Objective:To evaluate lung protection of remote limb ischemic preconditioning after pulmonary resection.Methods:Methods sixty adult patients scheduled for elective pulmonary resection, were randomly divided into control group(group C, 30 cases) and remote limb ischemic preconditioning(group RLIP, 30 cases) using a random number table. Before one-lung-ventilation(T0), at 30 minites, 1 hour and 2 hours of OLV(T1, 2, 3), 15minites after re-expansion of the collapsed lung(T4), blood samples were drawn from the radial artery and vein for blood gas analysis, alveolar-arterial oxygen gradient(A-aDO 2)、pulmonary shunt ratio(Qs/Qt)were calculated. Extraction time of closed thoracic drainage tube, length of hospital stay, the incidence of in-hospital complications after operation were recorded. Results:Compared to T0 , each group at T1-T4, A-aDO 2 were obviously increased. We found that at T3, A-aDO 2 of group C increased much more higher and statistically significant( P<0.05). Compared to group RLIP, Qs /Qt of group C were significantly increased at T2( P<0.05). Compared with C group, the expression of microtubule-associated protein 1 light chain 3B in lung tissues was significantly"up-regulated in RLIP group( P<0.05). Followed the extraction time of closed thoracic drainage tube, length of hospital stay, the incidence of in-hospital complications after operation there were no statistically different( P>0.05). Conclusion:Remote limb ischemic preconditioning had some protective effect after pulmonary resection, which mechanism may be related to enhancing autophagy in the operated 1ung tissues of the patients.

试验旨在探究VA5免疫增强剂对鸽新城疫病毒(NDV)灭活疫苗(ND4416株)的免疫增效作用.选取160只健康鸽随机分为4组:ND4416株灭活疫苗组(NDV组)、ND4416株灭活疫苗与免疫增强剂(VA5)混合组(NDV+VA5组)、La Sota灭活疫苗组(La Sota组)及生理盐水空白对照组(C组),进行免疫效力及免疫持续期试验.采用血凝抑制试验检测各组鸽免疫后不同时间点的血清抗体效价,结果显示,VA5免疫增强剂能够显著提高鸽NDV灭活疫苗血清抗体水平(P＜0.05).脾脏淋巴细胞转化试验结果显示,VA5能够有效刺激免疫鸽的淋巴细胞转化.在免疫后第30、90、180天,各组鸽子采用ND4416株进行攻毒试验,结果显示,NDV+VA5组在3个时间点的攻毒保护率均为100％,均高于La Sota组.免疫持续期试验结果显示,NDV+VA5组血清抗体水平在免疫后第21天达峰值11.201og2,第180天仍能达到7.501og2,攻毒保护率达100％,表明VA5免疫增强剂可延长鸽新城疫疫苗的免疫效力持续期至180 d.攻毒后体外排毒检测结果表明,VA5免疫增强剂能够缩短攻毒后鸽的体外排毒周期,减少体外排毒.综上,VA5免疫增强剂能够显著提高鸽NDV灭活疫苗的免疫效果,为研制效果更好的鸽NDV疫苗提供了基础依据,同时也为免疫增强剂的应用增加了试验数据.

非洲猪瘟(African swine fever,ASF)是由非洲猪瘟病毒(African swine fever virus,ASFV)感染家猪和野猪引起的一种高度传染性和致死性疫病.2018年8月1日,国家外来动物疫病中心接到辽宁省沈阳市发生疑似ASF病例的报告,经官方确认后于2018年8月3日向世界动物卫生组织(OIE)通报为我国首例非洲猪瘟.截至2019年4月22日,中国已相继暴发了129起ASF疫情,共计扑杀102万头猪.由于尚无针对ASF的有效疫苗,其传播媒介广泛分布及复杂的相互作用使得防控局势变得更为严峻.扑杀猪群和切断病毒的传播路径是目前减缓疫情传播的主要手段.因此,对ASF流行病学特点及其扩散因素的深刻理解,有助于病毒扩散模型的建立和进一步防控措施的实施.

