Association of serum 25-hydroxyvitamin D concentration with HLA-B, -DRB1 and -DQB1 genetic polymorphisms.

BACKGROUND/OBJECTIVES: The human leukocyte antigen (HLA) gene region associates with the risk for several autoimmune diseases, including type 1 diabetes. An association between vitamin D deficiency and several autoimmune diseases has been suggested. We tested the association between serum 25-hydroxyvitamin D (25OHD) concentrations and HLA alleles in pregnant Finnish women. SUBJECTS/METHODS: HLA-B (n=395), HLA-DRB1 (n=501) and HLA-DQB1 (n=475) alleles were genotyped in pregnant women (mothers of children who later developed type 1 diabetes and mothers of non-diabetic children). HLA-B alleles were divided into supertypes that share similar peptide-binding specificity. Serum 25OHD concentration had been previously measured in these women from sera collected during the first trimester of pregnancy. Multiple testing was controlled for using the false discovery rate method. RESULTS: An association was found between 25OHD concentration and HLA-B44 supertype (P=0.009); women with HLA-B44 supertype (B*18, B*37, B*40 and B*44 alleles) had lower 25OHD concentrations. No association was found between HLA-DRB1 or -DQB1 alleles and 25OHD concentration. CONCLUSIONS: In this study we found for the first time an association between HLA genetic polymorphisms and 25OHD concentration. In future studies, the mechanistic background of this association and the role of vitamin D in the regulation of HLA gene expression should be investigated.

Serum 25-hydroxyvitamin D level during early pregnancy and type 1 diabetes risk in the offspring.

AIMS/HYPOTHESIS: Vitamin D deficiency during the fetal period or infancy is one of the suggested environmental factors for type 1 diabetes and for its increasing incidence. To test this hypothesis we compared serum 25-hydroxyvitamin D (25(OH)D) levels during early pregnancy in mothers of children who subsequently developed type 1 diabetes (case mothers) with mothers of non-diabetic healthy children (control mothers) of the same age. METHODS: Children with type 1 diabetes were identified from the nationwide prescription register. 25(OH)D concentration was measured from serum samples collected during the first trimester of pregnancy from all Finnish women (Finnish Maternity Cohort). A total of 343 case mothers and 343 control mothers were included in the study. Samples were collected throughout the year. Samples from case and control mothers were matched on the day of collection. RESULTS: Mean 25(OH)D levels in case mothers (43.9 nmol/l) and control mothers (43.7 nmol/l) were not different. Of all mothers, 481 (70.1%) were vitamin D-deficient or -insufficient. CONCLUSIONS/INTERPRETATION: No difference was found in serum 25(OH)D concentrations during first trimester of pregnancy between mothers whose children later on developed type 1 diabetes, and mothers of non-diabetic ' healthy' children of the same age. It is difficult to detect possible effects of mothers' vitamin D deficiency during early pregnancy on the development of type 1 diabetes in the offspring in this population, as such a large proportion of mothers were vitamin D-deficient or -insufficient.

High frequency of cross-reacting antibodies against 2009 pandemic influenza A(H1N1) virus among the elderly in Finland.

Since May 2009, the pandemic influenza A(H1N1) virus has been spreading throughout the world. Epidemiological data indicate that the elderly are underrepresented among the ill individuals. Approximately 1,000 serum specimens collected in Finland in 2004 and 2005 from individuals born between 1909 and 2005, were analysed by haemagglutination-inhibition test for the presence of antibodies against the 2009 pandemic influenza A(H1N1) and recently circulating seasonal influenza A viruses. Ninety-six per cent of individuals born between 1909 and 1919 had antibodies against the 2009 pandemic influenza virus, while in age groups born between 1920 and 1944, the prevalence varied from 77% to 14%. Most individuals born after 1944 lacked antibodies to the pandemic virus. In sequence comparisons the haemagglutinin (HA) gene of the 2009 pandemic influenza A(H1N1) virus was closely related to that of the Spanish influenza and 1976 swine influenza viruses. Based on the three-dimensional structure of the HA molecule, the antigenic epitopes of the pandemic virus HA are more closely related to those of the Spanish influenza HA than to those of recent seasonal influenza A(H1N1) viruses. Among the elderly, cross-reactive antibodies against the 2009 pandemic influenza virus, which likely originate from infections caused by the Spanish influenza virus and its immediate descendants, may provide protective immunity against the present pandemic virus.

