diagnosis of difficult multibacillary leprosy cases : lessons from clinical care

Seven multibacillary leprosy and two suspected cases assisted in different situations during clinical care activities at the University in Rio de Janeiro city are described. All cases presented some difficulties for diagnosis, since they evolved with few or no cardinal signs or symptoms of leprosy. A serological test used as an auxiliary tool was helpful in the diagnosis or exclusion procedure of each case, facilitating academic discussions at the time of case examination. considering serology and bacilloscopy (skin smear) as the only rapid and relatively cheap available tests for confirmation of atypical MB leprosy, the advantages and disadvantages of their use were discussed. Both tests support the diagnostic procedure and the classification of cases for treatment purposes. The advantage of bacilloscopy is its capacity for diagnosis confirmation. The advantages of serology are: (a) its applicability for direct use by health workers, providing immediate results; (b) the potential for patient participation in the process; and (c) it provides a learning opportunity, allowing for improved teaching of leprosy pathogenesis. Key-words: PGL-I serology. Leprosy. Multibacillary leprosy. Transmission. Early diagnosis. 1. Departamento de Dermatologia, Faculdade de Medicina, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, Brasil. 2. KIT Biomedical Research, Royal Tropical Institute, Amsterdã, Holanda e Instituto de Patologia Tropical e Saúde Pública, Universidade Federal de Goiás, Goiânia, Goiás, Brasil. Endereço para correspondência: Dra. Maria Leide W. Oliveira. Hospital Universitário. Rua Professor Rodolpho Paulo Rocco 255, 5 andar, sala 05 B 11, cidade Universitária, Ilha do Fundão, 21941-913 Rio de Janeiro, RJ, Brasil. e-mail: mleide@hucff.ufrj.br O coeficiente de detecção da hanseníase no Brasil permaneceu alto e geralmente estável nos últimos anos, sendo foco de vários debates científicos e estudos de pesquisa. Dentre todos os fatores que contribuem para essa realidade, estão o longo período de incubação de Mycobacterium leprae e a existência de uma população extensa de pessoas infectadas pelo bacilo sem tratamento com poliquimioterapia (PQT) . Ou seja, a endemia oculta mantém indivíduos com alta carga bacilar mas sem sintomas clínicos. Vários estudos mostraram que a carga bacilar de um paciente de hanseníase tem correlação com a presença de anticorpos IgM contra o glicolipídeo fenólico I (PGL-I), específico da parede do Mycobacterium leprae: 15-40% dos casos paucibacilares (PB) são soropositivos, comparado com 80-100% dos casos MB. Embora a detecção desses anticorpos não possa ser utilizada para diagnóstico, é uma ferramenta útil para a confirmação de casos multibacilares. A sorologia é altamente sensível na detecção da presença de anticorpos contra Mycobacterium leprae na corrente sanguínea, mais do que a microscopia de lesões localizadas em alguma parte específica do corpo como especificado no exame de bacilos álcool-ácido resistentes (BAAR). A vantagem da baciloscopia é sua capacidade para confirmar o diagnóstico. No entanto, há várias limitações operacionais: é um exame invasivo composto por várias etapas, o que aumenta a possibilidade de erro, e leva tempo para concluir. A biopsia de pele é realizada em muitos casos e sua confiabilidade e as informações histopatológicas resultantes muitas vezes ajudam na classificação das formas da doença. Isso provavelmente contribui para uma demanda excessiva do exame observada no Brasil, mas raramente está disponível nas unidades básicas de saúde (UBS).