为了建立检测猪伪狂犬病病毒(PRV)黏膜免疫时特异的IgA抗体的间接ELISA方法,本研究采用PCR方法扩增了PRV gB基因截短片段,并将其连入原核表达质粒pGEX-6P-1,构建的表达质粒转化至大肠杆菌Transetta(DE3)菌株后,采用IPTG进行诱导表达,得到gB重组蛋白的可溶性表达,并用谷胱甘肽亲和层析柱进行蛋白纯化.以纯化的gB重组蛋白作为包被抗原,以山羊抗猪IgA-HRP为二抗,建立了猪伪狂犬病病毒IgA的间接ELISA检测方法.经ELISA反应条件优化,确定抗原包被浓度为2.4μg/mL,封闭液为10 g/L明胶,一抗37℃孵育1.5h,二抗1:20 000稀释后37℃孵育0.5h,TMB显色液显色10 min.该方法的批内和批间变异系数均小于10％,重复性较好.利用该间接ELISA方法对PRV黏膜免疫的仔猪进行检测,结果可以在鼻腔黏液中检测出特异的IgA抗体.本试验为PRV黏膜免疫提供了初步检测方法.

In order to understand the effect of Cu2+-modified palygorskite on Cu2+ contents in the tissues of mice,we measured the concentrations of residual Cu2+ in feces,blood,hair and organs of 32 mice by ICP spectrometer.The results showed that in the experimental groups,the concentration of Cu2+ in feces tended to increase as the test went on,which suggested that there might be Cu2+-modified palygorskite residues in the stomach and intestines.The mice in all the experimental groups excreted 20％ ～ 30％ of Cu2+ mainly through feces,and there were almost no Cu2+ residue in their blood,hair and other organs.

BACKGROUND:Marek's disease virus (MDV) resides in the genus Mardivirus in the family Herpesviridae. MDV is a highly contagious virus that can cause neurological lesions, lymphocytic proliferation, immune suppression, and death in avian species, including Galliformes (chickens, quails, partridges, and pheasants), Strigiformes (owls), Anseriformes (ducks, geese, and swans), and Falconiformes (kestrels).CASE PRESENTATION:In 2015, two red-crowned cranes died in Nanjing (Jiangsu, China). It was determined that the birds were infected with Marek's disease virus by histopathological examination, polymerase chain reaction (PCR), gene sequencing and sequence analysis of tissue samples from two cranes. Gross lesions included diffuse nodules in the skin, muscle, liver, spleen, kidney, gizzard and heart, along with liver enlargement and gizzard mucosa hemorrhage. Histopathological assay showed that infiltrative lymphocytes and mitotic figures existed in liver and heart. The presence of MDV was confirmed by PCR. The sequence analysis of the Meq gene showed 100% identity with Md5, while the VP22 gene showed the highest homology with CVI988. Furthermore, the phylogenetic analysis of the VP22 and Meq genes suggested that the MDV (from cranes) belongs to MDV serotype 1.CONCLUSION:We describe the first molecular detection of Marek's disease in red-crowned cranes based on the findings previously described. To our knowledge, this is also the first molecular identification of Marek's disease virus in the order Gruiformes and represents detection of a novel MDV strain.

