Microcephaly, intractable seizures and developmental delay caused by biallelic variants in TBCD: further delineation of a new chaperone-mediated tubulinopathy.

Microtubule dynamics play a crucial role in neuronal development and function, and several neurodevelopmental disorders have been linked to mutations in genes encoding tubulins and functionally related proteins. Most recently, variants in the tubulin cofactor D (TBCD) gene, which encodes one of the five co-chaperones required for assembly and disassembly of α/ß-tubulin heterodimer, were reported to underlie a recessive neurodevelopmental/neurodegenerative disorder. We report on five patients from three unrelated families, who presented with microcephaly, intellectual disability, intractable seizures, optic nerve pallor/atrophy, and cortical atrophy with delayed myelination and thinned corpus callosum on brain imaging. Exome sequencing allowed the identification of biallelic variants in TBCD segregating with the disease in the three families. TBCD protein level was significantly reduced in cultured fibroblasts from one patient, supporting defective TBCD function as the event underlying the disorder. Such reduced expression was associated with accelerated microtubule re-polymerization. Morpholino-mediated TBCD knockdown in zebrafish recapitulated several key pathological features of the human disease, and TBCD overexpression in the same model confirmed previous studies documenting an obligate dependency on proper TBCD levels during development. Our findings confirm the link between inactivating TBCD variants and this newly described chaperone-associated tubulinopathy, and provide insights into the phenotype of this disorder.

[Comparative analysis of ISSR markers polymorphism in populations of yak (Bos mutus) and in F1 hybrids between yak and cattle in the Sayan-Altai region].

The genetic variability in seven yak populations from the Sayan-Altai region and in F1 hybrids between yak and cattle (khainags) was investigated with the help of a technique that involves the use of inter simple sequence repeat (ISSR) markers generated with PCR primers (AG)9C and (GA)9C. Samples for the analysis were collected in Mongolia, Tuva, and Altai from 2008 through 2012. The examined yak populations differed in in the presence/absence of ISSR fragments, as well as in their frequency. In total, 46 ISSR fragments were identified using two marker systems; the proportion of polymorphic loci constituted 76% and 90% for the AG-ISSR and GA-ISSR markers, respectively. For the total sample of yaks, total genetic diversity (Ht), within-population diversity (Hs), and interpopulation diversity (Gst) constituted 0.081, 0.044, and 0.459 for the AG-ISSR and 0.137, 0.057, and 0.582 for the GA-ISSR markers, respectively. Based on ISSR finger printing, species- and breed-specific DNA patterns were described for the three groups of animals (yaks, cattle, khainags). For the domestic yak, the species-specific profile was represented by eight ISSR fragments. Genetic relationships between the yak populations, cattle breeds, and khainags were examined with the help of four different approaches used in the analysis of population structure: estimation of phylogenetic similarity, multidimensional scaling, principal component analysis, and cluster analysis. Clear evidence on the differentiation of the populations examined at the interspecific, as well as at intraspecific, level were obtained. Similar (relative); as well as remote (isolated), yak populations were identified. Khainags occupy an intermediate position between yak and cattle. However, the data on the ISSR-PCR marker polymorphism (genome polymorphism, population structure).indicate that part of the analyzed khainag genome was more similar to the yak genome than to the cattle genome.

[Comparison of ISSR polymorphism among cattle breeds].

Polymorphism analysis of DNA fragments flanked by (TC)9G and (CT)9G inverted dinucleotide microsatellite repeats in 766 animals of 19 cattle breeds and one breeding type revealed 66 fragments, of which 64 were polymorphic. The breeds proved to differ in the frequency and presence or absence of amplified DNA fragments at the genomic level, indicating that ISSR fingerprinting is informative for differentiating the PCR product spectra and cattle breeds. Multilocus ISSR polymorphism analysis identified the group of fragments that can be used as Bos taurus and B. indicus species markers to describe the standards of breeds, their genetic profiles, and breed-specific patterns. Based on ISSR polymorphism, a prototypal gene pool of cattle was constructed and the breeds closest to it were identified. Genetic diversity analysis made it possible to assume that an optimal mean heterozygosity is characteristic of cattle breeds and that deviations from this optimum are indicative of various processes occurring in the population (breed).

[Analysis of the genetic structure of Tuvinian short-fat-tailed sheep populations with the use of the ISSR-PCR method].

The genetic structure of populations of the Tuvinian short-fat-tailed sheep was studied with the use of the ISSR-PCR (Inter Simple Sequence Repeats) method in 18 farms of Tyva. Data on the spectrum of ISSR fragments of DNA were obtained using the (AG)9C primer. Analysis of intermicrosatellite polymorphism permitted us to determine genomic characteristics of the populations, their genealogical relations, and the parameters of genetic diversity within the populations and the breed as a whole. Three genetic notions were considered on the basis of the results of this analysis: gene pool profile, gene pool standard, and breed-specific pattern. The data obtained can be used to carry out population genetic monitoring, to develop a breeding strategy, and to conserve in situ the Tuvinian sheep breed and breeds of other domesticated species.

[Mitochondrial DNA polymorphism in Tuvinian population of reindeer Rangifer tarandus L].

Mitochondrial DNA variation was examined in one of the southern most populations of domestic reindeer, inhabiting Tyva Republic (Tuva). In Tuvinian population sequence polymorphism of the mitochondrial DNA D loop region was demonstrated. In a sample of 29 individuals 7 mitotypes were distinguished, pointing to the preservation of rather high level of genetic diversity in this population.

