A comparision of brain biopsy and CSF-PCR in the diagnosis of CNS lesions in AIDS patients

Twenty patients with AIDS who had intracranial lesions underwent both brain biopsy and cerebro-spinal fluid (CSF) examination to compare histological diagnosis with the polymerase chain reaction (CSF-PCR) for the identification of infectious agents. CSF-PCR was performed for herpes simplex virus, varicella zoster virus, cytomegalovirus (CMV), JC virus (JCV), Epstein-Barr virus (EBV), Toxoplasma gondii and Mycobacterium tuberculosis. A definitive diagnosis was obtained by brain biopsy in 14 patients (2 with astrocytoma, 12 with brain infection). CSF-PCR was positive for EBV DNA in 3 of 3 cases of primary cerebral lymphoma, positive for JCV DNA in 6 of 7 biopsy-proven (and one autopsy-proven) cases of progressive multifocal leukoencephalopathy (PML). CSF-PCR was positive for CMV DNA in one biopsy-proven and one autopsy-proven case of CMV encephalitis (the former also had PML) and positive for M. tuberculosis DNA in one case of tuberculous encephalitis. None of the five toxoplasmic encephalitis cases (one definite, four presumptive) were T. gondii DNA positive. There was close correlation between histology and CSF-PCR for CMV encephalitis, PML and PCL. Antitoxoplasma therapy affected the sensitivity of both histological and CSF-PCR methods. Received: 8 November 1995 Received in revised form: 9 July 1996 Accepted: 19 July 1996     

The regional distribution of adenosine-regulating enzymes in the left and right ventricle walls of control and hypertrophic heart

Summary The transmural distribution of the adenosine-generating enzyme 5-nucleotidase (5N) and of the adenosine-degrading enzymes adenosine deaminase (ADA), AMP deaminase (AMP-D) and adenosine kinase (Ado-K) were determined across the walls of left and right ventricles of control and hypertrophic rat hearts.The enzyme distribution across the left ventricle wall (but not across the right wall) of normal hearts was not uniform: 5N activity shows its highest levels in the subepicardial and in the subendocardial regions, whereas all the other enzyme activities show their lowest levels. A similar pattern of transmural distribution was also detected in other mammalian species (ox and pig).In the experimental cardiac hypertrophy, caused by two different types of chronic cardiac overload, the levels and the profiles of transmural distribution of 5N and ADA enzyme activities may significantly change across the rat left ventricle wall.     

Ultrastructural features of the muscle fibers in congenital hypothyroidism

Summary Muscles of three patients (2 years, 2 years and 7 months, and 11 years of age) affected by congenital hypothyroidism due to dysgenesis of the thyroid gland were studied by electron microscope. The biopsies were performed on the quadriceps muscle. Several alterations were found in the ultrastructure of the muscle fibers. The contractile material underwent atrophy by two recessive processes: by reduction and by degeneration. The former, which affected mainly the white fibers, led to the progressive detachment of myofilaments from the periphery of the myofibrils while they maintainad a normal arrangement in the center. Consequently, their diameter decreased and their interfibrillar spaces enlarged. The latter enlargment, involving the red fibers, caused areas of dedifferentiation involving either the contractile apparatus or the other cell organelles. Modifications of the sarcolemma and basement membrane were seen in the fibers affected by the atrophying processes of the contractile material. Changes of the ultrastructure of the sarcoplasmic reticulum and of the T system were observed with numerous morphological abnormalities of the mitochondria. In addition, accumulation of glycogen and striking changes of the satellite cells were found. The ultrastructural findings are reviewed and discussed in relation to the literature on hypothyroid myopathy.     

