Clinical evaluation of the enhanced Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test for rapid diagnosis of tuberculosis in prison inmates.

The reliability of the enhanced Amplified Mycobacterium Tuberculosis Direct Test (E-MTD; Gen-Probe, Inc., San Diego, Calif.) for rapid diagnosis of pulmonary tuberculosis (TB) was evaluated by testing 1, 004 respiratory specimens from 489 Texas prison inmates. Results were compared to those of mycobacterial culture (BACTEC TB 460 and Middlebrook 7H11 biplates), smear for acid-fast bacilli (AFB; auramine O), and clinical course. After chart review, three patients (nine specimens) who were on antituberculosis therapy before the study began were excluded from final analysis. Of the remaining 995 specimens, 21 were AFB smear positive: 13 grew Mycobacterium tuberculosis complex (MTBC), 6 grew nontuberculous mycobacteria, and 2 (from two patients diagnosed with TB and started on therapy after the study began) were culture negative. Twenty-eight specimens (20 patients) were positive for MTBC by culture and E-MTD. Seven specimens (seven patients) were positive by culture alone; three were from patients who had other E-MTD-positive specimens, two were false-positive cultures, and two were false-negative E-MTD results. Eight specimens were positive by E-MTD only; four specimens (four patients) were false-positive E-MTD results, and four specimens were from two patients with earlier E-MTD-positive specimens that grew MTBC. Thus, there were 22 patients with TB (10 smear positive and 12 smear negative). The sensitivity and specificity of the AFB smear for diagnosis of TB, by patient, were 45.5 and 98.9%, respectively. After resolving discrepancies, these same values for E-MTD were 90.9 and 99.1% overall, 100 and 100% for the smear-positive patients, and 83.3 and 99.1% for the smear-negative patients. Excluding the one smear-negative patient whose E-MTD-negative, MTBC culture-positive specimen contained inhibitory substances, the sensitivity of E-MTD was 95.2% overall and 90.9% in smear-negative patients. The specificity and positive predictive value of E-MTD can be improved, without altering other performance characteristics, by modifying the equivocal zone recommended by the manufacturer. These data suggest that E-MTD is a reliable method for rapid diagnosis of pulmonary TB, irrespective of the AFB smear result. Guidelines for the most appropriate use of E-MTD with smear-negative patients are needed.

Significance of p53 overexpression in bone marrow biopsies from patients with bone marrow failure: aplastic anemia, hypocellular refractory anemia, and hypercellular refractory anemia.

Among patients with bone marrow failure, differentiating acquired aplastic anemia (AA) from hypocellular refractory anemia (hypo RA) can be a difficult and challenging task. Morphological, cytochemical, immunocytochemical, and cytogenetic studies may provide tools for discriminating between both entities. In addition, differences in the pattern of proliferation and apoptosis of bone marrow cells in AA and in the myelodysplastic syndrome have been reported. Because of the correlation between p53 and apoptosis, we examined the overexpression of p53 on bone marrow biopsies in RA and AA. Our study included 14 patients with hypo RA, 14 patients with hypercellular (hyper) RA, ten patients with classic acquired AA, and 37 hematologically normal individuals. p53 was overexpressed in eight (57%) hypo RA patients and 11 (79%) hyper RA patients. All normal individuals and patients with AA showed no overexpression of p53 in their marrow. These results were statistically significant:p < 0.01 (AA vs hypo RA), p<0.001 (AA vs hyper RA), while the difference between hypo RA and hyper RA was not statistically significant. We conclude that p53 overexpression in bone marrow biopsies is a valuable tool for studying bone marrow failure and may provide additional information to help differentiate hypo RA from acquired AA.

Role of T lymphocyte subsets in immunity to spotted fever group Rickettsiae.

To evaluate the roles of CD4 and CD8 T lymphocytes in immunity to disseminated endothelial infection with Rickettsia conorii (Malish 7 strain), these T cell subsets were depleted or adoptively transferred into subsequently infected C3H/HeN mice. CD4 T lymphocyte-depleted and sham-depleted mice underwent a similar course of illness with a sublethal rickettsial dose, cleared the infection by day 10, and recovered on days 10 to 11. In contrast, mice depleted of CD8 lymphocytes or CD4 and CD8 lymphocytes died or remained persistently infected through day 15 with the ordinarily sublethal dose. Endothelium was the major site of rickettsial persistence, including sites in the vital organs, brain, and lungs of CD8 lymphocyte-depleted mice. In nondepleted animals, CD8 T lymphocytes were observed in apposition to endothelial cells on day 10 at the time of rickettsial clearance. Adoptive transfer of immune CD4 or CD8 T lymphocytes protected mice against a lethal dose of R. conorii in the disseminated endothelial target model. Nonimmune CD4 or CD8 lymphocytes and immune lymphocytes that had passed through columns that depleted both CD4 and CD8 lymphocytes failed to protect mice against R. conorii. These studies represent the first analysis of the role of T lymphocyte subsets in immunity to spotted fever group rickettsiae and the first demonstration that clearance of spotted fever group rickettsiae from endothelial cells requires immune CD8 T lymphocytes.