The aim of this research was to develop effective alternative therapies to reduce antibiotic use in animal agriculture. In this study, the efficacy of copper-modified palygorskite (CM-Pal) in preventing diarrhea caused by Salmonella was specifically examined both in vitro and in vivo. The CM-Pal was prepared with palygorskite (Pal) and copper nitrate. The antibacterial activity of the CM-Pal was detected by comparing the differences in cell numbers on plate count agar before and after adding the CM-Pal to Salmonella typhimurium cultures. Seventy ICR mice were then allocated into seven groups. Five groups (the treatment groups) were infected with S. typhimurium by intraperitoneal (i.p.) injection and were given Pal, CM-Pal, montmorillonite powder, gentamicin, and physiological saline, respectively. One group (the prevention group) was given CM-Pal before infection with S. typhimurium. Another group (the uninfected group) was not infected with S. typhimurium. The effects of Pal, CM-Pal, montmorillonite powder, and gentamicin on the treatment or prevention of diarrhea in the mice were examined by stool studies, fecal scoring, and assessment of growth performance and villus height. The CM-Pal had satisfactory anti-bacterial properties in vitro: the antibacterial rate was 100% after 2 h incubation with S. typhimurium NJS1 cultures (1×106 colony-forming units (CFU)/ml). In the in vivo experiment, the CM-Pal exerted superior effects in the treatment and prevention of diarrhea in mice compared with Pal, montmorillonite powder, and gentamicin. In the CM-Pal group, no mice showed signs of diarrhea at 24 h post infection (p.i.), and all mice fully recovered from infection. However, the Pal group, montmorillonite group, and gentamicin group only recovered after 48, 48, and 96 h, respectively. The villus height level in the CM-Pal treatment group recovered at 3 d p.i. However, the recovery time of the other groups was longer (at least 5 d). The CM-Pal prevention group had a better effect on weight gain than the other groups. This study suggested that CM-Pal may be an effective alternative to conventional antibiotics for the treatment and prevention of animal diarrhea caused by Salmonella.

The aim of this research was to explore methods of Babesia gibsoni artificial infection and to establish the basis of studying on canine babesiosis.Six heath domestic dogs were divided into two groups.Three dogs were carried out splenectomy and 3 dogs were no treated as control.All dogs were administered sickness dog blood which was infected by Babesia gibsoni.Clinical symptoms,laboratory examination(such as blood smear,complete blood count,PCR diagnosis of pathogen) were detected after infection.The results showed that the dogs infected with Babesia gibsoni by artificial method displayed the simile clinical symptoms with natural infected dogs.Babesia gibsoni could be found in blood smear.The dogs infected with B.gibsoni showed serious anemia according to results of complete blood count.The gene fragment of Babesia gibsoni could be detected in infected dogs using PCR method.Infection rate of red blood and degree of anemia in the splenectomy group were higher than control groups.It was suggested that the methods of Babesia gibsoni artificial infection was established successfully in this research.Splenectomy before infection was the better choice for Babesia gibsoni artificial infection.

The pigeon circovirus (PiCV) is a member of the genus Circovirus of the family Circoviridae.The PiCV is an immunosuppressive pathogen.In the present study,a polymerase chain reaction(PCR)specific to the PiCV was used to detect samples collected from racing pigeons in Jiangsu Province,China.Six out of 22 samples were positive in PCRs.Total DNA was extracted from the JS15-1 sample and the genome was amplified and sequenced.Complete genome sequences were submitted to the GenBank database (GenBank number KX431143).The full-length of the genome was 2034 bp and contained three main open reading frames.Homology analyses showed that the overall nucleotide identity between this genome and others accessed in the National Center for Biotechnology Information database is ranged from 85.3％ to 97.1％.The best identity was with Bel20,in which the identity of Rep gene nucleotides ranged from 92.1％ to 96.6％ and that of Cap gene nucleotides ranged from 72.2％ to 99.4％.A phylogenetic tree based on the full-length genome also showed that JS15-1 had the closest relationship with the Bel20 that emerged in Belgium in 2006.A phylogenetic tree based on the Rep gene showed that JS15-1 was located in the same branch as Bel20,PiCV/Japan/2010,and Ita4B.A phylogenetic tree based on the Cap gene showed that JS15-1 and Bel20 were in the same branch,and JS15-1 had the closest relationship with Bel20.Compared with Bel20,six amino-acid mutations were found at residues 8,10,53,69,95 and 202 in the Rep protein and one amino-acid was found at residue 33 in the Cap protein.A one amino-acid insertion was found at residue 34 in the Cap protein.These results suggest that JS15-1 may have originated from Belgium,and that JS15-1 was imported from a large number of breeders from Belgium within the last two decades.

