Molecular diversity of Brazilian strains of porcine circovirus type 2 (PCV-2)

Porcine circovirus 2 (PCV-2) is associated with a broad range of syndromes. In this study, eight pig tissue samples from two Brazilian states were analyzed using six PCR primer pairs amplifying a 1705-bp fragment of the PCV-2 genome. The NJ distance-based method was used for the phylogenetic analysis with the eight field strains herein, 15 GenBank sequences and using PCV-1 as an out-group. This yielded two major clusters (A and B) for this viral species, with the Brazilian strains segregating with European and Asian sequences. Nucleotide identity was 99.7 to 100% among the sequences. This information can be used in further studies of pathogenesis related to PCV-2 in Brazil.     

High frequency of porcine norovirus infection in finisher units of Brazilian pig-production systems

Tropical Animal Health and Production - Norovirus (NoV) is a member of the Caliciviridae family and is considered an emerging human enteric pathogen. NoVs are detected in farm animals such as...     

猪沙波病毒抗体间接ELISA检测方法的建立及初步应用

以原核表达重组蛋白作为抗原建立了猪沙波病毒(SaV)抗体的间接ELISA检测方法。使用矩阵滴定法确定抗原包被浓度为6.4μg/mL,血清稀释倍数为1∶50。试验选用1%BSA作为封闭液,血清和二抗的最佳作用时间分别为60 min和40 min。选择无致癌的TMB为底物,确定TMB最佳反应时间为10 min。通过对20份猪阴性血清样品的检测,计算阳性判定值为0.203。通过交叉试验、批内和批间重复性试验证明,本研究建立的SaV间接ELISA检测方法具有特异性高,重复性好的特点,可用于猪沙波病毒抗体的检测。     

Prevalence and genetic diversity of Anaplasma marginale strains in cattle in South Africa

Bovine anaplasmosis, caused by the tick-borne rickettsia Anaplasma marginale, is endemic in South Africa and results in considerable economic loss to the cattle industry. This study was designed to characterize strains of A. marginale at the molecular level from cattle raised in communal and commercial farms in the north-eastern and south-western regions of the Free State Province, South Africa, that varied in rainfall and vegetation. Seroprevalence to A. marginale was determined in 755 cattle by an Anaplasma spp. competitive enzyme-linked immunosorbent assay and ranged from 44% to 98% and was similar in both regions. While Anaplasma centrale was not targeted in this study, A. marginale infections were identified by species-specific msp1alpha polymerase chain reaction in 129 of 215 of the samples studied. Similar genetic diversity of A. marginale strains was found in both the north-eastern and south-western regions. The sequences of 29 A. marginalemsp1alpha amplicons from South African strains revealed considerable genetic diversity providing 14 new repeat sequences. However, 42% of MSP1a repeat sequences were not unique to this region. These results indicated the presence of common genotypes between South African, American and European strains of A. marginale. Cattle movement between different parts of South Africa was suggested by the presence of identical A. marginale MSP1a genotypes in north-eastern and south-western regions of the Free State Province. Control strategies for anaplasmosis in South Africa should therefore be designed to be protective against genetically heterogeneous strains of A. marginale.     

Genetic diversity of porcine reproductive and respiratory syndrome virus strains circulating in Hungarian swine herds

Analysis of 37 ORF5 sequences of Hungarian porcine reproductive and respiratory syndrome virus (PRRSV) strains revealed that most of them (35) belonged to the European genotype, forming distinct subgroups, reflecting the exceptional diversity of Eastern European strains. Twelve vaccine-like strains were also found in non-vaccinated animals. Two strains belonged to the American genotype showing 90-91% nucleotide identity to the "Quebec" Canadian reference strain. The analysis of the putative ectodomains and their N-linked glycosylation sites of the vaccine strain and its variants suggested selective pressure on the first ectodomain, by a consistent amino acid change on epitope B and by loosing a glycosylation site in the otherwise conserved N-46 position.     

河北省猪圆环病毒3型流行病学调查及遗传变异分析

为了解猪圆环病毒型(porcine circovirus3,PCV3)在河北省的流行情况及遗传变异特点,本研究在2017年2月至2018年7月间,从河北省不同地区60个猪场采集了310份不同临床症状表现的发病猪组织样本,应用PCR方法对其进行检测。结果显示该地区PCV3阳性率为16.5%(51/310),猪场阳性率为45.0%(27/60),对不同临床症状发病率分析显示PCV3在表现为繁殖障碍及腹泻猪群中具有较高的阳性率,且易与PCV2发生共同感染。对获得的8株PCV3的cap基因进行测序及同源性分析显示,所得8条序列之间的核苷酸相似性为98.0%～99.7%,与24条国内外参考株之间的核苷酸相似性为98.0%～100.0%。对cap基因所推导的氨基酸序列进行分析发现cap基因的主要突变位点发生在24,27,77及150位,其中24和27位主要发生了A24V和R27K变异。对32条序列进行进化树的构建发现,进化树可分为3a、3b和3c 3个基因群,河北分离株主要分布于3a基因群和3c基因群,表明了河北省PCV3流行株相对稳定。我们的研究结果为PCV3在河北省的流行及进化特点提供了理论参考,这些数据可为该地区PCV3的诊断及综合防控提供科学依据。     