The case detection rate of leprosy in Brazil has remained high and generally stable in recent years and has been the source of scientific debates and research studies 1 .Among all the factors that contribute to this, the long incubation period of Mycobacterium leprae and the existence of a large pool of unidentified infected individuals not on multidrug therapy (MDT) are two important ones 2 .Another possibility is the silent transmission by individuals with asymptomatic disease or infection 3 who present a high bacterial load.
Several studies have shown that the presence of IgM antibodies to the Mycobacterium leprae-specific phenolic glycolipid-I (PGL-I) correlates with the bacterial load of a leprosy patient: 15-40% of paucibacillary (PB) patients are seropositive, compared to 80-100% of multibacillary (MB) patients.While detection of these antibodies cannot be used for diagnosis, they are a useful tool for confirming the diagnosis of MB disease.
Serology is highly sensitive for detecting the presence of antibodies to Mycobacterium leprae in the bloodstream, rather than in the lesions from a specific part of the body, as determined by acid fast bacilli (AFB) in slit skin smears.The advantage of bacilloscopy is its capacity to confirm diagnosis.However, the operational drawbacks are that it is a time-consuming, invasive exam composed of several steps that increase the possibility of error.Skin biopsy is performed in many cases and its reliability and the resulting histopathological information often help classify the form of the disease.This probably contributes to the excessive demand of this exam observed in Brazil 4 , but it is rarely available in remote areas.

FIgURE 2
Wade 1000x: No acid fast bacilli in the perianexial infiltrate.
In our University Hospital (UH), the leprosy team assists out-patients referred by basic health units (BHU), as well as inpatients in dermatology, rheumatology, infectious diseases and general medicine.Additionally, university extension activities are promoted, including active case finding campaigns.They involve local health workers (HW) and the interaction of dermatology residents in communities where leprosy is prevalent.All of the cases presented were diagnosed at the UH and complementary exams were performed at its laboratory.
During these activities a serological test was used, the ML Flow test, which detects IgM antibodies to PGL-I.The test is stable at 28 o C for 3 years and can be performed by health workers at any level.The test was performed using 5µl of whole blood obtained by finger pricking and results were read after 5 minutes.The color intensity of the test line can be read as negative, or ranging from 1+ to 4+ 5 .The fact that each step of the test can be followed by the patient and trainees benefits the teaching regarding the pathogenesis of the disease and provides an opportunity for learning.
Through the use of case report lessons 6 , our group aimed to (a) highlight the potential benefit of the ML Flow test as an auxiliary tool in diagnosing difficult MB leprosy cases (lepromatous (LL) and borderlinelepromatous (BL), and (b) demonstrate that a problem still exists concerning MB leprosy diagnosis when clinical signs and symptoms are absent or are not easily recognized in general health services.

