Histoplasmin reaction. Comparison of a polysaccharide antigen to the filtrate antigen

This work was planned by taking into account all the knowledge accumulated from the immunological study of paracoccidioidomycosis. It aimed at comparing a polysaccharide antigen from Histoplasma capsulatum to a classic histoplasmin with the help of intradermal tests of delayed type of hypersensitivity. Tests were applied to 115 individuals in Santo Amaro, a town in the state of São Paulo. Positive results using classic histoplasmin were obtained in 46.0% cases whereas positive results using the polysaccharide antigen at its hightest concentration were obtained in 51.30% cases. The major conclusion in this investigation is that it is possible to use the polysaccharide antigen as histoplasmin instead of the filtrate antigen

Detection of mycoplasmas in urethral swabs from HIV-1 infected patients and control individuals using culture techniques and polymerase chain reaction

The objective of the present study was to determine the prevalence of certain mycoplasma species, i.e., Mycoplasma hominis, Ureaplasma urealyticum and Mycoplasma penetrans, in urethral swabs from HIV-1 infected patients compared to swabs from a control group. Mycoplasmas were detected by routine culture techniques and by the Polymerase Chain Reaction (PCR) technique, using 16SrRNA generic primers of conserved region and Mycoplasma penetrans specific primers. The positivity rates obtained with the two methods were comparable. Nevertheless, PCR was more sensitive, while the culture techniques allowed the quantification of the isolates. The results showed no significant difference (p < 0.05) in positivity rates between the methods used for mycoplasma detection.

SCHISTOSOMIASIS MANSONI IN THE "BAIXADA OCIDENTAL MARANHENSE", STATE OF MARANHÃO, BRAZIL: CROSS-SECTIONAL STUDIES PERFORMED IN 1987 AND 1993

A cross-sectional study on the prevalence of schistosomiasis mansoni in three sites of the "Baixada Ocidental Maranhense" was carried out in 1993 in: Alegre (in the municipality of São Bento), Aliança (in Cururupu) and Coroatá II (in the municipality of São João Batista). Results were compared to those of another study performed at the same sites and in similar conditions, in 1987. The entire population of the three sites, with few exceptions, was submitted to fecal tests using the Kato-Katz method and immediate intradermal tests for schistosomiasis in both studies. Subjects with positive results in one of these tests were clinically evaluated by a physical examination. In 1993, the total of 827 subjects were submitted to fecal examination and 826 to intradermal test. Schistosoma mansoni eggs were found in the feces of 154 (18.6%) subjects, while 478 (57.9%) subjects presented a positive intradermal test. Stool examination was carried out in 367 subjects in Alegre with a positivity rate of 14.9%; the intradermal test, performed in 366 subjects, was positive in 47.5% of the cases. In Aliança, 277 subjects had their feces examined and were submitted to an intradermal test, with a positivity rate of 34.4% and 70.7%, respectively. Finally in Coroatá II, 183 inhabitants submitted to fecal and intradermal tests had positivity rates of 2.2% and 59.0%, respectively. When the present data were compared to those obtained in the survey performed in 1987, a significant decrease in the prevalence of infection by S. mansoni was observed in Alegre and Coroatá II, and a prevalence increase in Aliança.

Prevalence of Trypanosoma cruzi and Leishmania chagasi infection and risk factors in a Colombian indigenous population

This study was carried out in order to obtain base-line data concerning the epidemiology of American Visceral Leishmaniasis and Chagas’ Disease in an indigenous population with whom the government is starting a dwelling improvement programme. Information was collected from 242 dwellings (1,440 people), by means of house to house interviews about socio-economic and environmental factors associated with Leishmania chagasi and Trypanosoma cruzi transmission risk. A leishmanin skin test was applied to 385 people and 454 blood samples were collected on filter paper in order to detect L. chagasi antibodies by ELISA and IFAT and T. cruzi antibodies by ELISA. T. cruzi seroprevalence was 8.7% by ELISA, L. chagasi was 4.6% and 5.1% by IFAT and ELISA, respectively. ELISA sensitivity and specificity for L. chagasi antibodies were 57% and 97.5% respectively, as compared to the IFAT. Leishmanin skin test positivity was 19%. L. chagasi infection prevalence, being defined as a positive result in the three-immunodiagnostic tests, was 17.1%. Additionally, 2.7% of the population studied was positive to both L. chagasi and T. cruzi, showing a possible cross-reaction. L. chagasi and T. cruzi seropositivity increased with age, while no association with gender was observed. Age (p<0.007), number of inhabitants (p<0.05), floor material (p<0.03) and recognition of vector (p<0.01) were associated with T. cruzi infection, whilst age ( p<0.007) and dwelling improvement (p<0.02) were associated with L. chagasi infection. It is necessary to evaluate the long-term impact of the dwelling improvement programme on these parasitic infections in this community.