High Mycobacterium bovis genetic diversity in a low prevalence setting

The genetic diversity among South African Mycobacterium bovis isolates from cattle was determined by genetic fingerprinting. The restriction fragment length polymorphism (RFLP) markers IS6110 and polymorphic GC-rich sequence (PGRS) as well as spoligotyping and determination of variable number of tandem repeats (VNTR) were used to characterize sub samples of 91 M. bovis field isolates. PGRS RFLP was the single most discriminatory method and combinations of typing methods, which included IS6110 and/or PGRS had the highest discriminatory power, able to reveal 29 distinct genotypes among 35 farms with no epidemiological link. Three of the farms were co-infected with two genetically unrelated strains. In contrast to reports from European and also other colonised countries on the African continent our findings are suggestive of a high genetic diversity of M. bovis in South Africa's cattle population, implying a variety of unrelated ancestor strains. Despite effective intervention through test-and-slaughter campaigns no indication of a 'founder effect' was apparent in the panel of isolates derived from all infected provinces.     

RT-PCR test for detecting porcine sapovirus in weanling piglets in Hunan Province, China

The prevalence of porcine sapovirus infection in weanling pigs was investigated in Hunan Province, China, between August 2006 and October 2007. A total of 153 diarrheic fecal samples from ten intensive pig farms from ten representative administrative regions in Hunan province were examined for porcine sapoviruses using RT-PCR. Twenty-two of 153 (14.37?%) samples were found to contain porcine sapoviruses. Phylogenetic analysis showed that all the porcine sapovirus isolates in Hunan Province belonged to the porcine sapovirus genogroup III. The results of the present investigation have implications for the control of porcine sapovirus infection in pigs in Hunan Province, China.     

Seroprevalence and molecular detection of porcine sapovirus in symptomatic suckling piglets in Guangdong Province,China

The seroprevalence and genetic identification of sapovirus (SaV) in symptomatic suckling piglets were investigated in Guangdong Province, China, between November 2011 and April 2013. Serum (n?=?960) and diarrheic fecal (n?=?101) samples collected from symptomatic suckling piglets in Guangdong Province were evaluated for antibodies against SaV using indirect enzyme-linked immunosorbent assay (iELISA). The overall seroprevalence of SaV in symptomatic suckling piglets was 61.9 % (594/960). Positive animals were found in all regions with seroprevalence ranging from 52 to 67.8 %, but the difference was not statistically significant (P?>?0.05). In addition, RNA of SaV was extracted from diarrheic fecal samples, and the partial polymerase gene was amplified by RT-PCR and then sequenced. Seven of 101 (6.9 %) samples were found to contain porcine SaV. Phylogenetic analysis showed that all the porcine SaV isolates belong to the porcine SaV genogroup III (GIII). This is the first report of SaV seroprevalence in symptomatic pigs in China.     

猪TLR2基因Bi-PASA遗传标记的建立及多态性研究

根据人和小鼠TLR2基因序列设计了特异性PCR引物,优化PCR条件后扩增到中国梅山猪、欧洲约克夏猪、PIC-2系及PIC-3系商品猪的TLR2基因690 bp的基因片段,并对其进行序列分析。序列分析结果显示,猪TLR2基因多态性程度低,发现在猪TLR2基因编码区第1 255位点上存在1个单碱基突变位点。利用双向特定等位基因PCR扩增法(B i-PASA)建立了检测猪TLR2基因变异的遗传标记。利用猪TLR2基因的B i-PASA标记,分析了TLR2基因在大河乌猪、撒坝猪和黔邵花猪中的基因频率和多态性。研究建立的B i-PASA遗传标记和基因变异信息,将为进一步分析猪TLR2基因变异与疾病抵抗力及经济性状的相关分析提供基础资料。     

Detection and genetic diversity of porcine rotavirus A,B and C in eastern Australian piggeries

