狐、貉和水貂犬瘟热病毒受体SLAM 的基因克隆及其真核表达

信号淋巴激活分子(SLAM)为犬瘟热病毒(CDV) 感染其宿主动物识别的细胞受体。本试验应用RT -PCR 从狐狸、貉和水貂的外周血淋巴细胞中克隆到其相应SLAM 基因。基因测序比较发现,狐狸、貉与同科的犬SLAM 基因编码区长度均为1 029 bp,核苷酸同源性高于98.6% ;而水貂SLAM 基因编码区长度为1 020 bp,与以上三种动物遗传关系较远(核苷酸同源性< 83.7%),但与海豹SLAM 基因遗传关系较近(核苷酸同源性为91.4% )。基于不同动物SLAM 基因序列的系统进化树分析显示,犬、狐狸、貉、水貂和海豹在进化树上构成了以CDV 为感染宿主的遗传分支。氨基酸序列比较显示,该5 种动物SLAM 分子上均存在一个长度为26 个氨基酸的信号肽序列,且在空间结构上影响宿主--病毒特异性的8 个关键氨基酸均完全保守。通过构建表达该狐狸、貉、水貂SLAM 基因的三种真核表达质粒,分别转染CRFK 细胞后,应用CDV 强毒感染试验证实,CDV 均能在三种转染细胞上产生明显的细胞病变效应(CPE),而未转染CRFK 细胞对照无CPE 产生,由此证实作为CDV细胞受体的狐、貉和水貂的SLAM,体外表达后能明显增强犬瘟热强毒株对非敏感细胞的感染能力。     

Detection of sex chromosome aneuploidy in dog spermatozoa by triple color fluorescence in situ hybridization

With the development of a direct visualization of sex chromosome in a single sperm by fluorescence in situ hybridization (FISH) technique, the frequency of aberration (aneuploidy) in spermatozoa in several mammals has been investigated. However, there is no report in the incidence of X-Y aneuploidy in the sperm population of dogs. Therefore, in this study, the aneuploidy in dog spermatozoa was examined by multicolor FISH using specific molecular probes for canine sex chromosomes and autosome. Semen from eight male Labrador retrievers was used as specimen. For decondensation of sperm nuclei, the specimen was treated with 1 M NaOH for 4 minutes at room temperature. Probes for chromosomes X, Y, and 1, labeled with SpectrumGreen, Cy3 and Cy5, respectively, were hybridized with decondensed spermatozoa. Fluorescence in situ hybridization signals in sperm heads were clearly detected in each specimen, regardless of the sperm donor. The FISH signal of at least one of the three probes was detected in all sperm heads examined. There was no significant difference between the theoretical ratio (50:50) and the observed ratio of X and Y chromosomes in spermatozoa of all the eight dogs. Mean percentage of sex chromosome aneuploidy was 0.127% (ranged between 0% and 0.316%). This percentage of canine sex chromosome aneuploidy was lower than the one reported in cattle, horses, river buffalo, and goats sperm, but higher than that observed in mice and sheep.     

Monophyly and affinities of albanerpetontid amphibians (Temnospondyli; Lissamphibia)

The Albanerpetontidae are Middle Jurassic-Miocene amphibians that have variously been regarded as caudates (salamanders), a clade distinct from caudates, or incertae sedis lissamphibians. Here I test for monophyly of the Albanerpetontidae and examine the affinities of the group, within the framework of a more inclusive Temnospondyli, by performing a cladistic analysis using 59 informative characters scored for four non-lissamphibian temnospondyl genera, stem- and crown-clade caudates, salientians (frogs), gymnophionans (caecilians), and the two recognized albanerpetontid genera Albanerpeton and Celtedens . Monophyly of the Albanerpetontidae is corroborated. I interpret synapomorphies of the marginal teeth (non-pedicellate; crowns chisel like, labiolingually compressed, with three mesiodistally aligned cuspules) in albanerpetontids as being associated with a shearing bite. Other synapomorphies evidently strengthened and increased the mobility of the skull, mandible, and cervical region for burrowing, feeding, or both. Nested sets of synapomorphies place the Albanerpetontidae within the Lissamphibia, as the sistertaxon of Caudata plus Salientia. None of the 17 characters previously advanced as albanerpetontid-caudate synapomorphies convincingly places the Albanerpetontidae within the Caudata or allies the two groups as sistertaxa. Albanerpetontids are better interpreted not as aberrant caudates, but as a distinct clade of lissamphibians in which numerous apomorphies are superimposed upon an otherwise primitive lissamphibian body plan.     

