In the present study, enterovirus RNA was detected in serum long before the diagnosis of clinical type 1 diabetes. The frequency of virus peaked during the 6-month window that preceded the first appearance of diabetes-associated autoantibodies. This temporal relationship has also been observed in our previous studies, suggesting that enterovirus infections may play a role in the initiation of the β-cell–damaging process (8
). Enterovirus RNA was also more common in the case than in the control children after the initial autoantibody seroconversion but was not detected in samples taken close to the presentation of diabetes. The absence of enterovirus RNA at the diagnosis of diabetes is in contrast to the majority of the retrospective case-control studies where altogether an average of 31% of the patients and 6% of the control subjects have been positive for enterovirus RNA (3
). The controversy might be due to methodological differences, e.g., the sensitivity of the PCR applied or the type of samples collected (16
). On the other hand, it may also reflect a true difference in enterovirus epidemiology. In fact, we have previously observed that enterovirus infections are less frequent in Finland compared with many other countries (17
). In any case, the present study emphasizes the importance of those infections that occur during the early stages of the diabetogenic pathway, possibly playing a role in the initiation of the β-cell–damaging process rather than the later stages of the process.
As enterovirus viremia usually lasts for a maximum of 2 weeks, the number of enterovirus episodes is largely underestimated if samples are collected at longer intervals. Accordingly, we can estimate the true number of enterovirus RNA–positive episodes (N of positive samples/detection period covered by the collected samples × total follow-up time), which would be 154 episodes in case and 254 in control children (mean 3.7 vs. 1.6 episodes per child). However, it is also possible that the difference between the case and control children reflects prolonged enterovirus episodes in the case group.
Our previous studies among DIPP children suggest that the detection of the first diabetes-associated autoantibody and enterovirus RNA in stools shows similar seasonal variation (19
). In the present study, the same seasonal pattern was seen in the detection of enterovirus RNA in serum (frequent in the autumn and winter). In addition, viral RNA was most common in serum at the age when autoantibodies were most frequently induced. Autoantibodies became detectable soon after detection of enterovirus RNA in the serum. In mouse models, this time interval has also been short (20
It has previously been shown that children are protected against enterovirus infections by maternal antibodies and that this protection is at least partly mediated by antibodies in breast milk. In Finland, children are breastfed for an average of 8 months (range 0.15–23months) (21
). This may explain partly the low frequency of viral RNA observed in infants aged <6 months. Viral RNA was most frequent in 12- to 18-month-old children, suggesting that there is a susceptibility period at that age.
Enterovirus persistence has been shown to play a role in chronic cardiomyopathies, and it may also be involved in type 1 diabetes (5
). In the present study, no signs of persistent systemic infection were seen; enterovirus genomes that were detected from repeatedly positive children represented different viral genotypes. However, in persisting infections, the virus replication can occur at a very low level and the virus may not be detectable in peripheral blood even though it may be present in the pancreas or other organs (4
The risk effect of enterovirus RNA on autoantibody development was stronger among boys than among girls. Boys are also known to be more susceptible to general complications of enterovirus infections. The higher number of enterovirus RNA–positive samples in children with the DR3-DQ2/DR4-DQ8 genotype merits further investigation to find out whether this genotype is particularly susceptible to diabetogenic effect enteroviruses.
In conclusion, the present study supports the hypothesis that enteroviruses play a role in the pathogenesis of type 1 diabetes. The presence of virus in serum was shown to be a risk factor for the development of β-cell–specific autoimmunity, which progress to clinical diabetes, especially among boys.