A PCR-colorimetric microwell plate hybridization assay for detection of Mycobacterium tuberculosis and M. avium from culture samples and Ziehl-Neelsen-positive smears

Differentiation between Mycobacterium tuberculosis and M. avium is essential for the treatment of mycobacterial infections. We have developed an easy and rapid detection assay for the diagnosis of mycobacterial diseases. This is a PCR-hybridization assay based on selective amplification of a 16S rRNA gene sequence using pan-Mycobacterium primers followed by hybridization of the amplification products to biotinylated M. tuberculosis and M. avium-specific probes. A total of 55 mycobacterial isolates were tested. For all isolates, results concordant with those of conventional identification methods were obtained. Moreover, we developed a method for extraction of DNA from Ziehl-Neelsen-positive smears which allows the recovery of intact target DNA in our PCR-hybridization assay. Our method was able to confirm all culture results for 59 Ziehl-Neelsen-positive smears from clinical specimens (35 sputum, 11 lymph node biopsy, 6 stool, 4 pus, 2 urine, and 1 pericardial fluid specimens). These data suggest that our PCR-hybridization assay, which is simple to perform and less expensive than commercial probe methods, may be suitable for the identification of M. tuberculosis and M. avium. It could become a valuable alternative approach for the diagnosis of mycobacterial infections when applied directly to DNA extracted from Ziehl-Neelsen-positive smears as well.     

Molecular epidemiology study of exogenous reinfection in an area with a low incidence of tuberculosis

In geographical areas with a low incidence of tuberculosis, recurrent tuberculosis is generally due to reactivation of the disease. However, the relative contribution of tuberculosis reinfection increases in parallel with the incidence of disease and is likely to depend on the epidemiological context: factors such as the spread of multidrug resistance, human immunodeficiency virus (HIV) infection, and immigration from developing countries could modify disease transmission in areas at low risk for tuberculosis. A molecular epidemiology study was performed in Lombardy, Northern Italy, where the incidence of tuberculosis is 17.5 cases per 100,000 persons. A total of 2,452 cases of culture-confirmed tuberculosis in 2,127 patients were studied. A group of 32 patients (1.5%), each of whom had two episodes of tuberculosis with cure as the outcome of the first episode and with more than 6 months between the two episodes, were studied by means of restriction fragment length polymorphism DNA fingerprinting analysis. For 5 of the 32 patients (16%), the DNA fingerprinting patterns of Mycobacterium tuberculosis strains responsible for the second episode did not match those of the corresponding isolates of the first episode, indicating exogenous reinfection. Two of these patients developed multidrug-resistant tuberculosis during the second episode, and in three cases the isolates belonged to clusters of M. tuberculosis strains spreading in the community. A fourfold-increased risk for reinfection was observed in immigrant patients compared to Italian subjects. In contrast, a higher risk of relapse rather than reinfection was evidenced in HIV-positive subjects and in patients infected with multidrug-resistant tuberculosis. Episodes of tuberculosis reinfection in areas with a low incidence of tuberculosis are rare compared to those in high-incidence geographical regions. In populations that have immigrated from high-risk areas, reinfection may represent a considerable contributor to the rate of recurrent tuberculosis. This finding emphasizes the importance of containing the spread of epidemic strains in close communities, in order to prevent changes in global tuberculosis trends for developed countries.     

Employing a systematic approach to biobanking and analyzing clinical and genetic data for advancing COVID-19 research

Within the GEN-COVID Multicenter Study, biospecimens from more than 1000 SARS-CoV-2 positive individuals have thus far been collected in the GEN-COVID Biobank (GCB). Sample types include whole blood, plasma, serum, leukocytes, and DNA. The GCB links samples to detailed clinical data available in the GEN-COVID Patient Registry (GCPR). It includes hospitalized patients (74.25%), broken down into intubated, treated by CPAP-biPAP, treated with O₂ supplementation, and without respiratory support (9.5%, 18.4%, 31.55% and 14.8, respectively); and non-hospitalized subjects (25.75%), either pauci- or asymptomatic. More than 150 clinical patient-level data fields have been collected and binarized for further statistics according to the organs/systems primarily affected by COVID-19: heart, liver, pancreas, kidney, chemosensors, innate or adaptive immunity, and clotting system. Hierarchical clustering analysis identified five main clinical categories: (1) severe multisystemic failure with either thromboembolic or pancreatic variant; (2) cytokine storm type, either severe with liver involvement or moderate; (3) moderate heart type, either with or without liver damage; (4) moderate multisystemic involvement, either with or without liver damage; (5) mild, either with or without hyposmia. GCB and GCPR are further linked to the GCGDR, which includes data from whole-exome sequencing and high-density SNP genotyping. The data are available for sharing through the Network for Italian Genomes, found within the COVID-19 dedicated section. The study objective is to systematize this comprehensive data collection and begin identifying multi-organ involvement in COVID-19, defining genetic parameters for infection susceptibility within the population, and mapping genetically COVID-19 severity and clinical complexity among patients.Subject terms: Genetics research, Viral infection     