CASES REPORTS And dISCUSSIOn
The detection of AFB in skin smears confirms active disease, while PGL-I serological tests detect antibodies against Mycobacterium leprae; this indicates infection that might never evolve to disease.Nevertheless, serology is useful for classifying patients as PB or MB in field conditions 7 and as a tool to assist in diagnosis, as illustrated in the following cases.The test was used for (1) helping identify differential diagnoses in cases 1, 2 and 3; (2) reinforcing clinical suspicion of MB asymptomatic cases in cases 6, 8, and 9, and (3) confirming the suspicion of active leprosy in cases 4, 5 and 7.The objective was to discuss the cases and report how the test was used.Table 1 summarizes the patients' histories and only relevant aspects of clinical features are described.
Case 1.The patient was referred to the UH to investigate recurrent erythema nodosum on the trunk and limbs lasting for 12 years.At the time of examination, he presented only erythematous and painful nodules spread over the entire body (Figure 1).The ML Flow test was performed and its negative result triggered further investigation for other infectious diseases as a possible explanation for the clinical symptoms.The patient was positive for the hepatitis C virus (HCV), which is also a cause of erythema nodosum.The histopathological exam showed no features of leprosy (Figure 2); therefore, leprosy was excluded.
Case 2. Admitted to the infectious disease department of the UH, this patient presented with fever and enlargement of the lymph nodes (cervical/inguinal), liver and spleen.AIDS was suspected and tuberculosis (TB) was diagnosed due to an AFBpositive sputum result.Soon thereafter, anti-TB treatment was initiated (rifampicin, isoniazid and pyrazinamide) but his symptoms worsened with a reactional episode, despite the fact that all three sputum cultures were negative for Mycobacterium Tuberculosis.
Therefore, confusion between the two diagnoses occurred due to the possibility of Mycobacterium leprae present in the upper respiratory tract.The positive ML Flow test performed during the dermatological consultation induced further examination of material from lymph node biopsy and slit skin smears.The skin infiltration observed in the face was doubtful and fibular nerve was painful upon palpation.No other signs or symptoms were observed.The biopsy was stained according to Wade and LL was confirmed (Figure 3).Multidrug therapy (MB) was initiated, combined with 200mg thalidomide for two months.After the regular 12 doses of MDT the patient was released from treatment.Leprosy and tuberculosis are both endemic in Brazil and, in most cases, show distinct clinical courses; however, differential diagnosis between these diseases can be challenging, especially in early lepromatous leprosy cases.
Revista da Sociedade Brasileira de Medicina Tropical 41(Suplemento II):27-33, 2008 Case 3. The patient was initially diagnosed with a drug allergy and treated with corticosteroids.After three episodes, the patient was sent to the skin disease out-patient clinic of the UH.He reported substantial weight loss (8kg), 3 flu-like episodes (nasal obstruction) and malaise in the preceding 6 months.Only a few red nodules on the legs were observed.Biopsy of a nodule was performed under suspicion of nodular vasculitis, tuberculosis, and/or drug allergy.In addition, a HIV test was performed, which was negative.After 20 days the patient returned and no longer showed any lesions or symptoms.Unexpectedly, the result of histopathology was erythema nodosum leprosum (ENL).The ML Flow was positive and MDT/MB treatment initiated.
Erythema nodosum is a syndromic event with many causal possibilities, among these is MB leprosy.This case is an example of the immunopathology of silent LL, showing that the type-2 reaction was triggered by viral infection.The biopsy was performed due to suspicion of adverse drug effects and leprosy was not considered.In a leprosy-endemic area, a rapid test used for screening could be used to test patients presenting recurrent erythema nodosum.In positive cases, bacilloscopy could be performed, thus avoiding biopsy.The patient was treated with MDT MB in the original health facility.
As an example, in Case 4, a positive ML Flow test reinforced the clinical suspicion of leprosy and the biopsy was avoided.The patient was referred to the UH due to thickened skin in the shoulder and right arm.The only symptom was paresthesia on an unspecific skin lesion (Figure 4), which was not easily recognized by inexperienced field workers.The clinical suspicion was BL.A skin smear examination was performed, the case was confirmed and treated with MDT MB in the health facility.
The following two cases are probably part of the silent MB cases mentioned in the literature as the main source of leprosy transmission 6 .Since no other family members presented MB leprosy, cases 5 and 6 probably acted as transmission agents for PB secondary cases.
Case 5.The patient was treated and cured of leprosy when he was 12 years old with the MB scheme used at that time (90 days rifampicin plus dapsone daily for two years).Two years before the present admission, his daughter was diagnosed with tuberculoid leprosy.Only a few months later, he observed an enlarged plaque on his abdomen (Figure 5).Skin smears performed in a basic health facility were negative.The biopsy at that time was inconclusive, but compatible with indeterminate leprosy.Referred to the out-patient clinic of the UH, evolution and minor infiltration of the initial plaque was observed and a new one appeared.Cotton wool, pinprick and monofilaments tests showed loss of sensation in the center of the lesion.A positive ML Flow test, clinical aspects and medical history reinforced the diagnosis of a MB relapse and MDT/MB was initiated immediately.The slit skin smears were reexamined and AFB were detected.The histopathological results confirmed an active BL leprosy case.The positive ML Flow test was helpful in diagnosing a possible reinfection/ relapse case.The