The absence or deficiency of DNA mismatch repair (MMR) activity results in microsatellite instability (MSI) in cancer. The avian leukosis virus (ALV) causes neoplastic disease in chickens. In this study, the status of MMR, MSI, the cell cycle and apoptosis were detected in DF-1 cells after avian leukosis virus subgroup A infection. Flow cytometry analysis results indicated that there was no significant difference in cell apoptosis between the control and infected groups. The percentage of cells in S and G2 phases were increased in the infected group. MSI and mutation of MSH2 and MLH1 gene exons were absent in DF-1 cells after infection. Levels of MSH2 and MLH1 mRNA were dramatically increased in DF-1 cells after infection. These results demonstrated that ALV RAV-1 infection may promote the expression of MSH2 and MLH1 genes rather than resulting in gene mutations. Mismatch repair functions were normal and may be have relationships with the arrest of S phase and G2 phase.

The aim of this research was to develop effective alternative therapies to reduce antibiotic use in animal agriculture. In this study, the efficacy of copper-modified palygorskite (CM-Pal) in preventing diarrhea caused by Salmonella was specifically examined both in vitro and in vivo. The CM-Pal was prepared with palygorskite (Pal) and copper nitrate. The antibacterial activity of the CM-Pal was detected by comparing the differences in cell numbers on plate count agar before and after adding the CM-Pal to Salmonella typhimurium cultures. Seventy ICR mice were then allocated into seven groups. Five groups (the treatment groups) were infected with S. typhimurium by intraperitoneal (i.p.) injection and were given Pal, CM-Pal, montmorillonite powder, gentamicin, and physiological saline, respectively. One group (the prevention group) was given CM-Pal before infection with S. typhimurium. Another group (the uninfected group) was not infected with S. typhimurium. The effects of Pal, CM-Pal, montmorillonite powder, and gentamicin on the treatment or prevention of diarrhea in the mice were examined by stool studies, fecal scoring, and assessment of growth performance and villus height. The CM-Pal had satisfactory anti-bacterial properties in vitro: the antibacterial rate was 100% after 2 h incubation with S. typhimurium NJS1 cultures (1×106 colony-forming units (CFU)/ml). In the in vivo experiment, the CM-Pal exerted superior effects in the treatment and prevention of diarrhea in mice compared with Pal, montmorillonite powder, and gentamicin. In the CM-Pal group, no mice showed signs of diarrhea at 24 h post infection (p.i.), and all mice fully recovered from infection. However, the Pal group, montmorillonite group, and gentamicin group only recovered after 48, 48, and 96 h, respectively. The villus height level in the CM-Pal treatment group recovered at 3 d p.i. However, the recovery time of the other groups was longer (at least 5 d). The CM-Pal prevention group had a better effect on weight gain than the other groups. This study suggested that CM-Pal may be an effective alternative to conventional antibiotics for the treatment and prevention of animal diarrhea caused by Salmonella.

Copper-modified attapulgite clay was made of attapulgite clay and copper nitrate solution by ion exchange.The effects of copper-modified attapulgite clay on prevention and treatment of pullorum diseases were studied by mixing it with feed.In the preventive test,one hundred one dayold chickens were randomly divided into five groups,control group,infection group,No.1 prevention group,No.2 prevention group and chloramphenicol prevention group.Control group was fed normal diet without any treatment.Infection group was fed normal diet from 1st day to 15th day and infected with Salmonella pullorum at the 7th day.Prevention groups were given different levels of copper-modified attapulgite clay or chloramphenicol as prophylactic administration from 1st day to 15th day.Prevention groups were also infected with Salmonella pullorum at the 7th day.In the treatment test,one hundred one-day-old chickens were randomly divided into five groups,control group,infection group,No.1 treatment group,No.2 treatment group and chloramphenicol treatment group.Control group was fed normal diet without any treatment.Infection group was given normal diet and infected with salmonella pullorum at the 7th day.Treatment groups were infected with Salmonella pullorum at the 7th day and then given different levels of copper-modi fied attapulgite clay or chloramphenicol as treatment administration.During the test,morbidity,mortality,protection rate were monitored and recorded.Salmonella infection of cecum and liver were determined by SS plate culture.The weights of chicken and feed intake were recorded for measuring the growth performance.The morphology of jejunal was examined using optical microscope by histopathology.The results of prevention test showed that morbidity,mortality and isolating rate of Salmonella in live and caecum were decreased when adding copper-modified attapulgite clay in feed (1％).The growth performance of chickens was improved.The effect of 1％ coppermodified attapulgite clay group was better than other groups.The results of treatment test showed that the mortality and isolating rate of Salmonella in live and caecum were decrease when adding copper-modified attapulgite clay in feed (1.5％).The cure rate and growth performance of coppermodified attapulgite clay group were improved than other group.It was suggested that coppermodified attapulgite clay was characterized by antibacterial activity and protection of intestinal mucosa.It can prevent and treat pullorum diseases,reduce damage induced by salmonella pullorum and improve productivity performance of chicken.

