Correction: Molecular basis for a new bovine model of Niemann-Pick type C disease.

[This corrects the article DOI: 10.1371/journal.pone.0238697.].

Molecular basis for a new bovine model of Niemann-Pick type C disease.

Niemann-Pick type C disease is a lysosomal storage disease affecting primarily the nervous system that results in premature death. Here we present the first report and investigation of Niemann-Pick type C disease in Australian Angus/Angus-cross calves. After a preliminary diagnosis of Niemann-Pick type C, samples from two affected calves and two obligate carriers were analysed using single nucleotide polymorphism genotyping and homozygosity mapping, and NPC1 was considered as a positional candidate gene. A likely causal missense variant on chromosome 24 in the NPC1 gene (NM_174758.2:c.2969C>G) was identified by Sanger sequencing of cDNA. SIFT analysis, protein alignment and protein modelling predicted the variant to be deleterious to protein function. Segregation of the variant with disease was confirmed in two additional affected calves and two obligate carrier dams. Genotyping of 403 animals from the original herd identified an estimated allele frequency of 3.5%. The Niemann-Pick type C phenotype was additionally confirmed via biochemical analysis of Lysotracker Green, cholesterol, sphingosine and glycosphingolipids in fibroblast cell cultures originating from two affected calves. The identification of a novel missense variant for Niemann-Pick type C disease in Angus/Angus-cross cattle will enable improved breeding and management of this disease in at-risk populations. The results from this study offer a unique opportunity to further the knowledge of human Niemann-Pick type C disease through the potential availability of a bovine model of disease.

Osteoarticular Infection in Three Young Thoroughbred Horses Caused by a Novel Gram Negative Cocco-Bacillus.

We describe three cases of osteoarticular infection (OAI) in young thoroughbred horses in which the causative organism was identified by MALDI-TOF as Kingella species. The pattern of OAI resembled that reported with Kingella infection in humans. Analysis by 16S rRNA PCR enabled construction of a phylogenetic tree that placed the isolates closer to Simonsiella and Alysiella species, rather than Kingella species. Average nucleotide identity (ANI) comparison between the new isolate and Kingella kingae and Alysiella crassa however revealed low probability that the new isolate belonged to either of these species. This preliminary analysis suggests the organism isolated is a previously unrecognised species.

Cellular development in muscle differs between Angus steers from low and high muscle score selection lines1.

This study assessed cellular characteristics of longissimus lumborum (LL) and semitendinosus (ST) muscles in steers genetically selected for low (Low) or high (High) muscling using live muscle scoring, and High steers with 1 copy of the loss-of-function 821 del11 MSTN allele (HighHet). We hypothesized High and HighHet have altered muscle cellular characteristics and mechanisms influencing muscling compared with Low steers. Angus steers 25 mo old comprising 14 High, 19 Low, and 11 HighHet were backgrounded to 20 mo of age, grain finished for 150 d, and then slaughtered. Body and carcass weights did not differ due to muscling line (P = 0.46). Weight of LL was 16% greater (P = 0.004) and total protein in LL was 18% greater (P = 0.012) in HighHet than Low steers. ST weight in HighHet was 10% and 13% greater than in High and Low steers (P = 0.007), respectively, and of total ST protein 12% and 17% greater in HighHet than High or Low (P = 0.002). Cross-sectional area (CSA) of LL was greater in HighHet than in High and greater in High than in Low (85.0 vs. 77.0 vs. 70.4 cm2, P < 0.001). Apparent number of myofibers and myofibers per unit CSA did not differ between the muscling lines in LL (P = 0.14) or ST (P = 0.47). Myofiber CSA was greater in the ST of Low than of High and HighHet for type 1 (36% and 31% respectively, P = 0.005) and 2A (22% and 25%, P < 0.001). HighHet steers had greater area of glycolytic (type 2X) relative to more oxidative myofiber types within LL (P = 0.02; 11% and 43% more than High and Low, respectively) and ST (P < 0.001; 27% and 75%). Concentration of RNA in LL was 13% and 10% greater (P = 0.005) in High than in Low and HighHet, respectively, and total amount of RNA in LL was 22% greater in High and 20% greater in HighHet than in Low (P < 0.001). The LL of High steers had less protein to RNA (P = 0.03; 57.4 vs. 65.6) and more RNA to DNA (P = 0.007; 9.03 vs. 7.83) than Low. HighHet steers had 11% more DNA in ST than High (P = 0.04) and 19% more RNA in ST than Low (P = 0.012). The shift towards glycolytic myofibers was consistent with loadings in a principal component that explained 39% of the variation in LL and 38% in ST. Overall, these findings show that selection for increased muscling using live cattle muscle scoring, and 1 copy of the 821 del11 MSTN allele, results in more glycolytic muscle. They also suggest that increased muscling of the High compared with Low steers may be associated with increased translational capacity in the LL.

Ichthyosis fetalis in Polled Hereford and Shorthorn calves.

Inherited forms of ichthyosis, or generalized scaling of the skin, have been reported in many animal species, including cattle, and are characterized by an autosomal recessive mode of inheritance. We investigated 2 calves affected with ichthyosis fetalis, a Polled Hereford and a Shorthorn. Both cases had hard white plaques on the skin consistent with excessive keratinization. This was confirmed by histopathology, which showed severe diffuse epidermal and follicular orthokeratotic hyperkeratosis. The known mutation (H1935R) in gene ABCA12, responsible for ichthyosis fetalis in Chianina cattle, was shown to be absent in both affected calves and their obligate heterozygous parents. These molecular findings indicate that allelic heterogeneity exists for this condition in cattle.

Internal restriction sites: quality assurance aids in genotyping.

Improvements to restriction fragment length polymorphism (RFLP)-based genotyping assays currently used for detection of mutations responsible for bovine ferrochelatase and myophosphorylase deficiencies, and equine hyperkalemic periodic paralysis (HYPP) are described. Reports of sporadic inhibition of restriction enzyme activity suggest a critical factor in RFLP-based genotyping assays should be assurance that restriction enzymes perform to specification with every sample. The RFLP genotyping assays that use either a mismatched recognition sequence in one or both of the oligonucleotides, or incorporate a second native site within the PCR amplicon, provide the mechanism by which efficiency of restriction enzymes can be assessed with every sample. The outcome is confirmation of the activity of the discriminating enzyme regardless of genotype.

Role of transmembrane segment 10 in efflux mediated by the staphylococcal multidrug transport protein QacA.

The staphylococcal multidrug exporter QacA confers resistance to a wide range of structurally dissimilar monovalent and bivalent cationic antimicrobial compounds. To understand the functional importance of transmembrane segment 10, which is thought to be involved in substrate binding, cysteine-scanning mutagenesis was performed in which 35 amino acid residues in the putative transmembrane helix and its flanking regions were replaced in turn with cysteine. Solvent accessibility analysis of the introduced cysteine residues using fluorescein maleimide indicated that transmembrane segment 10 of QacA contains a 20-amino-acid hydrophobic core and may extend from Pro-309 to Ala-334. Phenotypic analysis and fluorimetric transport assays of these mutants showed that Gly-313 is important for the efflux of both monovalent and bivalent cationic substrates, whereas Asp-323 is only important for the efflux of bivalent substrates and probably forms part of the bivalent substrate-binding site(s) together with Met-319. Furthermore, the effects of N-ethyl-maleimide treatment on ethidium and 4',6-diamidino-2-phenylindole export mediated by the QacA mutants suggest that the face of transmembrane segment 10 that contains Asp-323 may also be close to the monovalent substrate-binding site(s), making this helix an integral component of the QacA multidrug-binding pocket.