Serological diagnosis of tuberculosis.

OBJECTIVE: To evaluate the efficacy of ELISA for the detection of IgG antibodies against antigen 85 complex (Ag 85 complex) of Mycobacterium tuberculosis. METHODS: Children of either sex, 0-18 years of age, attending the outpatient department and admitted in the casualty and wards of the Department of Pediatrics, S.N. Medical College, Agra, were included in present study. The study was carried out on children with pulmonary and CNS tuberculosis along with matching controls (83 cases and 32 controls). Informed consents of their parents or guardians were taken. They were subjected to clinical examination, relevant laboratory investigations, tuberculin test and chest radiograph. Relevant body fluids were subjected to bacteriological tests; ELISA was applied to serum samples for detection of IgG antibodies against antigen 85 complex (Ag85). The result of ELISA was compared with bacteriological tests [Ziehl Neelson (ZN) staining for acid-fast bacilli, culture on Lowenstein Jensen (LJ) medium and culture on BacT/Alert 3D system]. RESULTS: ELISA tests showed a significantly higher sensitivity (59.1%) as compared with LJ medium culture method (19.3%), BacT/Alert 3D system (24.1%) and ZN staining (16.9%) in all patients (p<0.001). Specificity of ELISA test was 71.9%. CONCLUSION: In view of the convenience, low cost and good sensitivity, ELISA tests have a promising future in the diagnosis of childhood tuberculosis.

PCR and in-situ hybridization for diagnosis of leprosy.

OBJECTIVE: To assess the diagnostic value of Polymerase Chain Reaction (PCR) and in situ hybridization. METHODS: This prospective study was carried out in 22 patients RESULTS: The histopathological examination confirmed the clinical diagnosis in 27.2% cases only. In situ hybridization showed a positivity of 42.8% in early (I/BT) and 46.7% in BB/BL group. In situ hybridization thus enhanced the diagnosis by 18.1%. PCR targeting 36 kDa gene of M. leprae was performed on 15 cases. In these 15 cases, histopathology confirmed the diagnosis in 4 cases (26.6%) and PCR confirmed the diagnosis in 10 cases (66.6%), thus enhancing the diagnosis by 40%. CONCLUSION: 36 kDa PCR and in situ hybridization enhance the diagnosis of leprosy when compared to routine histopathology. They are important diagnostic tools for definitive diagnosis in early and doubtful cases of leprosy.

Diagnostic value of in situ Polymerase Chain Reaction in leprosy.

OBJECTIVE: This prospective study was carried out to assess the diagnostic value of in situ Polymerase Chain Reaction in leprosy, particularly in enhancing the histopathological diagnosis. METHOD: Clinical examination of 20 patients (< 16 yr) was done and skin smear for AFB was prepared. Biopsy of lesion site was taken for histopathological examination and in situ PCR testing. RESULTS: The histopathological examination confirmed the clinical diagnosis in 45% cases only; non-specific histopathology was reported in the remaining 55% cases. In situ PCR showed a positivity of 57.1% in early/localized form of leprosy (IIBT) and 61.5% in (BB/BL) group. When compared to histopathology examination, a significant enhancement of 15% in diagnosis was seen. With in situ PCR, the diagnosis could be confirmed in 4/11 (36.3%) cases with non-specific histopathological features, (which is common in early disease) in addition to confirmation of 8/9 (88.8%) histopathologically-confirmed tissue sections. Histopathology and in situ PCR, combined together, confirmed the diagnosis in 13/20 cases (65% of total cases). CONCLUSION: Thus, in situ PCR is an important diagnostic tool especially in early and doubtful cases of leprosy.

Study of gene probes in childhood leprosy.

The objective was to detect nucleic acids of M. leprae in skin lesions of leprosy patients and study the effect of treatment on these nucleic acids, using r-RNA gene probes, using a cross sectional study. The study was carried out at Department of Paediatrics, S.N. Medical College, Agra and Department of Microbiology, Central JALMA Institute for leprosy, Agra. The study included 32 cases of leprosy less than 16 years of age, divided into 3 groups viz. without treatment (12 cases), in middle of treatment (11 cases) and at the end of treatment (9 cases). All cases were subjected to a detailed history and thorough clinical examination. All of them had smear examination and lepromin test done and their skin biopsies were subjected to gene detection. Nucleic acids were isolated from skin biopsies of all cases by standard procedure. After dot blotting of these nucleic acids, they were hybridised with radioactive (p32) r-RNA probes. The results were interpreted after getting the X-ray films processed with background signals from controls. Majority of cases were between 13-16 years of age. As age advanced, the disease moved from tuberculoid end of spectrum towards lepromatous end (p < 0.05). Majority of paucibacillary (PB) cases were lepromin positive while majority of multibacillary (MB) cases were lepromin negative (p < 0.05). In specimens of untreated cases, 50% of PB specimens and 87.5% of MB specimens were positive for r-RNA probes. In multibacillary type 100% smear positive specimens and 67% smear negative specimens were positive for r-RNA probes. In patients during the middle of treatment positivity for r-RNA decreased and 20% of PB specimens and 16.6% MB specimens were positive. At the end of treatment (1 year for PB cases and 2 years for MB cases), the results of r-RNA were negative, which indicated that the treatment regimens used in the study were effective. This study supports the usefulness of r-RNA probes as a diagnostic and therapeutic tool in childhood leprosy.

