Summary.  Background: Type 3 von Willebrand disease (VWD) is an autosomal recessive bleeding disorder, characterized by virtually undetectable plasma von Willebrand factor (VWF) and consequently reduced plasma factor VIII levels. Genetic mutations responsible for type 3 VWD are very heterogeneous, scattered throughout the VWF gene and show high variability among different populations. Methods: Twenty‐five severe VWD patients were studied by direct sequencing of the 51 coding exons of the VWF gene. The total number of VWD type 3 families in Hungary is 24, of which 23 were investigated. Results: Fifteen novel mutations were identified in 31 alleles, five being nonsense mutations (p.Q1238X, p.Q1898X, p.Q1931X, p.S2505X and p.S2568X), four small deletions and insertions resulting in frame shifts (c.1992insC, c.3622delT, c.5315insGA and c.7333delG), one a large partial deletion (delExon1‐3) of the 5′‐region, four candidate missense mutations (p.C35R, p.R81G, p.C295S, p.C623T) and one a candidate splice site mutation (c.1730–10C>A). Six previously described mutations were detected in 17 alleles, including the repeatedly found c.2435delC, p.R1659X and p.R1853X. Only one patient developed alloantibodies to VWF, carrying a homozygous c.3622delT. Conclusion: We report the genetic background of the entire Hungarian type 3 VWD population. A large novel deletion, most probably due to a founder effect, seems to be unique to Hungarian type 3 VWD patients with high allele frequency. In contrast to previous reports, none of the five patients homozygous for the large partial deletion developed inhibitors to VWF. This discrepancy raises the possibility of selection bias in some of the reports.

The primary immune response to a viral antigen (tick-borne encephalitis, TBE) has been determined in haemophiliacs. Twelve HIV-negative and four clinically asymptomatic, HIV-positive haemophiliacs as well as 16 age-matched healthy controls were included in the study. Antibody responses after TBE vaccination were comparable in HIV-negative haemophiliacs and controls; however, antibody titres in HIV-infected haemophiliacs were significantly lower after completion of the three-dose vaccination schedule (geometric mean reciprocal antibody titres (SEM): controls, 193 (1.37), HIV-positive haemophiliacs, 13 (2.18), P < 0.005). TBE vaccination failed to induce a T cell proliferative response in the HIV-positive haemophiliacs. While in HIV-negative patients the antigen-specific lymphoproliferative responses after primary and one booster vaccination were comparable to those of the controls, cellular responses were decreased in HIV-negative haemophiliacs following a second booster immunization 19 months after primary immunization (3H-thymidine incorporation, delta dpm, mean +/- SEM: controls, 34662 +/- 7129, HIV-negative haemophiliacs, 14339 +/- 7420, P < 0.005). As the protective mechanisms for TBE infection are not yet completely understood, further work will be necessary to determine whether the decreased capacity to mount a sufficient long-term cellular memory response in HIV-negative haemophiliacs might be important for the protective effect of TBE vaccination in this population.

Deficiencies of blood coagulation factors VIII and IX (haemophilia A and haemophilia B) represent the most common inherited bleeding disorders with a wide range of causative mutations. Carrier and prenatal diagnostics are preferably performed by direct mutation detection; however, in certain situations, indirect family studies may also be useful. We aimed to utilize a combination of direct and indirect techniques for carrier and prenatal diagnostics in both haemophilias in a single national centre. Two hundred and eleven haemophilia A families were investigated by screening for inversions of introns 1 and 22, and by family studies using polymorphic markers. Twenty-eight haemophilia A and 39 haemophilia B families were investigated by Sanger-sequencing of the coding regions. Among severe haemophilia A families, frequencies of intron 22 and 1 inversions were 82 out of 145 (57%) and two out of 145 (1.4%). Sequencing of the entire coding region of the respective factor gene was performed and 12 (haemophilia A) and 5 (haemophilia B) previously unpublished disease-causing mutations were identified. For genetic markers used for haemophilia A indirect family testing, heterozygosity rates varied between 137 out of 327 [42% intragenic BclI restriction fragment length polymorphism (RFLP], 168 out of 254 (66% intragenic F8Civs13CA) and 202 out of 261 (77% extragenic DXS15CA) with a combined rate of 92% (intragenic markers) and 97% (all three markers). For male fetuses, prenatal diagnostics was provided to 43 haemophilia A families (n = 22 with direct mutation detection and n = 21 by indirect family testing) and to three haemophilia B families. The combination of direct and indirect molecular genetics approaches is a successful and cost-effective approach to provide carrier and prenatal diagnostics and risk assessment for inhibitor formation.

The occurrence of suboptimal vitamin D supply is significant, although less frequent than that in literature reports. In women taking oral contraceptives, serum 25-hydroxyvitamin D levels were higher, but serum intact parathyroid hormone concentrations were not decreased suggesting that the increased 25-hydroxyvitamin D levels may be the consequence of oestrogen-induced alterations of serum protein fractions.

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