A population-specific diabetogenic haplotype HLA-A2,Cw1,B56,DR4,DQ8 is associated with high birthweight in Finnish diabetic families.

Children with type 1 diabetes (T1D) susceptibility HLA genotypes are shown to have an increased birthweight. We investigated to what extent T1D-predisposing HLA haplotypes were associated with increased birthweight. A total of 1255 Finnish children comprising those with T1D and their non-diabetic siblings were investigated. A total of 342 children and their non-diabetic parents were HLA genotyped. Birthweight data were obtained from the national Medical Birth Registry. The population-specific diabetogenic haplotype HLA-A2,Cw1,B56,DR4,DQ8 was associated with high birthweight (P=0.0280) in families with a diabetic offspring. Other T1D-predisposing HLA haplotypes showed nonsignificant tendency with high birthweight. More infants with a birthweight >or=4000 g were born in families with a T1D offspring than in the general Finnish population (P=0.0139). The previously observed direct association between birthweight and T1D risk may be mediated through the modulating effects that T1D susceptibility HLA genes have on weight. High birthweight and subsequent weight gain may accelerate the ongoing pancreatic autoimmune process in genetically susceptible individuals. The high proportion of infants having a birthweight >or=4000 g in families with a diabetic offspring raises a concern of potential adverse health outcomes that high birthweight can have.

Facilitation of performance in a working memory task with rTMS stimulation of the precuneus: frequency- and time-dependent effects.

Although improvements in performance due to TMS have been demonstrated with some cognitive tasks, performance improvement has not previously been demonstrated with working memory tasks. In the present study, a delayed match-to-sample task was used in which repetitive TMS (rTMS) at 1, 5, or 20 Hz was applied to either left dorsolateral prefrontal or midline parietal cortex during the retention (delay) phase of the task. Only 5 Hz stimulation to the parietal site resulted in a significant decrease in reaction time (RT) without a corresponding decrease in accuracy. This finding was replicated in a second experiment, in which 5 Hz rTMS at the parietal site was applied during the retention phase or during presentation of the recognition probe. Significant speeding of RT occurred in the retention phase but not the probe phase. This finding suggests that TMS may improve working memory performance, in a manner that is specific to the timing of stimulation relative to performance of the task, and to stimulation frequency.

HLA-patterns in patients with multiple sclerosis and type I diabetes mellitus: evidence for possible mutual exclusion of both diseases.

BACKGROUND: Type I diabetes mellitus (T1DM) and multiple sclerosis (MS), both immune-mediated diseases, rarely co-exist in the same individual or co-segregate in families. HLA susceptibility genes for T1DM (DRB1*0401, DRB1*0404, DQB1*0302, DRB1*0301, DQB1*0201) rarely occur in MS patients. HLA genes known to confer "resistance" to T1DM (DRB1*1501, DQB1*0602-DQA1*0102) predispose to MS. To test the hypothesis of mutually exclusive HLA patterns, patients affected by T1DM plus MS were compared to those of patients affected by either of the diseases alone in a case-control study. METHODS: Blood was sampled for analysis of HLA class I and class II alleles from 66 patients of German ancestry, of whom 33 had T1DM plus MS, and 33 had MS-only. For comparison to patients with T1 DM-only we referred to published data. HLA typing was performed using conventional serology (immuno-magnetic beads) and genotyping (SSP-PCR Dynal(R) SSP low/high resolution kits). RESULTS: Individuals with co-existing MS plus T1DM displayed the expected T1DM associated HLA-pattern (75.8% carried DRB1*04, 69.7% carried DQB1*0302, 42% were DR4, DR3 heterozygous), but failed to display the expected MS associated HLA-pattern (0% carried DQB1*0602, 3.1% carried DQA1*0102). The expected MS associated HLA-pattern of Caucasoid patients, however, was found in the MS-only patients (42% carried DRB1*1501-DQB1*0602, 58% carried DQA1*0102), while the prevalence of T1DM susceptibility and 'resistance' alleles was not different from the general population. The allele frequency of DRB1*1501 was 16/66, 24.2% in the 33 MS-only patients, and 0% in the 33 MS plus T1DM patients. The allele frequency of DQB1*0602 was 16/66, 24.2% in the 33 MS-only patients, and 0% in the 33 MS plus T1DM patients. The allele frequency of DQA1*0102 was 18/66, 27.3%, in the 33 MS-only patients, and 1/66 1.5% in the 33 MS plus T1DM patients. CONCLUSION: These data confirm the hypothesis of mutually exclusive HLA-patterns of T1DM and MS, and are consistent with a low rate of co-morbidity of both diseases.

