Biomedical subjects
Publications and source records attributed to R Cisterna.
We present the case of an infertile patient, whose first attempt at IVF had to be postponed for 18 months due to a vulvovaginal yeast infection refractory to treatment. The main causative organism was a Candida glabrata strain resistant to all the imidazolic agents tested. The organism and the host's humoral status were studied in depth, looking for possible causes of the refractoriness to treatment.
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Anti-Candida albicans germ tube antibodies can be induced in rabbits immunized with different C. albicans extracts. Antigens responsible for the induction of those antibodies have molecular weights of approximately 230-250, 62, 43 and 41 kDa. These antigens are present in the cell wall of both C. albicans morphological forms, although their location seems to be different.
To investigate the association between human immunodeficiency virus (HIV) and Epstein-Barr virus (EBV), simultaneous determinations of HIV antigen (HIV Ag) p24 and EBV DNA were performed in lymphocyte culture supernatants from 63 individuals at risk of HIV infection. In vitro data, together with HIV immune status results, were subjected to a statistical analysis. HIV infection was identified in 49 patients (78%); of these, in vitro EBV DNA was found in 44 individuals (90%), while in only 3 of the 14 non-infected ones (21%). Statistical analysis demonstrated a close relationship between evidence of HIV infection and in vitro detection of EBV DNA (87.3% concordant with 95% confidence interval: 76.5%-94.5%). Furthermore, a strong dependence was revealed between the presence of EBV DNA and HIV Ag in culture (p less than 0.00001). These results indicate the existence of in vitro viral interactions, with likely in vivo implications in the pathogenesis and evolution of HIV infection.
The polymerase chain reaction (PCR) was used to detect hepatitis B virus DNA (HBV-DNA) in serum samples of 104 chronic HBV carriers. Of 22 patients positive for both HBV surface (HBsAg) and HBVe (HBeAg) antigens, seven were positive for HBV-DNA on dot blot hybridisation, and all 22 positive in the PCR. Of 41 HBsAg positive patients who had antibodies against HBeAg (anti-HBe), only three were positive for DNA-HBV on dot blot hybridisation, however DNA was detected in 30 of them with the PCR. Similarly, of 41 individuals with antibodies against HBsAg (anti-HBs), 23 yielded positive results in the PCR technique, although dot blot hybridisation detected HBV-DNA in only one patient. These results indicate that while serological and conventional DNA hybridisation assays are not sensitive enough to determine the infectivity of HBV chronic carriers, PCR is an accurate method for establishing the status and progression of disease in these patients.
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The existence of molecular transactivations between EBV and HIV-1, as well as reactivations of EBV latent infections in AIDS patients, have been recently documented. In order to shed more light on the putative association between EBV and HIV, and its role in the evolution to AIDS, we have determined simultaneously p24 protein and EBV DNA in culture supernatants of peripheral blood mononuclear cells from 47 individuals suspected of having HIV infection. The results of the in vitro assays were correlated with the clinical stage of the individuals and their serologic status to EBV. Statistical analysis showed a concordance between HIV infection and in vitro detection of EBV DNA (P < 0.002); particularly, a strong correlation between the presence of EBV DNA and p24 in culture was observed (P < 0.001). These results are consistent with the occurrence of viral interactions, manifested in vitro. However, in our series, the appearance of EBV DNA in culture was not concomitant with an elevation of anti-VCA IgG titers, anti-EA titers or the development of symptomatology, suggestive of a reactivation of a latent EBV infection or a progression of HIV infection. Therefore we conclude that, although interaction between both viruses may take place at the molecular level, there is no clear evidence of the repercussion that this event may have on the clinical course of HIV infection.
The potential of the transfer of natural plasmids between sewage strains has been studied. In vitro transfer was conducted at 37 degrees C in tryptone soya broth and sterile raw sewage as mating media. In situ transfer was carried out in sterile raw sewage within membrane diffusion chambers at 10.6 degrees C. When the recipient was a laboratory strain of Escherichia coli K-12, the in situ frequency values were significantly lower (P less than 0.001) than those obtained in vitro for the same mating pair. When the laboratory recipient was replaced with recipients from the same sewage source, frequency values decreased progressively from the optimum conditions to the most adverse. However, in situ frequency values were higher than those for the same donors mated with a laboratory recipient.
The influence of biotic and abiotic factors on plasmid transfer between Escherichia coli strains in terms of the variation in the number of transconjugants formed and the variation in transfer frequency was investigated. The density of parent cells affected the number of transconjugants, reaching a maximum when the cell density was on the order of 10(8) CFU ml-1. As the donor-to-recipient ratios varied from 10(-4) to 10(4), the number of transconjugants varied significantly (P less than 0.001), reaching a maximum with donor-to-recipient ratios between 1 and 10. The concentration of total organic carbon in the mating medium affects both the number of transconjugants and the transfer frequency, being significantly higher (P less than 0.001) when the total organic carbon concentration was higher than 1,139 mg of C liter-1. However, the transconjugants were detected even with less than 1 mg of C liter-1. Linear regression of log10 transconjugants versus mating temperature showed a highly significant regression line (P less than 0.001). Neither the transfer frequency nor the transconjugant number varied significantly in the range of pHs assayed. We can conclude that plasmid transfer by conjugation can take place within a wide range of conditions, even in such adverse conditions as the absence of nutrients and low temperatures.