Rotaviruses (RV) have a high prevalence in piggeries worldwide and are one of the major pathogens causing severe diarrhoea in young pigs. RV species A, B, and C have been linked to piglet diarrhoea in Australian pig herds, but their genetic diversity has not been studied in detail. Based on sequencing of the structural viral protein 7 (VP7) RVA G genotypes G3, G4 and G5, and RVC types G1, G3, G5, and G6 have been identified in Australian piggeries in previous studies. Although occurrence of RVB was reported in Australia in 1988, no further genetic analysis has been conducted. To improve health management decisions in Australian pig herds, more information on RV prevalence and genetic diversity is needed. Here, 243 enteric samples collected from 20 pig farms within Eastern Australia were analysed for the presence of RV in different age groups using a novel PCR-based multiplex assay (Pork MultiPath™ enteric panel). RVA, RVB, and RVC were detected in 10, 14, and 14 farms, respectively. Further sequencing of VP7 in selected RV-positive samples revealed G genotypes G2, G5, G9 (RVA), G6, G8, G14, G16, G20 (RVB), and G1, G3, G5, G6 (RVC) present. RVA was only detected in young (<10 weeks old) pigs whereas RVB and RVC were also detected in older animals (>11 weeks old). Interestingly, RVB and RVC G-type occurrence differed between age groups. In conclusion, this study provides new insights on the prevalence and diversity of different RV species in pig herds of Eastern Australia whilst demonstrating the ability of the Pork MultiPath™ technology to accurately differentiate between these RV species.     

Genetic diversity of porcine sapoviruses

Sapoviruses (SaVs) within the Caliciviridae family are an important cause of gastroenteritis in both humans and animals. Although the widespread occurrence of divergent human SaV strains has been reported, there have only been a few studies of porcine SaVs examining their genetic diversity. The aim of this study was to assess the genetic diversity of porcine SaVs in piglets with diarrhea in South Korea. Two hundred and thirty-seven fecal specimens from piglets with diarrhea were examined from 78 farms over a 2-year period from six provinces in South Korea. Overall, 69 (29.1%) of the samples from five provinces tested positive for porcine SaVs by either RT-PCR or nested PCR with the primer pairs specific to porcine SaVs. An analysis of the partial capsid gene (757bp) of 12 porcine SaVs detected from fecal samples showed genetic divergence, not only among the Korean porcine SaVs (67.7-99.1%), but also between Korean and American porcine SaVs (69.1-90.2%). Interestingly, one strain (Po/SaV/JN-MA113/05/Korea), formed a second porcine SaV/GIII genotype, and is tentatively called GIII/2. This strain had a 0.236-0.405 inter-cluster distance with the other strains in the same genogroup, which is comparable to the distances between the established GI and GII SaVs. Furthermore, two potential recombinant porcine SaVs were identified. In conclusion, porcine SaV infections are common in diarrheic piglets in South Korea. The infecting strains are genetically diverse, and include a newly recognized genotype and recombinant viruses within genogroup III.     

Unexpected detection of porcine rotavirus C strains carrying human origin VP6 gene

Background: Rotavirus C (RVC), a known etiological agent of diarrheal outbreaks, mainly inflicts swine population globally with sporadic incidence in human, cattle, ferret, mink and dog.

Objective: To demonstrate the presence of RVC in Indian swine population and characterization of its selected structural (VP6) and non-structural (NSP4 and NSP5) genes.

Methods: A total of 108 diarrheic samples from different regions of India were used. Isolated RNA was loaded onto polyacrylamide gel to screen for the presence of RVs through the identification of specific electrophoretic genomic migration pattern. To characterize the RVC strains, VP6 gene and NSP4 and NSP5 genes were amplified, sequenced and analyzed.

Results: Based on VP6 gene specific diagnostic RT-PCR, the presence of RVC was confirmed in 12.0% (13/108) piglet fecal specimens. The nucleotide sequence analysis of VP6 gene, encoding inner capsid protein, from selected porcine RVC (PoRVC) strains revealed more than 93% homologies to human RVC strains (HuRVC) of Eurasian origin. These strains were distant from hitherto reported PoRVCs and clustered with HuRVCs, owning I2 genotype. However, the two non-structural genes, i.e. NSP4 and NSP5, of these strains were found to be of swine type, signifying a re-assortment event that has occurred in the Indian swine population.

Conclusion: The findings indicate the presence of human-like RVC in Indian pigs and division of RVC clade with I2 genotype into further sub-clades. To the best of our knowledge, this appears to be the first report of RVC in Indian swine population. Incidence of human-like RVC VP6 gene in swine supports its subsequent zoonotic prospective.     