Phylogenetic analysis of avian paramyxovirus 1 strains isolated in Finland.

Eight strains of avian paramyxovirus type 1 (PMV-1) isolated in Finland during the last 3 decades were studied with reverse transcriptase polymerase chain reaction (RT-PCR) and subsequent sequence analysis of the region of 208 nucleotides covering the fusion (F) protein cleavage site. Both genetic and antigenic heterogeneity of the strains was significant. Direct epidemiological links between strains isolated during successive outbreaks, and also between strains isolated from wild fauna and from poultry or captive birds, were seen in this study. These results also support the previously published view that wild waterfowl serve as a reservoir for the apathogenic or low-pathogenic strains, allowing them to evolve further into pathogenic strains.

Sustained human chemosignal unconsciously alters brain function.

The human chemosignal, Delta 4,16-androstadien-3-one modulates psychological state without being consciously discernible as an odor. This study demonstrates that Delta 4,16-androstadien-3-one (androstadienone) alters cerebral glucose utilization both in subcortical regions and in areas of the neocortex not exclusively associated with olfaction. These widely distributed changes are consistent with modulation of an integrated neural network for regulation of emotional and attentional states. This is the first study to demonstrate the effects of a sustained chemosignal on brain metabolism and to show that they are similar to those of long acting chemical substances that affect psychological states. Moreover, this provides the first evidence that a human chemosignal has distributed effects on cortical processes and brain metabolism even when it is not detected consciously.

Acceptable protective efficacy of influenza vaccination in young military conscripts under circumstances of incomplete antigenic and genetic match.

Commercial inactivated parenteral influenza vaccines reduced febrile (> or = 38 degrees C) respiratory illness by 53% (95% CL: 41-63%) during a 3 week outbreak in 1998 when A/Sydney/5/97(H3N2)-like influenza viruses were shown to be the predominant etiological agents and an older antigenic variant, A/Nanchang/933/95, served as the vaccine virus. The calculatory efficacy for preventing virologically diagnosed influenza infections was 57% (95% CL: 40-68%). The study population consisted of 1374 young male military conscripts. Vaccination coverage on a voluntary basis was 67%. Vaccination was ineffective in preventing febrile illness during a second epidemic wave lasting 2 weeks when mainly adenoviruses were shown to have been circulating in the garrison. Out of the 36 nasopharyngeal aspirates positive for influenza A by antigen detection, 18 A/Sydney/5/97-like strains (10 from non-vaccinated and eight from vaccinated subjects) and two A/Nanchang/933/95-like strains (both from non-vaccinated subjects) were isolated in MDCK cell cultures. Intraepidemic variation was detected among the A/Sydney/5/97-like field strains in their HA1 sequences and reactivity in HI tests, but no evidence was obtained that this variation would have been of significance to the virus in breaking through the vaccination-induced immunity.

Arginine/lysine-rich structural element is involved in interferon-induced nuclear import of STATs.

