Hb A1c in relation to intrauterine growth among male adolescents in southern Brazil.

The fetal origins hypothesis states that nutritional deprivation in utero affects fetal development and contributes to the incidence of diseases associated with the metabolic syndrome in later life. This study investigated whether haemoglobin (Hb) A(1c), an indicator of blood glucose, varied among healthy male adolescents according to their fetal growth rate, in a middle-income setting. Participants were men aged 18 years, belonging to the 1982 Pelotas birth cohort. Complete data, including gestational age and Hb A(1c) at age 18 years, were available for 197 individuals. There was an inverse association between mean Hb A(1c) and birthweight for the gestational age, but not birthweight alone. The association remained significant after adjustment for family income and mother's education, as well as for body mass index at 18 years (P for trend=0.01 and 0.03, respectively).

Glycosylation and secretion of yellow fever virus nonstructural protein NS1.

The flavivirus genome codes for structural and nonstructural proteins which are produced from a common polyprotein precursor by proteolytic processing. The nonstructural protein NS1 is hypothesized to be involved in virus assembly and maturation as well as in protective immunity. This paper describes the synthesis of several forms of yellow fever (YF) virus NS1 protein during virus cultivation in vitro. These include cell-associated and secreted NS1 forms, the major difference among these being their N-linked glycosylation status. High-mannose and complex (or hybrid)-type moieties were identified on YF NS1 by analysis with endoglycosidases of known specificity. These studies are the basis for a systematic approach to determining the role of NS1 in the virus cycle and possibly in pathogenesis.

Complete nucleotide sequence of yellow fever virus vaccine strains 17DD and 17D-213.

The complete nucleotide sequence of the genome from two yellow fever (YF) virus vaccine strains, 17DD and 17D-213, has been determined. Comparison of these sequences with those of other YF viruses including the parental virulent Asibi strain allowed the identification of 48 nucleotide sequence differences which are common to all 17D substrains. This is a significant reduction from the 67 nucleotide changes originally reported as being 17D-specific and potentially related to viral attenuation. The 48 changes are scattered throughout the genome, 26 of which are silent and 22 led to amino acid substitutions. These 22 changes are bona fide candidates to test by mutating the infectious YF cDNA to investigate their role in viral attenuation.

[The use of pacifiers in children: fecal contamination and association with diarrhea]. / Uso de chupeta em crianças: contaminação fecal e associação com diarréia.

A cross-sectional study of 354 children under two years of age was carried out in two periurban slums, with poor sanitary and socioeconomic conditions, located in Pelotas, southern Brazil. Most (79%) of the children studied were current users of pacifiers, 15% had never used one and the remaining 6% were ex-users. Among current users, 38% sucked a pacifier most of the time ("constant users"). Of the pacifiers in constant use, 93% were cultured for evidence of fecal contamination. Fecal coliforms were present in 49% of these. Diarrhoea was reported in 35% of all the children in the two weeks preceding the survey. Among constant pacifiers users, 40% had had diarrhoea in the preceding fortnight; this proportion was 32% for occasional users and 37% for non-users. These differences were not statistically significant.

[The use of tapioca as coverage in viral titration]. / O uso da tapioca como cobertura na titulação viral.

Virus titration is an important step required on viral vaccines quality control. "Plaque assay", which employs several types of overlay media, is usually used on viral titrations. In this paper we describe the use of Tapioca as an overlay media. Firstly, the toxicity of Tapioca was tested on Vero cells inoculated or not with the Yellow Fever virus (YF) 17DD vaccine strain. Secondly, different batches of the 17DD virus using the Tapioca and Karaya gum as the overlay on Vero cells were tested when higher titres were obtained using Tapioca. Tapioca was also shown to be a suitable overlay to be used in thermostability and plaque reduction neutralization tests. Other systems could benefit from the use of Tapioca as an overlay, since it was possible to titer Measles virus in Vero cells. Tapioca is a cheap Brazilian product, is locally available, easy to use, and reliable. Its use is suggested.

Genetic variability among yellow fever virus 17D substrains.