Production of monoclonal antibodies anti-Taenia crassiceps cysticerci with cross-reactivity with Taenia solium antigens

We describe the production of the potential monoclonal antibodies (MoAbs) using BALB/c mice immunized with vesicular fluid (VF)-Tcra (T. crassiceps) antigen. Immune sera presented anti-VF-Tcra (<20kD) IgG and IgM antibodies with cross-reactivity with T. solium (Tso) antigen (8-12, 14, and 18 kD). After cell fusion, we selected 33 anti-Tcra and anti-Tso reactive IgM-clones and 53 anti-Tcra specific IgG-clones, 5 of them also recognizing Tso antigens. Two clones identified the 8-14 and 18kD peptides of VF-Tcra.

Diagnosis of hepatitis C virus in Brazilian blood donors using a reverse transcriptase nested polymerase chain reaction: comparison with enzyme immunoassay and recombinant protein immunoblot assay

Screening blood donations for anti-HCV antibodies and alanine aminotransferase (ALT) serum levels generally prevents the transmission of hepatitis C virus (HCV) by transfusion. The aim of the present study was to evaluate the efficiency of the enzyme immunoassay (EIA) screening policy in identifying potentially infectious blood donors capable to transmit hepatitis C through blood transfusion. We have used a reverse transcriptase (RT)-nested polymerase chain reaction (PCR) to investigate the presence of HCV-RNA in blood donors. The prevalence of HCV-RNA positive individuals was compared with the recombinant immunoblot assay (RIBA-2) results in order to assess the usefulness of both tests as confirmatory assays. Both tests results were also compared with the EIA-2 OD/C ratio (optical densities of the samples divided by the cut off value). ALT results were expressed as the ALT quotient (qALT), calculated dividing the ALT value of the samples by the maximum normal value (53UI/l) for the method. Donors (n=178) were divided into five groups according to their EIA anti-HCV status and qALT: group A (EIA > or = 3, ALT<1), group B (EIA > or = 3, ALT>1), group C (1<=EIA<3, ALT<1), group D (1<=EIA<3, ALT>1) and group E (EIA<=0.7). HCV sequences were detected by RT-nested PCR, using primers for the most conserved region of viral genome. RIBA-2 was applied to the same samples. In group A (n=6), all samples were positive by RT-nested PCR and RIBA-2. Among 124 samples in group B, 120 (96.8%) were RIBA-2 positive and 4 (3.2%) were RIBA-2 indeterminate but were seropositive for antigen c22.3. In group B, 109 (87.9%) of the RIBA-2 positive samples were also RT-nested PCR positive, as well as were all RIBA-2 indeterminate samples. In group C, all samples (n=9) were RT-nested PCR negative: 4 (44.4%) were also RIBA-2 negative, 4 (44.4%) were RIBA-2 positive and 1 (11.1%) was RIBA-2 indeterminate. HCV-RNA was detected by RT-nested PCR in 3 (37.5%) out of 8 samples in group D. Only one of them was also RIBA-2 positive, all the others were RIBA-2 indeterminate. All of the group E samples (controls) were RT- nested PCR and RIBA-2 negative. Our study suggests a strong relation between anti-HCV EIA-2 ratio > or = 3 and detectable HCV-RNA by RT-nested PCR. We have also noted that blood donors with RIBA-2 indeterminate presented a high degree of detectable HCV-RNA using RT-nested PCR (75%), especially when the c22.3 band was detected.

Susceptibility of Biomphalaria tenagophila and Biomphalaria straminea to Schistosoma mansoni infection detected by low stringency polymerase chain reaction

In order to determine Schistosoma mansoni infection rates in Biomphalaria tenagophila and B. straminea, low stringency polymerase chain reaction (LS-PCR) technique was used as a complementary method to light exposure technique. LS-PCR has already been standardized in our laboratory to detect the trematode DNA in B. glabrata. Higher S. mansoni infection rates were detected using conventional method and LS-PCR. The parasite DNA profile was detected in both species after 7-day exposure to miracidia, using LS-PCR. This technique enables early detection of schistosomiasis transmission focuses, in endemic areas, before the beginning of cercariae shedding.

Polymerase chain reaction (PCR) for the detection of Salmonella in artificially inoculated chicken meat

The aim of this study was to develop a polymerase chain reaction (PCR) protocol for the detection of Salmonella in artificially contaminated chicken meat. Tests were performed with different dilutions of Salmonella Typhimurium or Salmonella Enteritidis cells (10-7, 10-8 or 10-9 CFU/mL) inoculated in chicken meat samples, in order to establish the limits of detection, incubation times (0, 6, 8 and 24 hours of pre-enrichment in PBW 1%) and three DNA extraction protocols (phenol-chloroform, thermal treatment and thermal treatment and Sephaglass). The assay was able to detect until 10-9 CFU/mL of initial dilution of Salmonella cells inoculated in chicken meat, which allows detection of Salmonella within 48 hours, including 24 hours of pre-enrichment and using the phenol-chloroform DNA extraction protocol. As the results are obtained in a shorter time period than that of microbiological culture, this procedure will be useful in the methodology for detection of Salmonella in chicken.