猪圆环病毒2型山西分离株的遗传变异多样性分析

为了进一步研究山西猪圆环病毒2型(PCV2)的遗传变异多样性情况,对山西2013—2016年间分离的9株猪圆环病毒流行株(SXQX、SXCZ、SXJC、SXJX、SXLL、SXPY、SXPG、SXXY、SXTY2株)的全基因组进行了扩增、克隆和测序,已收录入GenBank中。并将其全基因序列、ORF2序列和推导出的Cap蛋白氨基酸序列与不同国家和地区的28株主要流行株进行了遗传变异多样性分析,结果显示:9株PCV2山西流行株全基因组核酸序列全长SXJX、SXXY、SXJC株为1 768bp,其余均为1 767bp,分别约占33%和67%。9株PCV2流行株之间核苷酸同源性为94.7%~99.8%,与国内外28株参考株同源性为93.9%~99.9%,与国产疫苗株同源性(LG、DBN-SXO7-2、SH株)为95.1%~99.8%;PCV2全基因组序列的进化分析表明:本试验分离的SXJX、SXJC、SXXY株属于PCV2a亚型的2D亚群,SXLL、SXPY、SXCZ株属于PCV2b亚型的1C亚群,SXTY14、SXQX、SXPG株属于PCV2b亚型的1A/1B亚群,可见近几年,山西PCV2流行株以PCV-2b亚型为主。分离的9株PCV2的ORF2之间核苷酸和Cap蛋白氨基酸同源性分别为90.0%~100.0%和87.1%~100.0%,与28株参考株之间核苷酸和氨基酸同源性分别为87.6%~100.0%和84.1%~100.0%,与国产疫苗株核苷酸同源性和氨基酸同源性分别为91.0%~100.0%和89.3%~100.0%,同源性都较高。9株分离株中,有6株分离株(SXQX、SXTY14、SXPG、SXXY、SXJC、SXJX)Cap蛋白氨基酸为233个,其中SXQX、SXTY14、SXPG株的ORF2位于1 033~1 734bp,SXXY、SXJC、SXJX株的ORF2位于1 034~1 735bp,3株分离株(SXCZ、SXLL、SXPY)Cap蛋白氨基酸为234个,其ORF2位于1 030~1 734bp,而与其他基因组序列为1 767bp分离株的ORF2相比,1 030~1 734bp位置多了3个碱基TCA,导致其氨基酸序列在234位置增加1个K(Lys)。另外还发现9株PCV2Cap蛋白除了唯一糖基化位点(NYS)(第143~145位氨基酸)呈高度保守外,还存在较多的氨基酸变异,这为山西地区PCV2的免疫预防等的研究提供理论依据,并为深入开展PCV2的分子致病机理提供基础理论数据。     

种群遗传变异及基因多样度分析

阐述了种群遗传学和生态遗传学研究的基本单位－种群的定义和意念含义,其一是种系变异含义,其二是生态适应含义,并概述了遗传变异的研究历史。从３个方面及质量性状变异,数量性状变异与基金变异,介绍了种群遗传变异的研究方法,重点介绍了基因变异的测量和亚种群基因多样度的测量,并且依据研究所得的遗传参数对中国苜蓿不同种群遗传结构和多样度进行了例证分析。     

Phylogenetic analysis of porcine rotavirus in Argentina: increasing diversity of G4 strains and evidence of interspecies transmission

Group A rotaviruses are one of the most frequently detected viral agents associated with neonatal diarrhea in piglets. In order to characterize rotavirus (RV) strains circulating in Argentinean swine, four porcine production farms located in Buenos Aires were studied. RV strains genotyped as P[6]G4, P[6]G8 and P[1]G6 were found in piglets under 30 days of age, without diarrhea. Phylogenetic and sequence analysis of the VP7 gene from G4 strains available in databases, reveals five porcine new lineages (III-VII) and three sublineages (VIIa-VIIc). The G4 porcine Argentinean strains were grouped with a porcine RV strain isolated in Brazil and another RV strain isolated from a child with diarrhea in Mexico, constituting an American lineage (VII). On the other hand, porcine G6 and G8 were closely related to RV's circulating in Argentinean cattle and South-American camelids, respectively. The fact that G4 porcine lineages were epidemiologically related to human strains, and G6 and G8 Argentinean porcine strains were found related to bovine and South-American camelids, respectively, suggests that pigs might play a crucial role as reservoir and generator of newly adapted emerging RV strains for human and other species.     

2012年猪嵴病毒流行病学调查及3D基因的遗传变异分析

为了解猪嵴病毒(PKV)的流行及变异情况,本研究根据PKV的3D基因保守区域核苷酸序列设计一对特异性引物,并利用RT-PCR方法对2012年来自24个省市84个猪场的病料样品共计236份进行3D基因的检测。结果显示:在236份病料中,共有63份为PKV阳性,阳性率为36.69%;而在检测的84个猪场中,共有36个猪场显示PKV阳性,猪场阳性率为42.85%。此外,对2012年分离的9个PKV部分3D基因进行测序分析,并与GenBank中登录的人、牛、猪以及鼠等嵴病毒株相关序列进行遗传变异分析。结果表明9个PKV 3D基因与国内外其他PKV株无论是在核苷酸水平上还是在氨基端水平上均具有较高的同源性,表明PKV的3D基因具有较高的保守性,可以作为病原检测的靶基因。