AN ELISA SUITABLE FOR THE DETECTION OF RABIES VIRUS ANTIBODIES IN SERUM SAMPLES FROM HUMAN VACCINATED WITH EITHER CELL-CULTURE VACCINE OR SUCKLING-MOUSE-BRAIN VACCINE

An indirect ELISA for determination of post-vaccination rabies antibody was applied. Purified rabies virus was used as antigen to coat plates, and staphylococcal protein A linked with horseradish peroxidase was used for detecting IgG antibody in human sera. Sera from humans, vaccinated with cell-culture vaccine or suckling-mouse-brain vaccine, were examined. ELISA results were compared to those obtained from the virus neutralization test. The mean and standard deviation of OD were determined for 126 negative sera (pre-vaccination) and for 73 sera from vaccinated persons showing antibody titers lower than 0.5 IU/ml. Results were defined as ELISA -positive, -negative or -doubtful. Establishment of a doubtful region reduced the number of sera otherwise classified as positive (false-positive sera). In this way, the sensitivity, specificity and agreement values were respectively 87.5%, 92.4% and 88.5%. No significant differences were observed in these values when the group vaccinated with cell-culture vaccine and the group vaccinated with suckling-mouse-brain vaccine were compared. It was shown that much of the disagreement between the values obtained by neutralization test and ELISA occurred in sera obtained at the beginning of the immunization process, and was probably due to the presence of IgM in the serum samples, detected only by the former test. This ELISA method can be used as a screening test in rabies laboratories regardless of the kind of vaccine used for immunization.

PREVALENCE OF HEPATITIS B AND HEPATITIS C MARKERS IN ALCOHOLICS WITH AND WITHOUT CLINICALLY EVIDENT HEPATIC CIRRHOSIS

We assessed the frequency of serological markers of hepatitis B virus (HBV) and hepatitis C virus (HCV) infections in 365 alcoholics by determining, by ELISA, the presence of HBsAg, anti-HBc, anti-HBs and anti-HCV. Fifty patients were cirrhotics and 315 had no evidence of hepatic cirrhosis; of the latter HBsAg was assessed in all, anti-HBc and anti-HBs in 130, and anti-HCV in 210. Among the alcoholics the frequencies of HBsAg (1.9%), anti-HBc (28.3%) and anti-HCV (3.8%) were higher (p<0.001) than among the controls (N=17,059), 0.4%, 4.0% and 0.4% respectively. The frequency of positive HBsAg was higher (p<0.001) in the cirrhotic patients (8.0%) than in alcoholics without cirrhosis (0.95%) and in controls (0.4%), and similar between the latter; of anti-HBc in alcoholics without cirrhosis (28.5%) was similar in cirrhotics patients (28.0%) and higher (p<0.001) than in the controls (4.0%); of anti-HBs in alcoholics without cirrhosis (20.8%) was similar to that of the cirrhotic patients (10.0%), and the anti-HCV was similar between alcoholics with (6.0%) and without cirrhosis (3.3%) and higher (p<0.001) than in controls (0.4%). We concluded that: a) alcoholics with or without cirrhosis have similar frequencies of infection with HBV and HCV between them, and higher than in nonalcoholics; b) alcoholics without cirrhosis had a frequency of HBV active infection (HBsAg+) which was similar to the controls, whereas among those who progressed to cirrhosis this frequency was significantly higher, what suggests that HBV may be implicated in the pathogenesis of cirrhosis in a few alcoholic individuals.

Chronic hepatitis C virus infections in Brazilian patients: association with genotypes, clinical parameters and response to long term alpha interferon therapy

The present study assessed the clinical significance of hepatitis C virus (HCV) genotypes and their influence on response to long term recombinant-interferon-alpha (r-IFN-a) therapy in Brazilian patients. One hundred and thirty samples from patients previously genotyped for the HCV and with histologically confirmed chronic hepatitis C (CH-C) were evaluated for clinical and epidemiological parameters (sex, age, time of HCV infection and transmission routes). No difference in disease activity, sex, age or mode and time of transmission were seen among patients infected with HCV types 1, 2 or 3. One hundred and thirteen of them were treated with 3 million units of r-IFN-a, 3 times a week for 12 months. Initial response (IR) was significantly better in patients with genotype 2 (100%) and 3 (46%) infections than in patients with genotype 1 (29%) (p < 0.005). Among subtypes, difference in IR was observed between 1b and 2 (p < 0.005), and between 1b and 3a (p < 0.05). Sustained response (SR) was observed in 12% for (sub)type 1a, 13% for 1b, 19% for 3a, and 40% for type 2; significant differences were found between 1b and 2 (p < 0.001), and between 1b and 3a (p < 0.05). Moreover, presence of cirrhosis was significantly associated with non response and response with relapse (p < 0.05). In conclusion, non-1 HCV genotype and lack of histological diagnosis of cirrhosis were the only baseline features associated with sustained response to treatment. These data indicate that HCV genotyping may have prognostic relevance in the responsiveness to r-IFN-a therapy in Brazilian patients with chronic HCV infection, as seen in other reports worldwide.