Immunofluorescence assay reactivity patterns of serum samples presenting indeterminate Western blot results for antibodies to HIV-1 and HTLV-I/II in Cordoba, Argentina

Serum samples (n: 110) from blood donors and high risk individuals from Cordoba, Argentina with indeterminate HIV-1 and HTLV-I/II Wb profiles were studied for specific antibodies to HTLV-I/II and HIV-1 by indirect immunofluorescence assay (IFA) and for the presence or absence of HIV-1 and HTLV-I/II specific bands by Wb. This study was carried out in order to characterize their putative reactions with HIV-1 and HTLV-I/II proteins and to resolve the retrovirus infection status of these individuals. Results indicated that blood donors sera displaying indeterminate HIV-1 or HTLV-I/II Wb patterns were not immunoreactive to HTLV-I/II and HIV-1 on IFA. However, a high rate of indeterminate HIV-1 and HTLV-I/II Wb samples from high risk individuals had positive HTLV-I/II and HIV-1 IFA results respectively. Our study supports the growing evidence that HTLV-HIV indeterminate seroreactivity in low risk population is due to a cross reaction against nonviral antigens, and in high risk populations the indeterminate samples show serological cross-recognition between HIV-1 proteins and HTLV-I/II proteins on Wb. These results point out the necessity to investigate the HTLV-I/II reactivity in indeterminate HIV-1 samples and viceversa in order to confirm the diagnosis. Finally, this study shows the potential usefulness of IFA in elucidating the status of HIV-1 and HTLV-I/II infection of individuals with indeterminate Wb profiles, thus enabling resolution of retrovirus infection status.

Chagas disease in the Brazilian Amazon: IV. a new cross-sectional study

A new conglomerate family sample of 194 dwellings with 996 resident persons were studied in the town of Barcelos, State of Amazonas, in order to re-evaluate the risk of Chagas disease. During the survey the persons were interviewed and in this occasion we showed to them a collection of Panstrongylus, Rhodnius and Triatoma, asking if they recognized and eventually have been bitten by this kind of bugs. At this time we collected 500 ul of blood in microtainer® tubes from 886 interviewed persons who gave permission after informed consent. A screening test for T. cruzi antibodies based on agglutination of colored polymer particles, sensitized with three different synthetic peptides of T. cruzi (ID-PaGIA Chagas Test)®, showed 13.2% of sera positivity, but only 6.8% were confirmed by indirect immunofluorescence, and ELISA with purified T. cruzi antigens. Two hundred and six interviewed persons (20.7%) recognized the triatomines, as "piaçavas' lice" and 62 (30%) confirmed that have been bitten by the bugs, 25.8% of them had a positive serology for T. cruzi infection. Electrocardiographic alterations were shown in 9.3% of the seropositives and in 11.9% of the seronegative cases. This was considered not statistically significant.

Comparative study of the fecundity and fertility of Biomphalaria glabrata (Say, 1818) and Biomphalaria straminea (Dunker, 1848) in a laboratory through self-fertilization and cross-fertilization

The aim of this study was to compare the fecundity and fertility of B. glabrata and B. straminea by cross- and self-fertilization. To attain this objective, laboratory-raised strains of B. glabrata and B. straminea were used. The former originated from natural breeding grounds in the municipality Paulista, state of Pernambuco, Brazil. The latter originated from irrigation ditches in the municipality of Petrolândia, in the same state. Snail populations of B. glabrata and B. straminea were maintained for 240 days in laboratory. Their fecundity was evaluated by noting the number of egg-masses, eggs and eggs per mass. Their fertility was evaluated by the number of viable eggs and the hatching rate. B. straminea was markedly more fecund than B. glabrata through cross- and self-fertilization, namely: greater egg-mass; higher egg production and more eggs per mass. Regarding fertility, there seemed to be no preferential period for occlusion to occur or a trend in the rhythm of producing viable eggs.

Single step polymerase chain reaction (PCR) for the diagnosis of the Leishmania (Viannia) subgenus

In Brazil, the main etiologic agent of Leishmaniasis that frequently presents with mucosal involvement belongs to the Viannia subgenus. The therapeutic conduct in this disease depends on the parasitological diagnosis, and classical methods are restricted in identifying the agent. In this paper we describe a polymerase chain reaction (PCR), which uses primers designed from mini-exons repetitive sequences. The PCR amplifies a 177bp fragment that can distinguish (Viannia) from (Leishmania) subgenus. This test could be a useful diagnostic tool.