Macrophage antitumor activity in vitro. Comparative analysis of cytolytic, cytostatic, and cytotoxic activities of mouse macrophages and human monocytes

Mouse peritoneal M phi and human blood monocytes were assayed for their antitumor activity in vitro with a cytolysis, a cytostasis and a cytotoxicity test performed in parallel. Both natural and stimulus-induced M phi antitumor capacities were assessed. Results indicate that natural cytolytic activity of unstimulated M phi is generally unable to restrict final tumor cell growth, since it is not coupled with cytostatic capacity. In contrast, exposure of M phi in vitro to either MAF or IFN-beta, besides augmenting M phi cytolytic capacity, induced a very significant cytostatic activity and thus efficiently restricted the survival of tumor cells.     

Acquired activated protein C resistance in postmenopausal women is dependent on factor VIII:c levels.

Activated protein C (APC) resistance is an established risk factor for venous thromboembolism. In 5% to 10% of patients with venous thromboembolism, the APC resistance phenotype is observed in the absence of factor V Leiden mutation. Moreover, some physiologic and pathologic conditions are associated with an "acquired" APC resistance, not caused by the Leiden mutation, such as inflammatory diseases, pregnancy, or oral contraceptive therapy. Several studies have demonstrated the effect of menopause on the hemostatic system, but no data are available about APC resistance. We found a high prevalence of APC resistance in postmenopausal women, not associated with factor V Leiden mutation. The mechanism that underlies this acquired APC resistance may be related to the higher levels of factor VIII, which showed a strong inverse correlation with APC resistance, whereas no correlation was found between the normalized APC ratio, factor V levels, and protein S values. Higher levels of factor VIII correlated with a marker of coagulation activation, such as prothrombin fragments 1 plus 2. Therefore, to identify women receiving hormone replacement therapy who have a greater risk for deep venous thrombosis, the APC resistance coagulation test should be used instead of the genetic study.     

IS1245 Restriction Fragment Length Polymorphism Typing of Mycobacterium avium Isolates: Proposal for Standardization

Mycobacterium avium has become a major human pathogen, primarily due to the emergence of the AIDS epidemic. Restriction fragment length polymorphism (RFLP) typing, using insertion sequence IS1245 as a probe, provides a powerful tool in the molecular epidemiology of M. avium-related infections and will facilitate well-founded studies into the sources of M. avium infections in animal and environmental reservoirs. The standardization of this technique allows computerization of IS1245 RFLP patterns for comparison on a local level and the establishment of M. avium DNA fingerprint databases for interlaboratory comparison. Moreover, by combining international DNA typing results of M. avium complex isolates from a broad spectrum of sources, long-lasting questions on the epidemiology of this major agent of mycobacterial infections will be answered.     

Changes in the transmural distribution of antioxidant enzyme activities across the left ventricle heart wall from rats fed ad libitum or food-restricted during growth and aging

Data on vulnerability to injury and on the larger age-related accumulation of lipofuscin in the subendocardial myocardium prompted us to investigate the changes in the levels and in the transmural distribution of catalase (C), glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) activities across the left ventricle heart wall of rats fed ad libitum a standard diet or submitted to intermittent feeding during growth and aging. Enzyme activities were assayed by standard techniques on subepicardial, midmyocardial or subendo- cardial tissue obtained by cutting the heart wall into 100-microm-thick sections at the cryostat. The levels of GSH-Px and of C (but not of SOD) activity increased with age and reached their highest values in the subendocardial region by adulthood or senescence, respectively. No effect was observed of intermittent feeding on age-related changes in enzyme levels and transmural distribution.