Correlation between Apoptosis and in Situ Immune Response in Fatal Cases of Microcephaly Caused by Zika Virus.

Zika virus (ZIKV) is a single-stranded positive-sense RNA flavivirus that possesses a genome approximately 10.7 Kb in length. Although pro-inflammatory and anti-inflammatory cytokines and apoptotic markers belonging to the extrinsic and intrinsic pathways are suggested to be involved in fatal cases of ZIKV-induced microcephaly, their exact roles and associations are unclear. To address this, brain tissue samples were collected from 10 individuals, five of whom were diagnosed as ZIKV positive with microcephaly and a further five were flavivirus-negative controls that died because of other causes. Examination of material from the fatal cases of microcephaly revealed lesions in the cerebral cortex, edema, vascular proliferation, neuronal necrosis, gliosis, neuronophagy, calcifications, apoptosis, and neuron loss. The expression of various apoptosis markers in the neural parenchyma, including FasL, FAS, BAX, BCL2, and caspase 3 differed between ZIKV-positive cases and controls. Further investigation of type 1 and 2 helper T-cell cytokines confirmed a greater anti-inflammatory response in fatal ZIKV-associated microcephaly cases. Finally, an analysis of the linear correlation between tumor necrosis factor-α, IL-1ß, IL-4, IL-10, transforming growth factor-ß, and IL-33 expression and various apoptotic markers suggested that the immune response may be associated with the apoptotic phenomenon observed in ZIKV-induced microcephaly.

In situ immune response and mechanisms of cell damage in central nervous system of fatal cases microcephaly by Zika virus.

Zika virus (ZIKV) has recently caused a pandemic disease, and many cases of ZIKV infection in pregnant women resulted in abortion, stillbirth, deaths and congenital defects including microcephaly, which now has been proposed as ZIKV congenital syndrome. This study aimed to investigate the in situ immune response profile and mechanisms of neuronal cell damage in fatal Zika microcephaly cases. Brain tissue samples were collected from 15 cases, including 10 microcephalic ZIKV-positive neonates with fatal outcome and five neonatal control flavivirus-negative neonates that died due to other causes, but with preserved central nervous system (CNS) architecture. In microcephaly cases, the histopathological features of the tissue samples were characterized in three CNS areas (meninges, perivascular space, and parenchyma). The changes found were mainly calcification, necrosis, neuronophagy, gliosis, microglial nodules, and inflammatory infiltration of mononuclear cells. The in situ immune response against ZIKV in the CNS of newborns is complex. Despite the predominant expression of Th2 cytokines, other cytokines such as Th1, Th17, Treg, Th9, and Th22 are involved to a lesser extent, but are still likely to participate in the immunopathogenic mechanisms of neural disease in fatal cases of microcephaly caused by ZIKV.

Oral live attenuated human rotavirus vaccine (RotarixTM) offers sustained high protection against severe G9P[8] rotavirus gastroenteritis during the first two years of life in Brazilian children

In a large Phase III trial conducted in 10 Latin American countries, the safety and efficacy of the live attenuated monovalent rotavirus vaccine RIX4414 was evaluated in 15,183 healthy infants followed up during the first two years of life. Belém was the only site in Brazil included in this multicentre trial. The study in Belém included a subset of 653 infants who were followed up until 24 months of age for protection against severe rotavirus gastroenteritis. These subjects were randomly assigned in a 1:1 ratio to receive two doses of vaccine (n = 328) or two doses of placebo (n = 325) at approximately two and four months of age. Of the 653 enrolled infants, 23 dropped out during the study period. For the combined two-year period, the efficacy of RIX4414 was 72.3% [95% confidence interval (CI) 37.5-89.1%] against severe rotavirus-related gastroenteritis, reaching a protection rate of 81.8% (95% CI 36.4-96.6%) against circulating wild-type G9 rotavirus strains. It is concluded that two doses of RIX4414 are highly efficacious against severe rotavirus gastroenteritis in Belém during the first two years of life and provide high protection against the worldwide emergence and spread of G9P[8] strains.