Quantitative trait loci mapping for canine hip dysplasia and its related traits in UK Labrador Retrievers

Background

Canine hip dysplasia (CHD) is characterised by a malformation of the hip joint, leading to osteoarthritis and lameness. Current breeding schemes against CHD have resulted in measurable but moderate responses. The application of marker-assisted selection, incorporating specific markers associated with the disease, or genomic selection, incorporating genome-wide markers, has the potential to dramatically improve results of breeding schemes. Our aims were to identify regions associated with hip dysplasia or its related traits using genome and chromosome-wide analysis, study the linkage disequilibrium (LD) in these regions and provide plausible gene candidates. This study is focused on the UK Labrador Retriever population, which has a high prevalence of the disease and participates in a recording program led by the British Veterinary Association (BVA) and The Kennel Club (KC).

Results

Two genome-wide and several chromosome-wide QTLs affecting CHD and its related traits were identified, indicating regions related to hip dysplasia.

Conclusion

Consistent with previous studies, the genetic architecture of CHD appears to be based on many genes with small or moderate effect, suggesting that genomic selection rather than marker-assisted selection may be an appropriate strategy for reducing this disease.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-833) contains supplementary material, which is available to authorized users.     

Altered miRNA expression in canine retinas during normal development and in models of retinal degeneration

Background

Although more than 246 loci/genes are associated with inherited retinal diseases, the mechanistic events that link genetic mutations to photoreceptor cell death are poorly understood. miRNAs play a relevant role during retinal development and disease. Thus, as a first step in characterizing miRNA involvement during disease expression and progression, we examined miRNAs expression changes in normal retinal development and in four canine models of retinal degenerative disease.

Results

The initial microarray analysis showed that 50 miRNAs were differentially expressed (DE) early (3 vs. 7 wks) in normal retina development, while only 2 were DE between 7 and 16 wks, when the dog retina is fully mature. miRNA expression profiles were similar between dogs affected with xlpra2, an early-onset retinal disease caused by a microdeletion in RPGRORF15, and normal dogs early in development (3 wks) and at the peak of photoreceptor death (7 wks), when only 2 miRNAs were DE. However, the expression varied much more markedly during the chronic cell death stage at 16 wks (118 up-/55 down-regulated miRNAs). Functional analyses indicated that these DE miRNAs are associated with an increased inflammatory response, as well as cell death/survival. qRT-PCR of selected apoptosis-related miRNAs (“apoptomirs”) confirmed the microarray results in xlpra2, and extended the analysis to the early-onset retinal diseases rcd1 (PDE6B-mutation) and erd (STK38L-mutation), as well as the slowly progressing prcd (PRCD-mutation). The results showed up-regulation of anti-apoptotic (miR-9, -19a, -20, -21, -29b, -146a, -155, -221) and down-regulation of pro-apoptotic (miR-122, -129) apoptomirs in the early-onset diseases and, with few exceptions, also in the prcd-mutants.

Conclusions

Our results suggest that apoptomirs might be expressed by diseased retinas in an attempt to counteract the degenerative process. The pattern of expression in diseased retinas mirrored the morphology and cell death kinetics previously described for these diseases. This study suggests that common miRNA regulatory mechanisms may be involved in retinal degeneration processes and provides attractive opportunities for the development of novel miRNA-based therapies to delay the progression of the degenerative process.

Electronic supplementary material

The online version of this article (doi:10.1186/1471-2164-15-172) contains supplementary material, which is available to authorized users.     

