The equine graying with age mutation of the STX17 gene: A copy number study using droplet digital PCR reveals a new pattern.

The equine graying with age causative mutation in the syntaxin-17 gene (STX17) has been known for over a decade, but proper genotyping of this variant remains challenging due to its molecular character (4.6-kb tandem duplication). Precise information on gray mutation status is important for horse breeders and veterinarians, since gray homozygous horses are more prone to developing aggressive melanoma tumors than heterozygotes. Since recent studies have confirmed that droplet digital PCR is a valuable technique for copy number analysis, we decided to investigate whether this method can be used for accurate genotyping of the horse graying-related variant and established the copy numbers of the 4.6-kb fragment in the available cohort (n = 75) of gray and nongray horses of various breeds. Surprisingly, we found that our STX17 genotype results varied from what has been previously published, suggesting that gray phenotype is associated with the presence of six (GG) or four (Gg) copies of studied region. All the examined nongray horses (gg) have the two copies of these fragments. This new pattern and its inheritance were also confirmed by an analysis conducted for the Polish Warmblood horse family. We noted no further copy number variation in the entire tested samples set. Our study confirmed the usefulness and accuracy of droplet digital PCR for genotyping STX17 gene variant. Further studies on a broader range of materials are needed to fully understand the origin and molecular structure of the graying causative mutation in the horse STX17.

TBX3 and ASIP genotypes reveal discrepancies in officially recorded coat colors of Hucul horses.

Although only a few specific pigmentation types are allowed within the Hucul horse registry, accurate determination of particular coat colors can be uncertain due to the presence of variation in color shades and segregation of multiple dun dilution variants. Herein, we genotyped the previously identified polymorphisms within two coat color loci TBX3 (T-box 3) and ASIP (Agouti Signaling Protein) in 462 Hucul individuals and compared the genotype predicted phenotypes with observed pigmentation types provided in the Polish Horse Breeders Association database. We identified disagreement between the predicted and recorded coat color in 157 horses (34%). The most common error was misclassification of horses with the nd1/nd1 and nd1/nd2 genotypes, what may be related with the occurrence of some 'intermediate' dilution phenotypes in such individuals. We have also proven that the frequency of the dominant dun dilution allele (D) (0.30) is higher than previously predicted by available studbooks. The D allele(s) is easily 'hidden' in various phenotypic groups including dark bay and black, therefore we hypothesized that the dun dilution effect itself is not as strongly epistatic in the Hucul horse as described in other horse breeds. This may be the result of an additional genetic modifier suppressing D allele phenotypic effect.

Polymorphism of the porcine leptin gene promoter and analysis of its association with gene expression and fatness traits.

We describe for the first time a 245 bp fragment of the porcine leptin gene promoter in the proximity of the transcription start site. Altogether, 720 pigs were screened with the PCR-SSCP technique for polymorphism in this region. Four SNPs, segregating as two haplotypes, have been identified, one of them (C113G) in the putative consensus site for the AP-2 transcription factor. This polymorphism was evenly distributed in the Duroc breed (n=21) and was absent in the Polish Landrace (n=248) and Pietrain breed (n=12). In the Polish Large White (n=191) and synthetic line 990 (n=243), allele G occurred with a very low frequency. The investigation was performed to test if the C113G SNP affects leptin mRNA level in subcutaneous fat and leptin protein concentration in serum. Additionally, the effect of this polymorphism on fatness traits was statistically analyzed. Although there was a trend toward decreased expression in GG animals, the differences were not significant between genotypes. We also found no evidence for an association of the LEP promoter genotype with the analyzed fatness traits.

Missense mutations in exon 4 of the porcine LEPR gene encoding extracellular domain and their association with fatness traits.

Three non-synonymous single nucleotide polymorphisms (T221C, T232A and C233T) were detected in exon 4 of the porcine leptin receptor (LEPR) gene. The T232A substitution could be identified as a (Tsp509I) restriction fragment length polymorphism. The frequency of genotype TT varied in six genetic groups from 0.62 (Duroc) to 0.99 (Polish Large White). Sequencing of exon 4, performed for 30 animals, revealed that only two intragenic haplotypes (TC and AT at nucleotide position 232-233) were present. The phenotypic effect of the Tsp509I polymorphism was tested for the Polish Landrace (n = 241) and a synthetic line 990 (n = 243). There was no statistical evidence for the direct effect of the LEPR polymorphisms on fatness traits. However, in Polish Landrace allele A at position 232 was associated with thicker backfat over shoulder.

Application of the pinless external fixator in severely burned patients.

In the treatment of patients with full thickness burns of the limbs adequate positioning is known as a critical condition for a favourable outcome after debridement and skin grafting. Up to now prevention of wound compression, prevention of shear forces and daily wound inspection were facilitated by using halofixators (HF) for the skull and AO external fixation (AO-EF) systems for the limbs. For the skull the HF is still the method of choice. However, in burned limbs the AO-EF has the considerable disadvantage that the pins penetrate the medullary cavity. Pin track infections, which occur quite frequently in burn patients, may thus lead to osteitis and additional impairment of the patient's compromised immune function. For these reasons, a new pinless fixation system has been introduced and tested in a series of eight patients with burns of the lower limbs. The special pin clamp system affects only the external subperiosteal portion of the bone. Application of the clamps is very fast since no drilling is necessary. The advantages of external fixation are retained without the associated disadvantages. Clinical examples are presented and discussed.

[Early functional treatment of rupture of the fibular ligament with a personally developed ankle splint]. / Frühfunktionelle Therapie der fibularen Bandruptur mit einer selbstentwickelten Knöchelschiene.

The inversion trauma of the ankle joint is one of the most important and frequent injuries. The treatment of recent rupture of the fibular ligament is now a point for discussion. Operative or conservative treatment is possible. The clinical evaluation of instability requires radiological control and stress test. With a new ankle splint we reduce pathological inversion of the ankle joint, so that we introduce a functional treatment. Our experience with this method will be demonstrated. 45 patients more than a half year after trauma reported good-to-excellent results. The objective evaluation shows in 95% good-to-excellent results. A secondary ligament reconstruction was not necessary.