Segurança, imunogenicidade e eficácia protetora de duas doses da vacina RIX4414 contendo rotavírus atenuado de origem humana / Safety, immunogenicity, and protective efficacy of two doses of RIX4414 live attenuated human rotavirus vaccine in healthy Brazilian infants

OBJETIVO: Determinar a segurança, imunogenicidade e eficácia de duas doses da vacina contra o rotavírus em lactentes brasileiros saudáveis. MÉTODOS: Foi realizado um estudo randomizado, multicêntrico, duplo-cego e controlado por placebo no Brasil, México e Venezuela. Os lactentes receberam duas doses orais de vacina ou placebo aos 2 e 4 meses de idade, juntamente com as imunizações de rotina, exceto a vacina oral contra poliomielite (VOP). O presente estudo relata apenas os resultados obtidos em Belém, Brasil, onde o número de indivíduos por grupo e os títulos da vacina viral foram os seguintes: 194 (104,7 unidades formadoras de focos - UFF), 196 (10(5,2) UFF), 194 (10(5,8) UFF) e 194 (placebo). A resposta de anticorpos anti-rotavírus (anti-RV) foi avaliada em 307 indivíduos. A gravidade clínica dos episódios de gastroenterite (GE) foi determinada através de um escore com 20 pontos, onde um valor > 11 foi considerado como GE grave. RESULTADOS: As taxas de sintomas gerais solicitados foram semelhantes tanto nos indivíduos que receberam a vacina como naqueles a quem se administrou placebo. Aos 2 meses após a segunda dose, ocorreu resposta em termos de IgA sérica para RV em 54,7 a 74,4 por cento dos vacinados. Não houve interferência na imunogenicidade das vacinas de rotina. A eficácia da vacina contra qualquer gastroenterite por rotavírus (GERV) foi de 63,5 por cento (IC95 por cento 20,8-84,4) para a maior concentração (10(5,8) UFF). A eficácia foi de 81,5 por cento (IC95 por cento 44,5-95,4) contra GERV grave. Em sua maior concentração (10(5,8) UFF), a RIX4414 conferiu uma proteção de 79,8 por cento (IC95 por cento 26,4-96,3) contra GERV grave causada pela amostra G9. CONCLUSÕES: A RIX4414 foi altamente imunogênica com baixa reatogenicidade, e não interferiu na resposta sérica à difteria, tétano, coqueluche, hepatite B e antígenos Hib. Duas doses da RIX4414 conferiram proteção significativa contra a GE grave causada pelo RV.

OBJECTIVE: To determine the safety, immunogenicity and efficacy of two doses of rotavirus vaccine in healthy Brazilian infants. METHODS: A randomized, multicenter, double-blind, placebo-controlled trial was conducted in Brazil, Mexico and Venezuela. Infants received two oral doses of vaccine or placebo at 2 and 4 months of age, concurrently with routine immunizations, except for oral poliomyelitis vaccine (OPV). This paper reports results from Belém, Brazil, where the number of subjects per group and the viral vaccine titers were: 194 (10(4.7) focus forming units - FFU), 196 (10(5.2) FFU), 194 (10(5.8) FFU) and 194 (placebo). Anti-rotavirus (anti-RV) antibody response was assessed in 307 subjects. Clinical severity of gastroenteritis episodes was measured using a 20-point scoring system with a score of > 11 defined as severe GE. RESULTS: The rates of solicited general symptoms were similar in vaccine and placebo recipients. At 2 months after the second dose, a serum IgA response to RV occurred in 54.7 to 74.4 percent of vaccinees. No interference was seen in the immunogenicity of routine vaccines. Vaccine efficacy against any rotavirus gastroenteritis (RVGE) was 63.5 percent (95 percentCI 20.8-84.4) for the highest concentration (10(5.8) FFU). Efficacy was 81.5 percent (95 percentCI 44.5-95.4) against severe RVGE. At its highest concentration (10(5.8) FFU), RIX4414 provided 79.8 percent (95 percentCI 26.4-96.3) protection against severe RVGE by G9 strain. CONCLUSIONS: RIX4414 was highly immunogenic with a low reactogenicity profile and did not interfere with seroresponse to diptheria, tetanus, pertussis, hepatitis B and Hib antigens. Two doses of RIX4414 provided significant protection against severe GE caused by RV.

