as a clinical indicator of resistance. 2

PATTERN OF RESISTANCE AND DRUG PERSPECTIVES
The 1 st National Survey on Resistance 2 (1996 to 1997) had a representative sample for the country: totaling 6,000 outpatient cases among never treated and retreated.Table 1 shows the pattern of resistance found.
Since 1945, with the development of pyrazinamide and later streptomycin, few drugs have been produced for TB treatment, especially when compared to other diseases. 26Table 2 summarizes all drugs produced up to the present.

FINAL CONSIDERATIONS
Avoiding resistance is known as the most important premise of tuberculosis treatment. 8,19,21Add to that the complexity of the disease per se, the diffi culty in handling patient with long treatment course, the lower effi ciency compared to conventional treatments, and the high costs of drugs and the propaedeutics necessary to follow-up.Additionally, there is the human suffering due to the severity of the consequences of this form of TB, that many times takes the patients away from the job market due to disability, in their most productive years. 1,18In this aspect, it is important to highlight that among TB cases reported to the Epidemiology Surveillance System of the Ministry of Health, the predominant age group (50.1% of cases) is between 20 to 40 years old, and that the frequency of HIV coinfection is lower than the national average among TB cases (Table 4).
In a prospective outlook for MDR TB the fi elds of prevention, diagnoses and treatment are considered as priorities for investments and research.
For transmission control, investments on introducing DOTS strategy are priority, together with the greater effi ciency of short course treatment, preventing the appearance of resistant forms; and the increased use of molecular epidemiology in describing micro epidemics and in detecting clusters.
The strategy for diagnoses would be: greater confi dence in sensitivity tests available, with strict quality control of the outcomes supplied; validation of new and fast methods; studies with molecular and immunologic markers; standardization of individual minimal inhibiting concentrations of drugs to determine resistance; studies to validate clinical markers of resistance based on therapy history.
For treatment the priorities would be: validation of sensitivity tests for newer drugs and monitoring of minimal inhibiting concentrations for each of them, aiming at increasing the effi ciency of regimens, and discovering drugs that can lead to new regimens.In this sense, the current period is extremely relevant, with studies on 7 drugs being conducted, from different pharmacological groups and in different phases, pre and post clinical; this had not occurred since the discovery of rifampicin, almost 40 years ago. 1,18According to Global Alliance for TB Drug Development, for the development of anti-TB drugs, the following are under study: one dyarilquinoline, two nitroimidazoles analogs and derivatives, together with synthase and translocase enzyme inhibitors.These studies also aim at creating treatment regimens with lower doses and shorter courses.
Another challenge is to reduce market prices, particularly of second and third line drugs.This may be reached through the actions of supranational institutions such as: Global Fund Against Aids, Tuberculosis, and Malaria, Global Alliance for TB Drug Development, Green Light Committee.
And, fi nally, for control, the priority will be to test and validate drugs following effi ciency and plausibility criteria.
The same phenomenon was seen in several Brazilian capitals, revealing the problem in the country.In Rio Grande do Sul, in 1960, cure

Table 1 .
Pattern of resistance found in the 1st National Survey on Resistance.³Brazil, 1996-1997.

Table 2 .
Drugs groups and medication produced since 1945, available for TB treatment.

Table 3 .
Groups and drugs in study and drug perspective for tuberculosis.

Table 4 .
Multidrug resistant tuberculosis cases reported to the Epidemiological National Surveillance of the Ministry of Health and frequency of HIV co-infection, between 1995 and 2007.

Table 3
summarizes the main substances and drug groups currently under study and the research lines for TB treatment.