Prevalence of hepatitis B and C in the sera of patients with HIV infection in São Paulo, Brazil

The objective of this study was to evaluate the prevalence of hepatitis B and C viruses in a group of HIV infected patients, followed at a single institution since 1996. 1,693 HIV positive patients (1,162 male, 531 female) were tested for HBV infection. Virological markers for HBV included HBsAg and total anti-HBc by ELISA. 1,457 patients (1,009 male, 448 female) were tested for HCV infection. Detection of HCV antibodies was carried out by ELISA. A sample of HCV antibody positive patients was tested for HCV by PCR to confirm infection. Of 1,693 patients tested for HBV, 654 (38.6%) and 96 (5.7%) were anti-HBc and HBsAg positive, respectively. Of 1,457 patients tested for HCV, 258 (17.7%) were anti-HCV positive. 82 of these patients were also tested by PCR and 81 were positive (98%). Of 1,411 patients tested for HBV and HCV 26 (1.8%) were positive for both viruses.

Hepatitis C virus in monozygotic twins

A case of a pregnant patient with chronic hepatitis C who gave birth to monozygotic twins that were infected with HCV is reported. One of the newborns was positive for HCV-RNA in blood sample collected 12 hours after delivery. The other newborn was negative for HCV-RNA at birth, but was detected HCV viremia at three months of age. The results have led to the conclusion that one of the twins was probably contaminated in the intrauterine period, while the other acquired the infection in the perinatal period. Both were negative for HCV-RNA and for anti-HCV in the serum samples collected at nine months of age. The report describes the changes in the laboratory tests conducted in mother and twins until 29 months after delivery.

Predictive factors for response to Lamivudine in chronic hepatitis B

BACKGROUND: Lamivudine has been shown to be an efficient drug for chronic hepatitis B (CHB) treatment. AIM: To investigate predictive factors of response, using a quantitative method with high sensitivity. METHODS: We carried out a prospective trial of lamivudine in 35 patients with CHB and evidence for viral replication, regardless to their HBeAg status. Lamivudine was given for 12 months at 300 mg daily and 150 mg thereafter. Response was considered when DNA was undetectable by PCR after 6 months of treatment. Viral replication was monitored by end-point dilution PCR. Mutation associated with resistance to lamivudine was detected by DNA sequencing in non-responder patients. RESULTS: Response was observed in 23/35 patients (65.7%) but only in 5/15 (33.3%) HBeAg positive patients. Only three pre-treatment variables were associated to low response: HBeAg (p = 0.006), high viral load (DNA-VHB > 3 x 10(6) copies/ml) (p = 0.004) and liver HBcAg (p = 0.0028). YMDD mutations were detected in 7/11 non-responder patients. CONCLUSIONS: HBeAg positive patients with high viral load show a high risk for developing drug resistance. On the other hand, HBeAg negative patients show a good response to lamivudine even with high viremia.

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.

Hepatitis B and C virus markers among patients with hepatosplenic mansonic schistosomiasis

PURPOSE: To evaluate the frequency and the consequences of the co-infection of hepatitis B and C viruses in patients with hepatosplenic schistosomiasis (HSS). METHODS: B and C serologic markers, exposure to risk factors, biochemical assays, upper gastrointestinal endoscopies, and abdominal ultrasonograms were evaluated in 101 patients with HSS from 1994 to 1997. Whenever possible, PCR was tested and histopathological studies were reviewed. RESULTS: At least one HBV virus marker was found in 15.8%, and anti-HCV was detected in 12.9% of the subjects. The seropositive subjects tended to be older than the seronegative ones. A history of blood transfusion was significantly related to the presence of anti-HCV. Three (18.75%) out of 16 subjects exposed to B virus were HBsAg positive. Eleven (84.6%) out of thirteen patients who were anti-HCV positive demonstrated viral activity. Patients with ongoing viral infection presented a higher average level of liver aminotransferases, a higher frequency of cell decompensation and a higher rate of chronic hepatitis. Portal hypertension parameters were not influenced by viral exposure. CONCLUSIONS: The rate of hepatitis B and C viruses serologic markers observed in the patients with HSS was higher than the control group. The co-infection was responsible for a higher frequency of cell decompensation.