Human herpesvirus-7 as a cause of exanthematous ilnesses in Belém, Pará, Brazil

Examinamos soros de 370 pacientes acometidos de doença exantemática, selecionados em Belém, norte do Brasil, com o propósito de se detectarem anticorpos IgM e IgG para o herpesvírus humano-7 (HHV-7). As amostras foram obtidas entre janeiro de 1996 e dezembro de 2002 e, posteriormente, processadas utilizando-se a técnica da imunofluorescência indireta (IFI). Taxas de anticorpos IgM e/ou IgG foram encontradas em 190 (51,4) desses pacientes. Observamos taxas de prevalência similares para os sexos feminino e masculino com: 52,5 e 50,3, respectivamente. O "status" sorológico foi definido pela presença de anticorpos IgG nos espécimes de 135 (36,5) pacientes. A par disso, em 55 (14,9) dos 370 pacientes foram detectados anticorpos IgM para o HHV-7. Taxas de anticorpos IgM e IgG para o HHV-7 foram similares (p > 0.05) quando comparamos indivíduos do sexo feminino e masculino: 14,4 versus 15,3 e 38,1 versus 35,0, respectivamente. Diferença estatisticamente significativa (p = 0,003) foi observada quando comparamos as taxas de anticorpos IgM para o HHV-7 nos indivíduos do grupo etário de 5-8 meses pertencentes ao sexo feminino e masculino. Taxas de prevalência variando de 4,6 (masculino, 5-8 meses de idade) a 93,3 (feminino, > 10 anos de idade) e 12,2 (masculino, 5-8 meses de idade) a 80,0 (masculino, 8-10 anos de idade) foram observadas no subgrupo positivo para IgG. Um subgrupo (n = 131) de pacientes com anticorpos IgM ou IgG foi examinado quanto a presença de DNA para o HHV-7 pela técnica da reação em cadeia da polimerase/ "nested" PCR. Infecção recente/ativa para o HHV-7 ocorreu em 11,0 (6/55) dos pacientes cujas amostras apresentaram anticorpos IgM específicos para o HHV-7. Em um subgrupo (n = 76) de pacientes com altos níveis de anticorpos IgG para o HHV-7 (título > 1: 160) não foi detectada a presença de DNA em seus soros pelo PCR/ "nested" PCR. Entre as seis infecções recentes/ativas, quatro indivíduos com menos de um ano e dois com 3 e 6 anos de idade apresentaram típico exantema súbito (E.S) definido por febre elevada (> 38,0 ºC) com duração de 24 a 72 horas, acompanhando-se de erupção cutânea maculopapular. Nossos resultados ressaltam a necessidade de procurarmos a infecção pelo HHV-7 em pacientes portadores de doença exantemática, particularmente naquelas apresentações típicas de E.S. O HHV-7 parece emergir como um novo patógeno associado a quadros exantemáticos em nossa região.

Rotavirus G serotypes and p[],G genotypes identified in cases of reinfection among children participating in a trial with rhesus-human reassortant tetravalent vaccine (RRV-TV) in Belém, Brazil.

