A Simple Reverse Transcription-Polymerase Chain Reaction for Dengue Type 2 Virus Identification

We show here a simplified reverse transcription-polymerase chain reaction (RT-PCR) for identification of dengue type 2 virus. Three dengue type 2 virus strains, isolated from Brazilian patients, and yellow fever vaccine 17DD, as a negative control, were used in this study. C6/36 cells were infected with the virus, and tissue culture fluids were collected after 7 days of infection period. The RT-PCR, a combination of RT and PCR done after a single addition of reagents in a single reaction vessel was carried out following a digestion of virus with 1% Nonidet P-40. The 50ml assay reaction mixture included 50 pmol of a dengue type 2 specific primer pair amplifying a 210 base pair sequence of the envelope protein gene, 0.1 mM of the four deoxynucleoside triphosphates, 7.5U of reverse transcriptase, and 1U of thermostable Taq DNA polymerase. The reagent mixture was incubated for 15 min at 37oC for RT followed by a variable amount of cycles of two-step PCR amplification (92oC for 60 sec, 53oC for 60 sec) with slow temperature increment. The PCR products were subjected to 1.7% agarose gel electrophoresis and visualized with UV light after gel incubation in ethidium bromide solution. DNA bands were observed after 25 and 30 cycles of PCR. Virus amount as low as 102.8 TCID50/ml was detected by RT-PCR. Specific DNA amplification was observed with the three dengue type 2 strains. This assay has advantages compared to other RT-PCRs: it avoids laborious extraction of virus RNA; the combination of RT and PCR reduces assay time, facilitates the performance and reduces risk of contamination; the two-step PCR cycle produces a clear DNA amplification, saves assay time and simplifies the technique

Prostaglandin A1 Inhibits Replication of Classical Swine Fever Virus

Prostaglandins (Pgs) have been shown to inhibit the replication of several DNA and RNA viruses. Here we report the effect of prostaglandin (PgA1) on the multiplication of a positive strand RNA virus, Classical Swine Fever Virus (CSFV) in PK15 cells. PgA1 was found to inhibit the multiplication of CSFV. At a concentration of 5 µg/ml, which was nontoxic to the cells, PgA1 inhibitis virus production in 99%. In PgA1 treated cells the size and number of characteristic Classical Swine Fever focus decreased in amount.

The Effects of a DNA Virus Infection on the Reproductive Potential of Female Tsetse Flies, Glossina morsitans centralis and Glossina morsitans morsitans (Diptera: Glossinidae)

Reproductive anomalies associated with the tsetse DNA virus infection in the female tsetse hosts, Glossina morsitans centralis Machado and Glossina morsitans morsitans Westwood, inoculated with the virus during the 3rd instar larval stage were studied and the data compared to those obtained from the control females injected with sterile physiological saline. Virus infected flies had significantly longer first and second pregnancy cycles (P<0.0001) and produced pupae that were of significantly less weight in milligrams (P<0.0001) compared to controls. Transmission of the virus to progeny was not absolute and only 21% of G. m. centralis and 48% of G. m. morsitans first progeny flies from infected females developed salivary gland hypertrophy as a result of transmission from mother to progeny. The virus infected females produced significantly fewere pupae compared to the controls during the experimental period (P<0.00001).

Prognosis of schistosomiasis mansoni patients infected with hepatitis B virus

A clinical study on the evolution of patients with schistosomiasis mansoni has been conducted since 1983 at the outpatient clinic of the Infectious and Parasitic Disease Service in the Clementino Fraga Filho University Hospital in Rio de Janeiro, Brazil, comparing prevalence of positive tests for HBsAg, anti-HBsAg, and anti-HBc among patients infected with Schistosoma mansoni coming from various regions of Brazil and with different clinical forms of the disease. A non-significant predominance of HBsAg, anti-HBsAg, and anti-HBc was detected among patients with the hepatosplenic form of schistosomiasis, who presented a more severe clinical evolution with a higher frequency of hematemesis and/or melena, in addition to the development of macronodular cirrhosis and a worse prognosis as compared to patients with the toxemic form, schistosomiasis-infection and the hepatointestinal form.