Signal transducers and activators of transcription (STATs) are latent cytoplasmic transcription factors, which mediate interferon (IFN), interleukin, and some growth factor and peptide hormone signaling in cells. IFN stimulation results in tyrosine phosphorylation, dimerization, and nuclear import of STATs. In response to IFN-gamma stimulation, STAT1 forms homodimers, whereas IFN-alpha induction results in the formation of STAT1.STAT2 heterodimers, which assemble with p48 protein in the nucleus. Phosphorylation as such is not sufficient to target STATs into the nucleus; rather, the dimerization triggered by phosphorylation is essential. Although IFN-induced nuclear import of STATs is mediated by the importin/Ran transport system, no classic nuclear localization signal (NLS) has been found in STATs. In the three-dimensional structure of STAT1, we observed a structural arginine/lysine-rich element within the DNA-binding domain of the molecule. We created a series of point mutations in these elements of STAT1 and STAT2 and showed by transient transfection/IFN stimulation assay that this site is essential for the nuclear import of both STAT1 and STAT2. The results suggest that two arginine/lysine-rich elements, one in each STAT monomer, are required for IFN-induced nuclear import of STAT dimers. Import-defective STAT1 and STAT2 proteins were readily phosphorylated and dimerized, but they functioned as dominant negative molecules inhibiting the nuclear import of heterologous STAT protein.

Molecular epidemiology and evolution of coxsackievirus A9.

Genetic relationships between 35 clinical isolates of coxsackievirus A9 (CAV9), collected during the last five decades from different geographical regions, were investigated by partial sequencing. Analysis of a 150 nucleotide sequence at the VP1/2A junction region identified 12 CAV9 genotypes. While most of the strains within each genotype showed geographical clustering, the analysis also provided evidence for long-range importation of virus strains. Phylogenetic analysis of a longer region around the VP1/2A junction (approximately 390 nucleotides) revealed that the designated genotypes actually represented phylogenetic lineages. The phylogenetic grouping pattern of the isolates in the analysis of the VP4/VP2 region was similar to that obtained in the VP1/2A region whereas analysis of the 3D region indicated a strikingly different grouping, which suggests that recombination events may occur in the region encoding the nonstructural proteins. Analysis of the deduced amino acid sequences of the VP1 polypeptide demonstrated that the RGD (arginine-glycine-aspartic acid) motif, implicated in the interaction of the virus with integrin, was fully conserved among the isolates.

Evidence of recombination among enteroviruses.

Human enteroviruses consist of more than 60 serotypes, reflecting a wide range of evolutionary divergence. They have been genetically classified into four clusters on the basis of sequence homology in the coding region of the single-stranded RNA genome. To explore further the genetic relationships between human enteroviruses and to characterize the evolutionary mechanisms responsible for variation, previously sequenced genomes were subjected to detailed comparison. Bootstrap and genetic similarity analyses were used to systematically scan the alignments of complete genomic sequences. Bootstrap analysis provided evidence from an early recombination event at the junction of the 5' noncoding and coding regions of the progenitors of the current clusters. Analysis within the genetic clusters indicated that enterovirus prototype strains include intraspecies recombinants. Recombination breakpoints were detected in all genomic regions except the capsid protein coding region. Our results suggest that recombination is a significant and relatively frequent mechanism in the evolution of enterovirus genomes.

Relationships between simian and human enteroviruses.

Partial sequences from two genomic regions of simian enteroviruses were analysed and their relatedness to other picornaviruses was compared. Of the 18 simian viruses included in the analysis, sequences were obtained from eleven strains for at least one genomic region. In the 5' non-coding region, SV6, SV19, SV26, SV35, SV43 and SV46 (simian viruses) and BA13 (baboon virus) clearly grouped together with human enteroviruses, whereas SV4, SV28 and SA4 (South African isolate) were more distantly related. In the 3D RNA polymerase-coding region, SV26, SV35, SV43 and SV46 could be clearly identified as enteroviruses and fell into the previously defined cluster A, which contains such human viruses as coxsackievirus A16 and enterovirus 71. However, although SV6 and BA13 were also enterovirus-like, they did not belong to any known genetic cluster of human enteroviruses. Moreover, while SV18 could be recognized as a picornavirus, it did not directly group with members of the genus Enterovirus.

A new picornavirus isolated from bank voles (Clethrionomys glareolus).