[Polymorphism of ISSR-PCR markers in Tuvinian population of reindeer Rangifer tarandus L].

The results of the first analysis of the reindeer population inhabiting the south of Siberia, the Republic of Tyva (Tuva), performed by use of inter-simple sequence repeat (ISSR) technique, are reported. Using ISSR-PCR with two primers, a total of 71 fragments of different size were detected. Furthermore, for the population examined the indices of the mean pairwise similarity and the mean heterozygosity were calculated. The approach used can be recommended as a useful tool for the genetic diversity monitoring in Tuvinian population of reindeer.

Infectious stunting and leg weakness in broilers: I. Pathology and biochemical changes in blood plasma.

A syndrome of stunting and leg weakness could be reproduced experimentally by inoculation of 1-day-old broilers with homogenised intestines from affected birds. Inoculated birds kept in isolators showed highly impaired growth until 3 weeks p.i. Birds produced mucoid yellowish coloured droppings and at post mortem thin liquid intestinal contents were found. Biochemical examination of blood plasma showed low plasma carotenoid concentrations and an increased alkaline phosphatase (ALP) activity, mainly caused by one isoenzyme, which was most likely of intestinal origin. These findings implicate infectious causal agents with the intestines as the site of primary involvement. Bone abnormalities consisted of rickets-like changes at the age of 3 weeks, whereas a distinct dyschondroplasia was seen at 4 weeks. The syndrome could also be transmitted to uninoculated birds kept in contact with birds inoculated at 1 day of age. Birds inoculated at 7 days of age also showed greatly impaired growth but developed no macroscopical bone disorders. Inoculation at 14 days of age did not result in impaired growth or bone abnormalities. Following inoculation with REO virus, isolated from a field case, no bone abnormalities occurred. However, a shortlived impaired growth, diarrhoea, increased plasma ALP activity and decreased carotenoid concentration were observed. The rapid spread of the disease and the role of REO virus are discussed.

Egg production in relation to the results of a long term serological survey of 73 flocks of fowl.

Seventy-three flocks of fowl were tested at regular intervals for the presence of precipitins to fowl adenovirus (AV) and infectious bronchitis virus (IBV), haemaggluinating inhibiting antibodies to BC in 14 virus, and of agglutinins to Mycoplasma gallisepticum (M.g.) and Mycoplasma synoviae (M.s.). In all the eight flocks affected with Egg Drop Syndrome (EDS '76), egg production problems were associated with increasing numbers of BC14 virus reactors and AV reactors. In flocks showing production problems other than EDS'73 without any apparent cause, the average percentage of AV reactors increased significantly after the rearing period; this was not true of IBV reactors. BC14 reactors were either absent or present only once, in small numbers and with low titres, during the test period. The average percentage of AV reactors did not increase after the rearing period either in normally producing flocks or in flocks with production problems for which other diseases or dietary errors plausibly accounted for these problems. All this suggests a pathogenic role of AV in production problems. One can conclude from the high percentage of reactors in all groups of flocks that subclinical IBV infections are common. The percentage of IBV reactors during the laying period of flocks with EDS'76 was significantly higher than that of normally producing flocks. It is therefore suggested that subclinical IBV infection could be among the factors causing stress, acting as a trigger for EDS'76. All M.g.-infected flocks showed production problems; M.s. infections could not be related to egg production disturbances.

Egg production in relation to the results of a long term serological survey of 73 flocks of fowl.

Summary Seventy-three flocks of fowl were tested at regular intervals for the presence of precipitins to fowl adenovirus (AV) and infectious bronchitis virus (IBV), haem-agglutinating inhibiting antibodies to BC14 virus, and of agglutinins to Mycoplasma gallisepticum (M.g.) and Mycoplasma synoviae (M.s.). In all the eight flocks affected with Egg Drop Syndrome (EDS '76), egg production problems were associated with increasing numbers of BCI4 virus reactors and AV reactors. In flocks showing production problems other than EDS '76 without any apparent cause, the average percentage of AV reactors increased significantly after the rearing period; this was not true of IBV reactors. BC14 reactors were either absent or present only once, in small numbers and with low titres, during the test period. The average percentage of AV reactors did not increase after the rearing period either in normally producing flocks or in flocks with production problems for which other diseases or dietary errors plausibly accounted for these problems. All this suggests a pathogenic role of AV in production problems. One can conclude from the high percentage of reactors in all groups of flocks that sub-clinical IBV infections are common. The percentage of IBV reactors during the laying period of flocks with EDS '76 was significantly higher than that of normally producing flocks. It is therefore suggested that subclinical IBV infection could be among the factors causing stress, acting as a trigger for EDS '76. All M.g.-infected flocks showed production problems; M.s. infections could not be related to egg production disturbances.

Dropped egg production, soft shelled and shell-less eggs associated with appearance of precipitins to adenovirus in flocks of laying fowls.

Two outbreaks of dropped egg production and production of soft shelled and shell-less eggs are described. The outbreaks were selected from a larger number of flocks with similar problems in the field. The drop in egg production was closely correlated with the appearance of precipitins to adenovirus in the laying birds. Birds were also infected with Mycoplasma synoviae. A number of other infectious causes of production problems could be excluded in these outbreaks. In particular infectious bronchitis was not a factor and, in contrast to experiences following infections with this virus, production recovered nearly completely within 6 to 10 weeks. The observations suggest that in the field infections with adenovirus may cause production problems as serious as those caused by infectious bronchitis virus.