Risk Factors for Human T Cell Lymphotropic Virus Type I among Injecting Drug Users in Northeast Brazil: Possibly Greater Efficiency of Male to Female Transmission

It was observed in the city of Salvador, State of Bahia, the highest seroprevalence of human T cell lymphotropic virus type 1 (HTLV-I) infection in Brazil as demonstrated by national wide blood bank surveys. In this paper, we report results of an investigation of drug use and sexual behavior associated with HTLV-I infection among male and female injecting drug users (IDUs) in Salvador. A cross sectional study was conducted in the Historical District of Salvador from 1994-1996 (Projeto Brasil-Salvador) and 216 asymptomatic IDUs were selected using the snowball contact technique. Blood samples were collected for serological assays. Sera were screened for human immunodeficiency virus (HIV-1/2) and HTLV-I/II antibodies by ELISA and confirmed by Western blot. The overall prevalence of HTLV-I/II was 35.2% (76/216). The seroprevalence of HTLV-I, HTLV-II and HIV-1 was for males 22%, 11.3% and 44.1% and for females 46.2%, 10.3% and 74.4% respectively. HTLV-I was identified in 72.4% of HTLV positive IDUs. Variables which were significantly associated with HTLV-I infection among males included needle sharing practices, duration of injecting drug use, HIV-1 seropositivity and syphilis. Among women, duration of injecting drug use and syphilis were strongly associated with HTLV-I infection. Multivariate analysis did not change the direction of these associations. Sexual intercourse might play a more important role in HTLV-I infection among women than in men.

Comparison between Human Immunodeficiency Virus Positive and Negative Patients with Tuberculosis in Southern Brazil

The objective of this study is to determine the different characteristics of human immunodeficiency virus (HIV) positive and negative patients treated for tuberculosis (TBC) in a tertiary hospital in Southern Brazil. We conducted a retrospective cohort study over a 5-year period, from January 1992 through December 1996. We reviewed medical charts of patients from our institution who received TBC treatment. We reviewed 167 medical charts of patients with confirmed TBC. HIV positivity was detected in 74 patients. There were statistically significant difference between HIV positive and negative patients in sex and age. HIV-infected patients showed significantly more signs of bacteremia than HIV-negative patients. Extra-pulmonary TBC was present respectively in 13 (17.6%) and 21 (22.6%) HIV positive and negative patients. There was a significant difference between chest radiograph presentation in HIV positive and negative patients. There were significantly lower hematocrit, hemoglobin, leukocyte and lymphocyte levels in HIV-positive compared to HIV-negative patients. Outcome was significantly different in the two groups with a death rate of 36.5% and 10.8% in HIV-positive and in HIV-negative patients. The difference between HIV positive and negative patients may have importance in the diagnosis, management and prognosis of patients with TBC.

Antigenic and Genetic Characterization of Twenty-six Strains of Human Respiratory Syncytial Virus (Subgroup A) Isolated During Three Consecutive Outbreaks in Havana City, Cuba

Twenty-six human respiratory syncytial virus strains (subgroup A) isolated from three outbreaks in Havana City during the period 1994/95, 1995/96 and 1996/97 were analyzed to determine their antigenic and genetic relationships. Analyses were performed by monoclonal antibodies and restriction mapping (N gene) following amplification of the select region of the virus genome by polymerase chain reaction. All isolated strains were classified as subgroup A by monoclonal antibodies and they showed a restriction pattern NP4 that belonged to subgroup A. Thus the results obtained in this work, showed a close relation (100%) between antigenic and genetic characterization of the isolated strains in our laboratory. These methods permit the examination of large numbers of isolates by molecular techniques, simplifying the researchs into the molecular epidemiology of the virus.