FIgURE 6
Wade40x -Perianexial granulomatous infiltrate.12 doses MDT/MB were completed and the patient is currently undergoing corticoid therapy for leprosy reactions.
Case 6.The patient was examined because he was a household contact of 2 paucibacillary (PB) leprosy children diagnosed during a leprosy elimination campaign in a peripheral area of RJ City.Neither leprosy skin lesions nor nerve enlargement were detected.He presented acrocyanosis and discrete erythema on the upper legs that could have been related to his cigarette addiction; however, since he was a candidate as the index case of the two PB cases, MB leprosy was also a feasible diagnosis.Therefore, the ML flow test was performed and the positive result justified additional investigation.Bacilloscopy and biopsy showed periaxial granulomatous infiltration (Figure 6) with AFB in the Wade stain confirming BL.The test results changed the patient's attitude, because he initially refused to go to the health facility for examination and the team had to go to his residence.Following the test result, he went willingly for further testing.This exemplifies that the use of the ML Flow test under field conditions reinforced clinical suspicion of silent MB leprosy and encouraged both: (i) the field health worker to refer the case for further investigation and (ii) the patient to go to the reference unit.

Case 7.
A woman brought her daughter to the UH, because leprosy was suspected at the local level, and a LL case was confirmed.Therefore, the woman was examined as a household contact and leg ulcers were observed (Figure 7A).She reported current treatment for varicose ulcers for almost a year.She also reported recurrent ulcers and painful nodules suggestive of erythema nodosum in both legs during the preceding 16 years, especially during her four pregnancies (Figure 7B).The question of active or inactive disease was raised and the strong positive serology result demanded for further investigation.Bacilloscopy and biopsy confirmed the LL case, possibly spreading AFB for 16 years.The patient had discrete loss of sensation in both plantar regions (tibialis posterior nerve region).Despite the residual aspects of her skin lesions and the presence of madarosis and a nasal ulcer without correspondent facial skin infiltration, she reported she had never been diagnosed or treated for leprosy.This was confirmed by an investigation of the leprosy case notification system of RJ.The ML Flow test was strongly positive and she was diagnosed with active LL and treated with MDT MB.The diagnosis was confirmed by histopathology (Figure 8A  and B).Her 18 year-old son had previously been examined and diagnosed as an LL case.Another son was examined as her contact and the case is presented below.Thus, she was the index case of three children with MB leprosy.Case 8.The 11 years old son of case 7 also exemplified silent MB leprosy and the importance of investigating nasal symptoms and acrocyanosis as indicative of LL leprosy.He presented nasal obstruction, moderate acrocyanosis, livedo reticularis in both legs (Figure 9) and possible skin infiltration of the earlobes and face, but no typical leprosy lesions.The ML Flow test was strongly positive and active LL was confirmed by skin smears and histopathology.The patient presented recurrent reactions that were difficult to control.
Case 9. A leprosy case?This case was a household contact of three LL cases diagnosed in 2003/04.In 2003 she was examined and was negative in both dermatological/neurological aspects and serological testing.In 2004 she presented with acrocyanosis, livedo reticularis and one erythematous plaque on her left thigh with loss of sensation.The ML Flow test was performed and showed a reversal to a positive result (4+).LL was suspected of subclinical infection could result in leprosy disease.The decision to treat for at least 6 months was based on the fact that disease could manifest itself and it probably would be a MB case 3 , thus the early treatment would benefit the patient.Perhaps, if no treatment had been provided directly after seroconversion, during follow-up of this case, the appearance of signs and symptoms of leprosy would have been observed.Nevertheless, this case illustrates how careful health professionals should be when interpreting the ML Flow test results.

COnClUSIOnS
One criticism to learning practices in medical training is the excess of complementary exams to confirm clinical suspicions of leprosy, even when patients present clear cardinal signs and symptoms of leprosy 4 .This could negatively influence the practices of new doctors at basic health care levels, where they must diagnose leprosy using minimal technology.In fact, 70% of

TABlE 1
Summary of the clinical symptoms and tests results of the cases.