Hepatitis C virus seroprevalence and risk factors among patients with HIV infection

The objective of this study was to evaluate the prevalence and risk factors associated with HCV infection in a group of HIV seropositive patients. We analyzed the medical records of 1,457 patients. All patients were tested for HCV infection by third generation ELISA. Whenever possible, a sample of the positive patients was also tested for HCV by PCR. HCV positive patients were analyzed according to their risk factors for both infections. The prevalence of anti-HCV positive patients was 17.7% (258 patients). Eighty-two (82) of these patients were also tested by PCR and 81 were positive for HCV virus (98%). One hundred fifty-one (58.5%) were intravenous drug users (IDU); 42 (16.3%) were sexual partners of HIV patients; 23 (8.9%) were homosexual males; 12 (4.7%) had received blood transfusion; 61 (17.5%) had promiscuous sexual habits; 14 (5.4%) denied any risk factor; 12 (4.7%) were sexual partners of IDU. Two hundred four patients mentioned only one risk factor. Among them, 28 (10.9%) were sexual partners of HIV-positive patients. Although intravenous drug use was the most important risk factor for co-infection, sexual transmission seemed to contribute to the high HCV seroprevalence in this group of patients.

Sexual transmission of hepatitis C virus

In order to contribute to a better understanding of the forms of acquisition of hepatitis C virus (HCV) in Brazil, with special emphasis on sexual transmission, we determined the presence of HCV infection in regular partners and in non-sexual home communicants of blood donors seen at Fundação Pró-Sangue Hemocentro de São Paulo from January 1992 to July 1996. Of 154 blood donors with HCV infection (index cases), 111 had had regular partners for at least 6 months. Sixty-eight of 111 partners were evaluated for HCV infection. Of these, 8 (11.76%) were considered to have current or previous HCV infection; a history of sexually transmissible diseases and index cases with a positive HCV-RNA test were more prevalent among partners with HCV infection. Of the 68 index cases whose partners were studied, 56 had non-sexual home communicants. Of the total of 81 home communicants, 66 accepted to be evaluated for HCV infection. None of them was HCV-positive, suggesting that the high prevalence of HCV infection among partners may be attributed at least partially to sexual transmission.

Aseptic meningitis in a large MMR vaccine campaign (590,609 people) in Curitiba, Paraná, Brazil, 1998

The aseptic meningitis after Measles-Mumps-Rubella vaccine (MMR) is a well recognized complication, and different incidences have been observed in several studies. We retrospectively analyzed forty cases of aseptic meningitis, during a large public immunization campaign (1998) in Curitiba, Southern Brazil (590,609 people), admitted in our Service. The vaccine utilized was Leningrad-3-Zagreb mumps strain, Edmonston-Zagreb measles strain, and RA 27#3 rubella strain. In all county, a total number of 87 cases were reported, resulting in a incidence of 1.7 cases per 10,000 given doses . The mean age was 23.7 ± 12.8 years. The female:male ratio was 1.35:1. Severe headache with meningismus (92.5%), fever (87.5%), nausea/vomiting (82.5%) were the most common clinical findings. Three cases (7.5%) developed mild mumps. All patients underwent cerebrospinal fluid (CSF) tap with the following findings: mononuclear pleocytosis from 100 to 500 cells/mm³ in 17 cases (42.5%; 257.5 ± 260.6 cells/mm³); increased protein 28 cases (67.5%; 92.1 ± 76.9 mg/dL); glucose was normal in all cases (56.8 ± 11.2 mg/dL) except in 4 (10%) cases, which presented less than 44 mg/dL. All serological tests (latex to bacterial meningitis, Cryptococcus, cysticercosis, VDRL) and bacteriological cultures were negative. Virus identification were also negative in 8 samples. None of the patients had neurological deficits or related symptoms after one year of onset. We believe the benefit of vaccination clearly outweights the incidence of benign vaccine-associated meningitis.