Group A rotaviruses are the most important agents of severe diarrhea in children and infants worldwide. The aim of present study was to identify rotavirus G serotypes and P[],G genotypes in cases of reinfection among children who participated in a vaccine trial with the tetravalent rhesus-human reassortant rotavirus vaccine (RRV-TV 4 x 10(4) pfu/dose) in Belém, Brazil. From July 1990 to June 1992, 540 children received, at their first, third and fifth months of life, oral doses of either vaccine or placebo. A total of 90 rotavirus diarrheal episodes among children who completed the three-dose vaccination schedule were recorded. We studied 11 reinfection rotavirus cases among five children (three female and two male). Fecal specimens were tested by using a enzyme immunoassay (IDEIA Rotavirus), followed by EIA with monoclonal antibodies to determine infecting serotypes Gl, G2, G3 and G4 and subgroups I and II. The viral dsRNA was extracted and electrophoresed through polyacrylamide gel and then subjected to reverse-transcription-polymerase chain reaction and nested-PCR for the determination of Gl, G2, G3, G4, G5 and G9 and P[4], P[6], P[8] and P[9] rotavirus genotypes. A total of 11 cases of reinfection (12 per cent) occurred among five children, three from the placebo group and two from the vaccine group. In four of the cases of reinfection G serotypes and P[],G genotypes were as follows: for the first and second infections, respectively: (1) G2/P[4],G2 and Gl/P[4],G1; (2) G2/P[4],G2 and G2/P[6],G5; (3) G2/P[4],G2 and G1/P[8],G1; and (4) G2/P[8],G1 and G1/P[8],G1. A fifth child had three successive infections caused by serotypes/genotypes G1/P[8],G1, in the first and second infections, and G2/P[4],G2 in the third infection. The common genotypes and unusual genotypes were detected in 8 (73 per cent) and 3 (27 per cent) of the isolates, respectively. With regards to the clinical severity, in two children a score indicated moderate/severe disease in both first and second infections. One child had three successive infections; the first episode was moderate/severe, the second very severe and the third was not available. In contrast, in two other children, the first episode was very severe, and the second episode was moderate/severe in one child and data was not available for the other child. The results obtained in the present investigation underscore the need to broaden our knowledge of the immunity in rotavirus reinfection. This should be useful regarding future rotavirus vaccination strategies in Brazil.

Nutritional status in relation to the efficacy of the rhesus-human reassortant, tetravalent rotavirus vaccine (rrv-tv) in infants from Belém, Para state, Brazil

The rhesus-human reassortant, tetravalent rotavirus vaccine (RRV-TV) was licensed for routine use in the United States of America but it was recently withdrawn from the market because of its possible association with intussusception as an adverse event. The protective efficacy of 3 doses of RRV-TV, in its lower-titer (4 x 10(4) pfu/dose) formulation, was evaluated according to the nutritional status of infants who participated in a phase III trial in Belém, Northern Brazil. A moderate protection conferred by RRV-TV was related to weight-for-age Z-scores (WAZ) greater than -1 only, with rates of 38 percent (p = 0.04) and 40 percent (p = 0.04) for all- and- pure rotavirus diarrhoeal cases, respectively. In addition, there was a trend for greater efficacy (43 percent, p = 0.05) among infants reaching an height-for-age Z-score (HAZ) of > -1. Taking WAZ, HAZ and weight-for-height Z-score (WHZ) indices <= -1 together, there was no significant protection (p > 0.05) if both placebo and vaccine groups are compared. There was no significant difference if rates of mixed and pure rotavirus diarrhoeal cases are compared in relation to HAZ, WAZ and weight-for-height Z-score (WHZ) indices. Although a low number of malnourished infants could be identified in the present study, our data show some evidence that malnutrition may interfere with the efficacy of rotavirus vaccines in developing countries

Epidemiology of rotavirus gastroenteritis in the Amazon region and the need for a specific vaccine

In the Amazon region, rotaviruses account for at least 30 per cent of all episodes of acute gastroenteritis among hospitalized children and are associated with nearly 1O per cent of cases of infantile acute diarrhea at community level. All four rotavirus serotypes are shown to infect children in our region, serotype l being predominant (about 50 per cent). Sequential infections in the same child, caused by different serotypes, are commonly noted. No clear seasonal variation on the occurrence of rotavirus diarrhea has been recorded, as cases are readily detected throughout the year. Rotavirus diarrhea cases have been found to be, in general, more severe than those of other aetiology. On the other hand, it has been noted that early (children less than 4 months of age) rotavirus infections are more likely to be asymptomatic (p = 0.021). Occurrence of rotavirus infections among Amazonian Indian populations seems to be very common. An explosive outbreak of rotavirus diarrhea affected possibly 88 per cent of both children and adults of the Tiryió population, Northern Pará State. In addition, rotavirus antibody was detected in 54.7 per cent of 1,299 sera collected from Amerinds belonging to 13 relatively isolated communities in the Amazon region. In the light of the above mentioned findings it was suggested that our region would be suitable for a field trial with a rotavirus-candidate vaccine. A study is therefore underway aiming to compare safety, immunogenicity and efficacy of a rhesus-human reassortant rotavirus (RRV-tetravalent) vaccine and placebo in 500 healthy infants living in the peripheral area of Belém.