A new method of Candida subspecies discrimination on Sabouraud-triphenyltetrazolium agar is reported. Five hundred sixty-two strains of Candida and Torulopsis glabrata, previously identified by conventional mycological methods, were studied. Each strain received a three-letter code and a number based on its colonial morphology. Sixteen morphotypes were found for Candida albicans, 6 were found for Candida parapsilosis, 4 were found for both Candida guilliermondii and Candida krusei, and 12 were found for Candida tropicalis. None of the 56 T. glabrata strains studied grew on this agar. A reproducibility of 95% was found for C. albicans. The simplicity and low cost could make this method useful for typing Candida spp.
BACKGROUND: The purpose of this work was to study the molecular basis of beta-lactamase production in ampicillin-resistant strains of Salmonella spp. METHODS: It was performed analytical isoelectric focusing of beta-lactamases produced by a group of 33 strains selected in basis of their resistance phenotype. Plasmid profile analysis and assays of transferable drug resistance were developed. The study was completed by hybridization experiments with an intragenic TEM probe which allowed the location of the bla-TEM gene. RESULTS: By analytical isoelectrofocusing we found that 26 out of the 27 ampicillin-resistant strains produced beta-lactamases with pl 5.4 and/or 5.6 corresponding to TEM-1 and/or TEM-2 type. Analysis of plasmid DNA revealed in almost all strains plasmids ranging in size from 1.1 to 125 Mdal. This plasmids were responsible of the resistance and, moreover, were able to transfer the resistance by conjugation mechanisms. Southern blot analysis detected the gene that code the TEM beta-lactamase at the 125, 8 and 5.8 Mdal plasmids. CONCLUSIONS: Resistance to ampicillin in the strains of Salmonella studied was due to the presence of TEM type beta-lactamases coded by conjugative plasmids. These plasmids coded also resistance to other antimicrobial agents. Our results showed that the use of a DNA probe to the detect TEM-type beta-lactamases using a non radioactive probe, could be a suitable alternative to isoelectric focusing.
The evaluation of the presence of p24 antigen on the membrane of peripheral blood mononuclear cells from 31 HIV infected individuals is presented. The study was performed by indirect immunofluorescence and flow cytometry and the data were analyzed by the Kolmogorov-Smirnov test. Values obtained [D/s(n)] result from the comparison of the fluorescence histograms of each sample with a control one. Cases showing p24 Ag on peripheral blood mononuclear cells also presented percentages of CD3, HLA-DR positive cells significantly higher than p24 negative ones. In addition, D/s(n) values were superior in symptomatic patients than in asymptomatic ones, which indicate the existence of a correlation between flow cytometry results, viral replication and clinical course. Nevertheless in this study, as well and in previous ones, a high degree of cross reactivity between the anti-p24 monoclonal antibody employed and normal lymphocytes has been observed. This reactivity is localized preferentially in the CD4 positive subset.
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HBV infection has been investigated in 47 anti-human HIV positive patients in relation to a similar group of 33 anti-HIV negative patients. Serological HBV markers were found in 87% of anti-HIV positive patients. The difference in markers of viral replication (HBeAg, HBV-DNA) was not statistically significant between the two groups. It has been suggested that HBV infection of peripheral blood mononuclear cells could be a cofactor implicated in the development of immunodeficiency due to HIV. For this reason we have investigated the presence of HBV-DNA in peripheral blood mononuclear cells by in situ hybridization. Although its detection was more frequent in anti-HIV positive patients than in anti-HIV negative ones (p < 0.05), it was not related to clinical state of immunodeficiency. With regard to serological HBV markers, HBV-DNA was detected in peripheral blood mononuclear cells from antiHBc w/o antiHBs patients. This fact means the virus may persist in this cells after recovery and suggest they could serve as additional reservoirs of HBV. These cells, that contain the HBV genome, could be implicated in the perpetuation, reactivation of the infection and in its transmission.
BACKGROUND: The presence of hepatitis delta virus was investigated in liver biopsies by in situ cyto-hybridization with a probe labelled with digoxigenin. METHODS: The techniques developed included extraction of plasmid DNA by lysis by alkali, electroelution, electrophoresis in agarose gel and digoxigenin labelling, and the application in liver tissue. RESULTS: Viral RNA was detected in 6 of the 10 patients, and the reactivity was exclusively restricted to the nucleolus of the hepatocytes. CONCLUSION: This method reveals as a sensitive and quick diagnostic procedure, which allows to study the intensity of the infection as well as the serological state of the patient.
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BACKGROUND: Pneumonia due to pathogenic bacteria is common in HIV infected patients. However, a global overview has not been yet performed and also the effectiveness of a non-invasive approach has not been tested. PATIENTS AND METHODS: An open study was conducted in 1989 including all HIV infected patients with a presumptive or definite diagnosis of bacterial pneumonia. Fiberoptic bronchoscopy procedures were used only in patients with a difficult diagnosis and/or problems in their clinical evolution. Clinical and biological data were compared among different etiologic groups, and their evolution was also recorded in relation with the use of non-invasive procedures. RESULTS: Among a total of 302 admissions of HIV infected patients, 113 developed a primary or secondary respiratory tract infection. Forty-three of those were bacterial pneumonia episodes. Etiologic diagnosis was achieved with non invasive methods in 18 cases (42%). Blood cultures were the most effective method used. Streptococcus pneumoniae was the most frequently identified microorganisms (9 cases, 21%). The identification of other bacteria was linked with more severe forms of disease regarding immunologic (leukopenia, p less than 0.001; T4 lymphopenia, p less than 0.05), and respiratory parameters (hypoxemia, p less than 0.05). Two deaths were directly related with pneumonia of known etiology. CONCLUSIONS: Bacterial pneumonia is the most frequent respiratory infection in HIV infected patients. An initial non-invasive approach yields a reasonable percentage of etiologic diagnosis and is not related with a poorer outcome.
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