Novel and frequent mutations of hepatitis B virus coincide with a major histocompatibility complex class I-restricted T-cell epitope of the surface antigen.

We examined the full-length hepatitis B virus (HBV) envelope (surface antigen or HBV small surface antigen [HBsAg]) sequences of 12 different liver samples from 10 different hepatoma-containing chronic carriers. Surprisingly, novel and frequent mutations occurred predominantly at amino acids 40 and 47 of HBsAg, in addition to within a known protective B-cell epitope (so-called group a determinant of HBsAg 124-148). Approximately 58% of chronic carriers contain mutations at the group a determinant. The mutation frequency at the hotspot codons 40 and 47 is approximately 83%, 1 order of magnitude higher than at the known polymorphic sites of subtype-specific determinants at codons 122 and 160, which is approximately 4%. This new mutational domain is found to coincide with a major histocompatibility complex class I-restricted T-cell epitope. The potential biological significance of this novel mutation in the immunopathogenesis of HBV chronic carriers is discussed.

Recurrent massive hemothorax in Rendu-Osler-Weber syndrome.

We report a case of Rendu-Osler-Weber syndrome, occurring as sudden death after two episodes of massive hemothorax. Autopsy revealed massive hemothorax resulting from spontaneous rupture of one of three subpleural arteriovenous malformations. Review of the patient's hospital records showed that she had had a massive spontaneous hemothorax 13 years earlier that was managed conservatively. This case emphasizes the importance of early therapeutic (surgical or radiologic) intervention in the treatment of pulmonary arteriovenous malformations.

Pulmonary malakoplakia in acquired immunodeficiency syndrome: an ultrastructural study of morphogenesis of Michaelis-Gutmann bodies.

Malakoplakia is an unusual inflammatory reaction to a variety of infections, characterized by the accumulation of macrophages containing the target-like calcospherites, the Michaelis-Gutmann body (MGB). We report three patients with acquired immunodeficiency syndrome with pulmonary malakoplakia associated with Rhodococcus equi infection; two patients were diagnosed at autopsy and one by examination of a transbronchial biopsy specimen. All three patients had pulmonary bacterial cultures and light and electron microscopic examination. The patients were 33-, 41-, and 43-year old men, human immunodeficiency virus-positive for 2, 6, 8 years, respectively. The two patients diagnosed at autopsy had cavitary lesions, and the patient diagnosed by biopsy specimen had nodular lesions on chest radiographs. Histologically, the lungs had well-circumscribed areas of infiltration with benign macrophages with granular cytoplasm, scattered MGBs, and numerous gram-positive coccobacilli. Electron microscopic examination showed intracellular coccobacilli, from 990 X 702 to 972 X 648 nm in diameter, with thick, homogenous cell walls, trilaminar cytoplasmic membranes, and dense cytoplasm with from one to five vacuoles. Electron microscopic studies showed that the bacteria within the pulmonary macrophages had thicker cell walls, less prominent nucleoid areas, and more vacuoles than the bacteria in cultures from the sputum and blood. The mature MGB ultrastructurally had a concentric, trilaminate structure with central mineralized core and was without recognizable bacterial forms. Early MGBs, however, consisted of a circular, electron-dense core containing bacteria, ultrastructurally similar to the R. equi seen in the culture. Pulmonary malakoplakia in patients with the acquired immunodeficiency syndrome might thus represent an acquired macrophage dysfunction of the intracellular digestion of phagocytized bacteria. The bacteria within the macrophages, however, seemed to have thicker cell walls compared with those in culture, and thus might be protected from enzyme digestion. It seems that MGBs are formed around the undigested bacteria as an alternative pathway for bacterial destruction, because R. equi was identified within the cores of early MGBs but not the mature or late stage MGBs.

Splenic lymphoma with circulating villous lymphocytes.

This report describes the occurrence of splenic lymphoma with villous lymphocytes (SLVL) in a 56 year old white female with a family history of chronic lymphocytic leukaemia. Other unusual features included a marked lymphocytosis with counts up to 224 x 10(9)/l and marked clumping of lymphocytes in EDTA anticoagulated blood. The neoplastic cells were CD19+, CD20+, CD22+, CD22+, IgM+, lambda+, kappa-, CD5-, and CD10-. The spleen had nodular infiltrates of B lymphocytes in the region of the white pulp with minimal red pulp involvement. Electron microscopy of peripheral blood lymphocytes revealed cells with polar cytoplasmic processes. This report underlines the need for detailed analysis, including morphology and immunophenotyping, for each patient with a small B cell lymphoproliferative disorder.