Human Immunodeficiency Virus Seroprevalence among Inmates of the Penitentiary Complex of the Region of Campinas, State of São Paulo, Brazil

Six hundred and ninety three male inmates from three penitentiaries, two (A and B) maximum-security systems and one (C) minimum-security facility, located in Campinas, State of São Paulo, Brazil were studied for the presence of human immunodeficiency virus (HIV) antibodies, using a cross-sectional design. The search for anti-HIV antibodies in 693 samples of sera collected was carried out by two serological tests: (a) the Microparticle enzyme immunoassay-HIV-1 and HIV-2 (MEIA) (Abbott Laboratories) and (b) the Western Blot-HIV-1 (WB) (Cambridge Biotech Corporation) to confirm positive results with MEIA. Sera reactivity for HIV antibodies was 14.4%. The highest frequency of anti-HIV antibodies was found in the A and B maximum-security prisons: 17% and 21.5%, respectively. In prison C, the frequency of reagents was 10.9%. Seventy three inmates, initially negative in the MEIA test, were checked again five and seven months later. Three of them, all from the maximum-security facilities, became reactive in the MEIA test, with confirmation in the WB, suggesting that serological conversion had occurred after imprisonment.

Hepatitis C virus prevalence among an immigrant community to the Southern Amazon, Brazil

A community-based random survey was conducted in a southern Brazilian Amazonian county aiming to investigate hepatitis C virus (HCV) infection prevalence and the association of demographic variables and lifestyle behaviours. Seven hundred eighty individuals were serologically screened with a third generation enzyme-linked immunosorbent assay to detect anti-HCV antibodies between 1994/1995. Positive samples were retested for confirmation with a line immunoassay (LIA, Inno-LIA HCV Ab III). Most of these subjects were low income and came from southern Brazilian states (65.8). Two point four percent (IC 95% 1.2%- 4.6%) of the subjects had LIA-confirmed anti-HCV antibodies reactivity. The age-specific prevalence of HCV antibodies slightly increased with age, with the highest prevalence after the age of 40 years. The results of multivariate analysis indicate a strong association between HCV antibodies and previous surgery and history of intravenous drug use. There were no apparent association with gender, hepatitis B virus markers, blood transfusion, and sexual activity. Mean time living in Amazon did not differ between confirmed and negative anti-HCV individuals. The present data point out an intermediate endemicity of HCV infection among this immigrant community to the Amazon region and that few HCV infected participants presented known risk factors.

Frequency of CCR5 Delta-32 Mutation in Human Immunodeficiency Virus (HIV)-seropositive and HIV-exposed Seronegative Individuals and in General Population of Medellin, Colombia

Repeated exposure to human immunodeficiency virus (HIV) does not always result in seroconversion. Modifications in coreceptors for HIV entrance to target cells are one of the factors that block the infection. We studied the frequency of Delta-32 mutation in ccr5 gene in Medellin, Colombia. Two hundred and eighteen individuals distributed in three different groups were analyzed for Delta-32 mutation in ccr5 gene by polymerase chain reaction (PCR): 29 HIV seropositive (SP), 39 exposed seronegative (ESN) and 150 individuals as a general population sample (GPS). The frequency of the Delta-32 mutant allele was 3.8% for ESN, 2.7% for GPS and 1.7% for SP. Only one homozygous mutant genotype (Delta-32/Delta-32) was found among the ESN (2.6%). The heterozygous genotype (ccr5/Delta-32) was found in eight GPS (5.3%), in one SP (3.4%) and in one ESN (2.6%). The differences in the allelic and genotypic frequencies among the three groups were not statistically significant. A comparison between the expected and the observed genotypic frequencies showed that these frequencies were significantly different for the ESN group, which indirectly suggests a protective effect of the mutant genotype (Delta-32/Delta-32). Since this mutant genotype explained the resistance of infection in only one of our ESN persons, different mechanisms of protection must be playing a more important role in this population.

Hepatitis B virus exposure in human immunodeficiency virus seropositive Cuban patients

In order to estimate the prevalence of serological markers of exposure to Hepatitis B Virus (HBV), 295 subjects were selected at random from the National Registry of human immunodeficiency virus positive subjects. Evidence of exposure to HBV was defined as: testing Hepatitis B surface antigen (HBsAg) and anti-Hepatitis B core antigen (anti-HBc) positive or anti-HBc positive only. Overall, 133 (45.5%) were positive for anti-HBc and 15 (5.1%) resulted positive to HBsAg. Significant statistical association was found between male sex and exposure to HBV (p<0.01). Homosexual or bisexual behavior was found to be strongly associated to HBV exposure (p<0.001). In conclusion, the prevalence of HBV serological markers is higher in Cuban HIV positive subjects compared to the Cuban general population.