湖北郧西黄龙洞更新世晚期人类牙齿

对2004—2006年在湖北省郧西县黄龙洞发现的7枚更新世晚期人类牙齿进行了观测与分析, 在此基础上与相关的化石人类及近代现代人类标本进行了对比。本研究发现:黄龙洞人类牙齿总体特征与现代人接近, 同时也保留部分可能属于更新世晚期人类的特点, 包括前部牙齿(侧门齿与犬齿)尺寸及粗壮程度都明显大于现代人。黄龙洞人类牙齿呈现的铲形门齿、双铲形门齿及臼齿釉质延伸说明,当时人类已经具有了东亚人群的典型牙齿形态特征。     

Number of teeth and risk of root caries

Objective: This study aimed to investigate the association between number of natural teeth and prevalence of root caries. Design: A cross‐sectional design was adopted. A stratified multi‐stage purposive sampling method was used to select a wide range of elderly people from all social classes, educational levels and sexes. Data was collected through interviews and clinical examinations. Setting: Urban area in Chiang Mai‐Thailand. Participants: 549 elderly dentate aged 60‐74 years. Main Outcome Measures: Teeth with decay and filled roots (DF‐T). Results: The prevalence of root caries was 18.2% with a mean DF‐T of 0.58 (S.D. 2.02). Results of multiple logistic regression showed that number of teeth, DMF‐S scores, sex, and socio economic factors were statistically significantly associated with root caries (P<0.05). Adjusted odds ratios demonstrated a 14% increase in the chance of having root caries when the number of teeth increased one unit. Similarly, a 3% increase was observed for each unit increase in the DMF‐S scores (p<0.001). Being male, having more than four years of education and earning more than 1500 baht/month increased the chances of root caries by 76%, 139% and 85% respectively (p<0.001). Recession and age were not statistically significantly associated with root caries. Conclusions: Having more teeth, higher DMF‐S scores, earning more than 1500 baht/month, having more than 4 years of education and being male increased the likelihood of having root caries.     

大熊猫犬瘟热病毒附着或血凝蛋白基因的序列分析

首次对犬瘟热病毒（ＣＤＶ）大熊猫（ＧＰ）毒株附着或血凝蛋白（Ｈ）基因进行了序列测定并与疫苗株Ｏｎｄｅｒｓｔｅｐｏｏｒｔ进行了比较。我们设计合成了４对引物,对ＧＰ株进行了ＲＴ－ＰＣＲ扩增与测序。Ｈ蛋白基因全长为１９４６ｂｐ,开放阅读框架（ＯＲＦ）始于２１－２３位的ＡＴＧ,终止于１８４２－１８４４位的ＴＧＡ,编码６０７个氨基酸,该基因序列已被ＧｅｎＢａｎｋ。将ＧＰ毒株与ＧｅｎＢａｎｋ中疫苗弱毒株Ｏｎｄ     

表达狂犬病毒糖蛋白抗原的重组犬2型腺病毒的构建及鉴定

为研制一种预防犬科动物狂犬病的新型疫苗,将含有狂犬病毒ERA株糖蛋白基因(Rabies glycoprotein,Rgp)表达盒的穿梭质粒pVAXΔE3Rgp中的Rgp表达盒克隆入犬2型腺病毒(Canine adenovirus type2,CAV2)骨架质粒pPoly2-CAV2中,获得重组质粒pPoly2-CAV2-ΔE3-Rgp,释放其基因组,转染MDCK细胞系,获得E3缺失区(Deletion of early protein3,ΔE3)含有Rgp表达盒的重组病毒CAV2-ΔE3-Rgp。该重组病毒能在MDCK细胞上产生典型的腺病毒细胞病变。通过酶切、PCR、基因测序,表明该重组病毒含有完整的Rgp表达盒。通过RT-PCR、Western blot等检测,表明该重组病毒能够表达Rgp抗原。用该重组病毒免疫犬,3次免疫后,可以诱导犬产生特异的抗CAV2HI抗体,其效价超过1∶256和抗狂犬病病毒(Rabies virus,RV)中和抗体,其效价超过0.50IU/mL。试验结果表明,获得的重组病毒免疫犬后,能够产生抗狂犬病毒和腺病毒的高效价保护性抗体,是一种有潜力的犬科动物狂犬病毒腺病毒二联疫苗候选株。