A previously unknown picornavirus was isolated from bank voles (Clethrionomys glareolus). Electron microscopy images and sequence data of the prototype isolate, named Ljungan virus, showed that it is a picornavirus. The amino acid sequences of predicted Ljungan virus capsid proteins VP2 and VP3 were closely related to the human pathogen echovirus 22 (approximately 70% similarity). A partial 5' noncoding region sequence of Ljungan virus showed the highest degree of relatedness to cardioviruses. Two additional isolates were serologically and molecularly related to the prototype.

Foodborne outbreaks of hepatitis A in a low endemic country: an emerging problem?

This paper describes 2 outbreaks of hepatitis A infection in Finland, a very low endemic area of hepatitis A infection, where a large proportion of the population is now susceptible to infection by hepatitis A virus (HAV). The first outbreak involved people attending several schools and day-care centres; the second employees of several bank branches in a different city. The initial investigation revealed that both were related to food distributed widely from separate central kitchens. Two separate case-control studies implicated imported salad food items as the most likely vehicle of infection. HAV was detected in the stool of cases from both outbreaks using reverse-transcriptase polymerase chain reaction; however, comparison of viral genome sequences proved that the viruses were of different origin and hence the outbreaks, although occurring simultaneously, were not linked. Foodborne outbreaks of HAV may represent an increasing problem in populations not immune to HAV.

Genetic and phylogenetic clustering of enteroviruses.

Genetic and phylogenetic analysis of enteroviruses showed that in the 5'NCR enteroviruses formed three clusters: polioviruses (PVs), coxsackievirus A type 21 (CAV21), CAV24 and enterovirus type 70 (ENV70) formed one cluster; coxsackievirus B isolates (CBVs), CAV9, CAV16, ENV71, echovirus type 11 (EV11), EV12 and all partially sequenced echoviruses and swine vesicular disease virus (SVDV) belonged to another cluster and bovine enteroviruses (BEVs) formed the third cluster. In the capsid coding region five clusters were seen: PVs, CAV21 and CAV24 formed one cluster (PV-like); ENV70 formed a cluster of its own; all CBVs, CAV9, EV11, EV12 and SVDV formed the third cluster (CBV-like); CAV16, CAV2 and ENV71 belonged to the fourth cluster (CAV16-like) and BEVs formed their own cluster (BEV-like). In the 3'NCR the same clusters were seen as in the coding region suggesting a close association of the 3'NCR with viral proteins while the cellular environment may be more important in the evolution of the 5'NCR. Secondary structures were predicted in the 3'NCR, which showed two different patterns among the five clusters. A potential pseudoknot region common in all five clusters was identified. Although the BEV-like viruses formed a separate cluster in all genomic regions, in the coding region they seem to be phylogenetically related to the CAV16-like viruses.

Recent influenza B viruses in Europe: a phylogenetic analysis.

BACKGROUND: Influenza B virus evolution is currently in a unique situation having two cocirculating main lineages B/Yamagata/16/88 (YM/88)-like and B/Victoria/2/87 (VI/87)-like viruses. Continuation of this bifurcation would mean development towards distinct forms resembling the HA subtypes of influenza A viruses. OBJECTIVE: We wanted to examine both intraepidemic heterogeneity and recent evolution in these two lineages. The initial purpose was to determine the geographic distribution of the two sublineages of the VI/87-like viruses in Europe in 1989-1990 under circumstances of low epidemic activity. Due to the outbreaks of YM/88-like viruses since 1991, the study was extended to contain the evolution of these viruses and their genetic relationship with the vaccine strains of that time. STUDY DESIGN: The HA1 gene sequences of 33 influenza B strains isolated in ten European countries since 1989 were determined and compared with those available through databases or personal contacts. RESULTS: The two main lineages, YM/88-like and VI/87-like viruses, both continued to circulate. In both lineages, changes in the potential glycosylation sites were observed. Two sublineages of the VI/87 lineage cocirculated during the 1989-1990 season with somewhat different geographic distributions. A high degree of intraepidemic heterogeneity was observed, as well as examples of conserved nucleotide sequences. CONCLUSIONS: It is important to follow the evolution and circulation of VI/87-like viruses. Current vaccines give poor or no protection against VI/87-like viruses in immunologically unprimed children or even in primed adults (Levandowski et al., 1991, Pyhala et al., 1994). Changes in the potential glycosylation pattern in the latest virus isolates of both main lineages have occurred and it is interesting to see the significance of these changes to viral evolution.