The complete nucleotide sequence of the genome from two yellow fever (YF) virus strains, 17DD and 17D-213 was determined. Comparison of these sequences with those of other YF viruses, including the parental virulent Asibi strain, allowed the identification of 48 nucleotide sequence differences which are 17D strain-specific and potentially related to viral attenuation. Another 43 nucleotide sequence differences were not common to all 17D substrains and are therefore substrain specific. Of the 21 changes between 17DD and Asibi 15 only five led to amino acid substitutions whereas 13 substrain differences common to all 17D-204 substrains produced six amino acid substitutions. Since the exact passage histories of these viruses is known it was possible to calculate, for each strain, the number of accumulated changes per passage. Based on these data the 17DD strain was the most genetically stable virus.

Molecular analysis of yellow fever virus 17DD vaccine strain.

The Oswaldo Cruz Foundation produces most of the yellow fever (YF) vaccine prepared worldwide. As part of a broader approach to determine the genetic variability in YF 17D seeds and vaccines and its relevance to viral attenuation the 17DD virus was purified directly from chick embryo homogenates which is the source of virus used for vaccination of millions of people in Brazil and other countries for half a century. Neutralization and hemagglutination tests showed that the purified virus is similar to the original stock. Furthermore, radioimmune precipitation of 35S-methionine-labeled viral proteins using mouse hyperimmune ascitic fluid revealed identical patterns for the purified 17DD virus and the YF 17D-204 strain except for the 17DD E protein which migrated slower on SDS-PAGE. This difference is likely to be due to N-linked glycosylation. Finally, comparison by northern blot hybridization of virion RNAs of purified 17DD with two other strains of YF 17D virus revealed only genome-sized molecules for all three viruses. These observations suggest that the vaccine phenotype is primarily associated with the accumulation of mutations.

The early use of yellow fever virus strain 17D for vaccine production in Brazil--a review.

The use of yellow fever (YF) virus 17D strain for vaccine production adapted in Brazil since its introduction in 1937 was reviewed. This was possible due to the availability of official records of vaccine production. The retrieved data highlight the simultaneous use of several serially passaged 17D substrain viruses for both inocula and vaccine preparation that allowed uninterrupted production. Substitution of these substrain viruses became possible with the experience gained during quality control and human vaccination. Post-vaccinal complications in humans and the failure of some viruses in quality control tests (neurovirulence for monkeys) indicated that variables needed to be reduced during vaccine production, leading to the development of the seed lot system. The 17DD substrain, still used today, was the most frequently used substrain and the most reliable in terms of safety and efficacy. For this reason, it is possible to derive an infectious cDNA clone of this substrain combined with production in cell culture that could be used to direct the expression of heterologous antigens and lead to the development of new live vaccines.

Heterogeneity in envelope protein sequence and N-linked glycosylation among yellow fever virus vaccine strains.

We have compared the deduced envelope (E) protein sequences of two biologically well-characterized yellow fever (YF) virus vaccine strains. The 17DD strain has been produced in Brazil for more than 50 years and used to successfully vaccinate millions of people worldwide. The 17D-213 is a candidate vaccine strain produced in tissue culture which has previously passed the monkey neurovirulence assay for testing human YF vaccines. Nucleotide sequence analysis of polymerase chain reaction-amplified cDNA revealed a number of mutations which were strain- and substrain-specific. A major difference of 17DD and 17D-213 as compared to 17D-204 and Asibi was the existence of a potential N-linked glycosylation site located at amino acid residues 153 and 151 of 17DD and 17D-213, respectively. These acceptor sites are apparently utilized for the addition of high-mannose carbohydrate chains as shown by endoglycosidase analyses of immunoprecipitated E proteins. Glycosylated E protein is also used to assemble YF vaccine virions. This work and eventual complete nucleotide sequence analysis of both vaccine strains should help to define possible changes involved in YF virus attenuation and allow their biological importance to be determined using a recently developed system for generating YF virus from cDNA. In addition, these data provide an estimate on the extent of genetic variability among YF 17D seeds and vaccines.