Diagnostic value of gene probes and its correlation with clinical profile of leprosy in children.

Clinico-bacteriological profile of 73 leprosy patients below 16 years of age was studied. Majority of the patients were males and fell in 11-16 years age group (p < 0.05). Skin lesions were present in all cases on both exposed as well as unexposed areas and their number increased with advancing age. Cutaneous sensations were affected in most of the patients while nerve thickening was observed in 41. As age increased, the disease moved from the tuberculoid end of spectrum towards the lepromatous end (p < 0.05) and the positivity of the skin smears increased (p < 0.05). Majority of the paucibacillary cases were lepromin positive while most multibacillary cases were lepromin negative (p < 0.01). Two M. leprae specific gene probes were applied in 42 cases to assess their diagnostic value. Eighty one per cent cases were picked up by the probes indicating presence of active bacilli. These included all lepromin positive cases, all smear positive cases, and most of smear negative cases (p < 0.05). Seven children with inconclusive histology were also positive. Drug treatment and inadequate size of biopsy sample could explain the negative probe results in 19% cases. This study highlights the immense potential of gene probes in diagnosing leprosy in children.

A clinico-hematological profile of hemolytic-uremic syndrome.

Hemolytic uremic syndrome (HUS) is defined as microangiopathic hemolytic anemia, thrombocytopenia and uremia. It is an important cause of acute renal failure (ARF) in children all over the world. The present study was carried out to assess the incidence, clinical presentation, hematological and biochemical profile of children presenting with HUS from 1987 to 1990. Out of the 100 cases who presented with ARF 22 had HUS. A majority of these children were males below 1 year of age, and had a prodromal phase of mainly gastrointestinal manifestations lasting for about a week. Anemia was a constant feature followed by bleeding diathesis, mainly melena and purpura. Neurological manifestations included altered sensorium, irritability, coma, hypertensive encephalopathy and convulsions. Renal problems mainly included oliguria, hypertension, hematuria and edema. Investigations revealed thrombocytopenia and microangiopathic hemolytic anemia in all cases. Evidence of disseminated intravascular coagulation (DIC) was observed in 3 cases as decreased fibrinogen levels, increased fibrinogen degradation products and deranged clotting studies. Blood biochemistry revealed azotemia in all cases, hyponatremia in 5 cases, hypernatremia in 3 cases and hyperkalemia in 12 cases. Stool culture showed the presence of Shigella in 8, E. coli in 6 and Klebsiella in 4 cases. Out of 22 cases of HUS, 15 were treated conservatively; of these 2 died. Both of these deaths were due to DIC 7 children were put on peritoneal dialysis; only 1 child died in this group. Factors affecting the outcome were duration of oliguria, levels of blood urea and presence of encephalopathy.(ABSTRACT TRUNCATED AT 250 WORDS)

A clinico-bacteriological profile of leprosy in children.

Clinico-bacteriological profile of 106 leprosy patients below 15 years of age was studied. Majority of the patients were males and fell in the 10-15 years age group (p less than 0.01). Nearly 89% had not received any prior treatment because of financial constraints. Seventy per cent gave a positive history of contact with adult patients who were mainly of the lepromatous variety (p less than 0.01). Skin lesions were present in 103 cases, mainly on the exposed areas and their number was found to increase significantly with advancing age (p less than 0.01). These lesions were hypopigmented patches in 71% of the children and erythematous in the rest. Cutaneous sensations were affected in most of the patients while nerve thickening was observed in 45. Positivity of the skin smears increased significantly as the number of skin lesions per patient increased (p less than 0.05). With advancing age, the disease moved from the tuberculoid end of the spectrum towards the lepromatous end (p less than 0.01).