HA1 domain of influenza A (H3N2) viruses in Finland in 1989-1995: evolution, egg-adaptation and relationship to vaccine strains.

The HA1 gene sequences of 22 MDCK cell-derived influenza A (H3N2) strains, ten of their egg-derived counterparts and three vaccine strains were determined. Antigenic and sequence differences between the epidemic and vaccine strains were recorded, most striking in 1992/93; a minority of the amino acid differences in 1989-95 was involved in egg-adaptation. Changes in the assortment of amino acid substitutions produced during egg-adaptation of field strains may account for the difficulty encountered in isolating these viruses in embryonated eggs. Six revertant amino acids, characteristic of field strains prevalent in 1969-71 were recorded in 1994/95. Their genome sequence was interpreted to have been maintained over the interval years among low abundant sequences of the viral quasispecies. Potential changes of carbohydrate moieties were recorded in two glycosylation sites, suggesting that oligosaccharides at these sites are not necessarily advantageous for the H3N2 subtype virus currently.

Stage-specific regulation of murine Hsp68 gene promoter in preimplantation mouse embryos.

In early mouse embryos, the major inducible heat shock gene, hsp68, is spontaneously and transiently activated at the two-cell stage and becomes heat-inducible around blastocyst stage. We have probed mouse embryo's ability to activate the promoter of this gene during preimplantation development by expression analysis of DNA constructs containing a reporter lacZ gene driven by hsp68 (hsp70A1) 5'-regulatory sequences of various length: (i) a full-length promoter (construct phsplacZ); (ii) a heat shock element (HSE)-deleted promoter (p delta 1hsplacZ); and (iii) a minimal, proximal promoter (p delta 2hsplac Z). When analyzed in transfected L-cells, phsplacZ was heat-inducible, while neither p delta 1hsplacZ nor p delta 2hsplacZ was. Developmental activity of the full-length construct was first analyzed after genome integration in transgenic embryos and found to follow endogenous hsp68 expression in terms of spontaneous activation at the 2-cell stage, down-regulation at the 4-cell stage, and acquisition of heat inducibility at the 16/32-cell stage. In transient expression experiments, injected phsplacZ, p delta 1hsplacZ, and p delta 2hsplacZ were expressed at similar levels by 2-cell embryos, independently of construct topology and injection stage. At the 4-cell stage, however, phsplacZ and p delta 1hsplacZ were expressed at similar levels, while p delta 2hsplacZ was inactive. Only phsplacZ became heat-inducible in late morulas. We conclude that in early mouse embryos, developmental activity of episomic hsp68 promoter depends on proximal sequences at the 2-cell stage and on putative enhancer sequences at the 4-cell stage, while HSEs appear dispensable during early cleavage.

Evolution of the HA1 domain of human influenza A (H1N1) virus: loss of glycosylation sites and occurrence of herald and conserved strains.

Thirty-one strains of human influenza A (H1N1) viruses isolated in Europe, mostly in Finland, from 1978-1992 were compared with respect to their nucleotide sequences coding for the HA1 portion of haemagglutinin. In 1984, at least two sublineages of H1N1 subtype viruses co-circulated in Finland. The viruses isolated after 1986 formed three sequential phylogenetic clusters. Loss of glycosylation sites, on the globular head of the HA1 portion suggests that oligosaccharides at these sites are not necessarily advantageous for the human virus. Isolation of a herald strain in Finland in June 1988 raised the question as to whether the virus was able to survive in Europe throughout the non-epidemic summer period. Demonstration of highly conserved strains, found over two continents in 1988, is further evidence of the existence of infection chains whose viruses have not been subjected to random sampling or selection events.