One strain of pigeon Newcastle disease virus(NDV) was isolated from a diseased racing pigeon loft suspected to be infected with NDV in Nanjing.This NDV isolate was named Pi/NJ/CH/4416/2016 (ND4416) and purified by using specific pathogen-free(SPF) eggs.Biological tests showed that this isolate showed high pathogenicity to pigeons,but lower to chickens.The resulting RNA samples were converted to cDNAs,followed by PCR amplification and sequencing of F and HNgenes.The cleavage site of the fusion protein(F) was 112RRQKRF117,a feature generally associated with virulent NDV strains.Phylogenetic analysis based on F and HNgene sequences showed that this strain belonged to Class Ⅱ,Ⅵb subgenotype.In addition,Pi/NJ/CH/4416/2016 and other strains prevailing in China were more closely related to the PPMV-1/Belgium/11-09620/2011(JX901124.1) strain.It supported that epidemic pigeon NDV isolates in China might originate from Belgium.Pi/NJ/CH/4416/2016 had a far genetic relationship from the classic vaccine strain La Sota (AF077761.1).Bioinformatics analysis revealed that a linear epitope tope consisting of amino acids 491-500 of HNprotein of this pigeon strain was different from La Sota.In addition,hemagglutination inhibition test revealed a noticeable antigenic difference between them.

Porcine infectious anemia caused by Mycoplasma suis is a global disease and results in serious economic losses. To determine the prevalence of M. suis infection in eastern China, a cross-sectional serologic study was conducted with 3458 porcine serum samples randomly obtained from January 2014 to August 2016. The samples were tested with a blocking enzyme-linked immunosorbent assay developed in our laboratory. The seroprevalence was 33.3% in the complete sample set and was 25.9%, 37.8%, and 37.8% in 2014, 2015, and 2016, respectively. The seroprevalence was distinctly higher in summer (39.9%) and autumn (42.0%) than in spring (28.9%) and winter (23.3%). Shanghai was the region with the highest seroprevalence (54.2%) and Jiangsu the region with the lowest (23.0%). The seroprevalence was markedly higher in boars (47.1%), multiparous sows (47.0%), and replacement gilts (39.2%) than in piglets (24.2%), fattening pigs (17.2%), and nursery pigs (12.5%). These data demonstrate that the prevalence of M. suis infection is increasing yearly in eastern China.

The aim of this research is to study the transcription levels of the mismatch repair related genes of MMR (MSH2,MSH3,MLH1 and PMS1),BRCA1,p53 genes mRNA in DF-1 cells when infected with the avian leukosis virus subgroup J (ALV-J).DF-1 cells were infected with the ALV-J NX0101 strain and the p27 antigen was detected to confirm the infection.The total RNA was extracted to detect the expression of the mismatch repair related genes by real-time PCR.The transcription levels of MSH2,MSH3,MLH1,PMS1,and p53 mRNA were increased in DF-1 cells after ALV-J infection,but the levels of BRCA1 mRNA were decreased.These results demonstrated that MSH2,MSH3,MLH1,PMS1,BRCA1 and p53 genes response to the recognition and repair of DNA damage.This study provides a new idea for studying the mechanism of tumorigenesis induced by ALV-J.

