Early human enterovirus infections in healthy Swedish children participating in the PRODIA pilot study.

Human enteroviruses (HEV) are common, especially in childhood and during the enterovirus season, causing mainly asymptomatic infections but also mild and severe illnesses. Numerous studies have shown the association between HEV infections and type 1 diabetes. Here, the prevalence of HEV infections was studied in healthy Swedish children with increased HLA-associated risk for type 1 diabetes participating in the PRODIA pilot study in which children were randomized to receive probiotics or placebo during the first 6 months of life. Stool specimens collected from 197 children in every 3 months from the age of 3 to 24 months were screened for HEV using traditional viral culturing method and identified with reverse transcriptase polymerase chain reaction (RT-PCR) and sequencing of the partial VP1 coding part of the viral genome. Altogether 4.8% (52/1,094) of the specimens were HEV-positive and 22.3% (44/197) of the children excreted HEV during the follow-up. HEV-A and HEV-B were present in 2.1 and 2.7% of the specimens, respectively. HEV-C and HEV-D viruses were not detected. In total, 17 different HEV serotypes were detected and the most common findings were CV-A9 (13.5%), CV-A16 (11.5%), and CV-A2 (9.6%). The majority of the infections (92.3%) were during the enterovirus season extending from July to December. Probiotic treatment did not affect significantly the risk of HEV infections during the 2-year follow-up although a trend for transient decrease for HEV positivity (HEV-A and/or HEV-B) by the age of 12 months was observed in children who received probiotics [OR 0.40; 95% confidence interval 0.15 to 1.08; P-value 0.071, generalized estimating (GEE) analysis]. According to the results, HEV-A findings were nearly as common as HEV-B findings among the healthy children participating in this study. Also it was shown that serotypes belonging to HEV-A species can be detected by means of viral culturing.

Comparison of poliovirus recombinants: accumulation of point mutations provides further advantages.

The roles of recombination and accumulation of point mutations in the origin of new poliovirus (PV) characteristics have been hypothesized, but it is not known which are essential to evolution. We studied phenotypic differences between recombinant PV strains isolated from successive stool specimens of an oral PV vaccine recipient. The studied strains included three PV2/PV1 recombinants with increasing numbers of mutations in the VP1 gene, two of the three with an amino acid change I-->T in the DE-loop of VP1, their putative PV1 parent and strains Sabin 1 and 2. Growth of these viruses was examined in three cell lines: colorectal adenocarcinoma, neuroblastoma and HeLa. The main observation was a higher growth rate between 4 and 6 h post-infection of the two recombinants with the I-->T substitution. All recombinants grew at a higher rate than parental strains in the exponential phase of the replication cycle. In a temperature sensitivity test, the I-->T-substituted recombinants replicated equally well at an elevated temperature. Complete genome sequencing of the three recombinants revealed 12 (3), 19 (3) and 27 (3) nucleotide (amino acid) differences from Sabin. Mutations were located in regions defining attenuation, temperature sensitivity, antigenicity and the cis-acting replicating element. The recombination site was in the 5' end of 3D. In a competition assay, the most mutated recombinant beat parental Sabin in all three cell lines, strongly suggesting that this virus has an advantage. Two independent intertypic recombinants, PV3/PV1 and PV3/PV2, also showed similar growth advantages, but they also contained several point mutations. Thus, our data defend the hypothesis that accumulation of certain advantageous mutations plays a key role in gaining increased fitness.