江苏省某种鹅场15日龄雏鹅疑似感染鸭疫里氏杆菌,无菌采取脑、心脏、肝和脾等病料,进行病原分离纯化,通过细菌生化试验、血清型鉴定及设计特异性扩增鸭疫里氏杆菌外膜蛋白A(OmpA)基因的引物,进行PCR扩增,从8份病料中分离到2株血清2型鸭疫里氏杆菌.药物敏感性试验表明,2个分离株对红霉素和阿奇霉素的敏感性高于其他药物.对OmpA基因PCR产物进行基因序列测定,在NCBI上进行基因序列同源性比对,其基因序列与多株已发表的序列的同源性达到99％.进化分析表明,分离株均与D2、G2株具有很近的亲缘关系.

To evaluate the effect of immunopotentiator VA5 on immunity potency of inactive vaccine of pseudorabies virus (PRV), inactive vaccine was mixed with VA5. The comparison test of the immunity potency was conducted by using conventional inactive vaccine of Bartha-K61 strain of PRV, inactive vaccine mixed with VA5 and attenuated vaccine of Bartha-K61 strain. Blood samples were collected at 14 and 35 days after the first immunization, respec-tively. Neutralization experiments and cytokine detection were performed. The results showed that, compared with live and inactive vaccine of Bartha-K61 strain of PRV, the inactive vaccine with VA5 could induce higher neutrali-zing antibody against the heterologous strain of PRV and increase the expression levels of IL-2 and IFN-γ mRNA in porcine peripheral blood lymphocytes ( PBMC) . Neutralization test showed that VA5 could significantly increase the level of neutralizing antibody against the epidemic variation strain of PRV. This experiment showed that VA5 could improve the humoral and cellular immunity of inactive PRV vaccine, and provide a basis for paving the better im-mune effect of inactivated PRV vaccine.

为构建新型H7亚型禽流感HA基因的原核表达载体，并在大肠杆菌中进行表达。从NCBI数据库下载H7亚型禽流感HA全基因序列，合成HA基因；定向克隆到原核表达载体pET－32a（＋）的多克隆位点中，构建重组原核表达质粒 pET －32a（＋）－HA；转化入大肠杆菌 BL21（DE3）中经 IPTG 诱导表达，表达产物经 SDS －PAGE 和Western－blot法鉴定。结果显示，重组质粒经双酶切及基因测序鉴定构建正确；重组的HA融合蛋白约为84 ku，大小与预期融合蛋白大小一致；重组HA融合蛋白可以与His标签单克隆抗体特异性结合，与H7阳性血清有较强的反应原性。成功构建了HA基因原核表达载体，并在大肠杆菌BL21（DE3）中获得重组HA融合蛋白表达。

Koi herpesvirus (KHV) infection is associated with high mortalities in both common carp (Cyprinus carpio carpio) and koi carp (Cyprinus carpio koi) worldwide. Although acute infection has been reported in both domestic and wild common carp, the status of KHV latent infection is largely unknown in wild common carp. To investigate whether KHV latency is present in wild common carp, the distribution of KHV latent infection was investigated in two geographically distinct populations of wild common carp in Oregon, as well as in koi from an Oregon-based commercial supplier. Latent KHV infection was demonstrated in white blood cells from each of these populations. Although KHV isolated from acute infections has two distinct genetic groups, Asian and European, KHV detected in wild carp has not been genetically characterized. DNA sequences from ORF 25 to 26 that are unique between Asian and European were investigated in this study. KHV from captive koi and some wild common carp were found to have ORF-25-26 sequences similar to KHV-J (Asian), while the majority of KHV DNA detected in wild common carp has similarity to KHV-U/-I (European). In addition, DNA sequences from IL-10, and TNFR were sequenced and compared with no differences found, which suggests immune suppressor genes of KHV are conserved between KHV in wild common carp and koi, and is consistent with KHV-U, -I, -J.

The recombinant swine poxvirus rSPV/H3-2A-H1 co-expressing HA1 genes of H3N2 and H1N1 subtype SIV has been constructed and identified. Inoculations of rSPV/H3-2A-H1 yielded ELISA and neutralization antibodies against SIV H1N1 and H3N2, and elicited potent H1N1 and H3N2 SIV-specific INF-γ response from T-lymphocytes in mice and pigs in this study. Complete protection against SIV H1N1 or H3N2 challenge in pigs was observed.