Duration of serum antibody responses following vaccination and revaccination of cattle with non-living commercial Pasteurella haemolytica vaccines

This study was designed to determine the duration of serum antibody responses to Pasteurella haemolytica whole cells (WC) and leukotoxin (LKT) in weanling beef cattle vaccinated with various non-living P. haemolytica vaccines. Serum antibodies to P. haemolytica antigens were determined periodically through day 140 by enzymelinked immunosorbent assays. At day 140, cattle were revaccinated, and antibody responses periodically determined through day 196. Three vaccines were used in two experiments (A and B), OneShot™, Presponse^® HP/tK, and Septimune^® PH-K. In general, all three vaccines between 7 and 14 days induced antibody responses to WC after vaccination. Antibodies to LKT were induced with OneShot and Presponse. Revaccination at days 28 and 140 usually stimulated anamnestic responses. Serum antibodies to the various antigens remained significantly increased for up to 84 days after vaccination or revaccination. The intensity and duration of antibody responses were variable depending on the experiment and vaccines used. Vaccination with OneShot usually stimulated the greatest responses to WC. Vaccination with OneShot or Presponse resulted in equivalent primary anti-LKT responses. In experiment B, spontaneous seroconversion was found in numerous calves on day 112. Revaccination of those cattle at day 140 resulted in markedly variable antibody responses such that several groups had no increase in antibody responses.     

Molecular and biochemical mechanisms ofPasteurella haemolyticaleukotoxin-induced cell death

Pasteurella haemolyticaleukotoxin (LKT) is a member of the RTX family of pore-forming toxins that kill bovine immune cells. Several studies have suggested that RTX toxins kill target cells by the induction of apoptosis. In the present study, BL3 bovine leukaemia cells were exposed to LKT and assessed by molecular and flow cytometric techniques that measure different aspects of apoptotic cell death. The intoxicated cells demonstrated morphological, light scatter and Hoechst 33258 staining characteristics consistent with cells undergoing apoptosis. The cells also exhibited internucleosomal DNA fragmentation and poly (ADP-ribose) polymerase (PARP) cleavage, both indicators of apoptosis. LKT-treated cells bound annexin-V-FITC indicating that phosphatidylserine groups were translocated from the inner to the outer leaflet of the cell membrane. The effect of LKT on cells was dose dependent and inhibitable by incubation with anti-LKT monoclonal antibody. Finally, an early step for induction of apoptosis appears to be the binding of LKT to a β2 integrin since pre-incubating cells with anti-β2 integrin antibodies inhibited LKT-induced apoptosis. This study provides new insights into understanding the pathogenesis of bovine pasteurellosis and could lead to the development of both preventative and therapeutic strategies for disease management.     

云南省普洱市思茅区大面积野鼠自毙疫情实验室检测结果与分析

目的 调查引起云南省普洱市思茅区大面积野鼠自毙疫情的病原体,以便为采取相应措施提供科学依据。方法 通过现场流行病学调查,采集119只自毙野鼠样本,解剖取脏器进行鼠疫耶尔森菌病原体分离、噬菌体裂解实验、F1抗原、caf1基因等病原学、免疫学及分子生物学检测,以排除鼠疫。同时,脏器样本接种绵羊血琼脂平板、巧克力平板、麦康凯平板、沙氏琼脂培养基等营养培养基和选择性培养基,分离培养细菌、真菌类病原体;选取10只组织腐败较轻的自毙鼠,按肺组织、脑组织、腹腔脏器组织分为3份样本后,研磨、离心,再以0.22μm滤膜过滤,收集离心上清液、滤膜滤出液和滤膜截留残渣共9份样本,腹腔接种雄性、体质量30~40g昆明小鼠,每组2只,每只0.5ml。另接种0.5ml Hank’s液2只鼠作对照。共接种昆明小鼠20只。死亡小鼠解剖观察病理变化,取脏器组织接种上述培养基进行细菌分离培养。结果 鼠疫耶尔森菌病原学、免疫学和分子生物学检测均为阴性;自毙野鼠脏器镜检、分离培养及接种实验动物均检出G- 短小杆菌,经梅里埃VITEK 2GN、GP卡、16S rRNA扩增鉴定为多杀巴斯德菌 (Pasteurella multocida,Pm) ,概率为99%;分离到的G+粗大杆菌为腊样芽孢杆菌,概率为99%。结论 多杀巴斯德菌可能是这次发生野鼠大面积死亡的病原菌。该病原体的生物学特性及危害值得进一步关注。     

Generation of an attenuated Salmonella-delivery strains expressing adhesin and toxin antigens for progressive atrophic rhinitis,and evaluation of its immune responses in a murine model

An expression/secretion plasmid containing genes encoding the FimA, CP39, PtfA, ToxA and F1P2 antigens associated with porcine pneumonic pasteurellosis and progressive atrophic rhinitis (PAR) was constructed and harbored in an attenuated Salmonella Typhimurium, which was used as the vaccine candidate. The immune responses induced by this delivery strain were investigated in a murine model. Each antigen secreted from the delivery strain was confirmed by Western blot analysis. Thirty BALB/c mice were divided equally into two groups; group A were intranasally inoculated with the mixture of the five delivery strains, and group B were inoculated with sterile PBS. In group A, all antigen-specific serum IgG were significantly increased compared to those of group B from the 2nd week post-inoculation (WPI) till the 8th WPI. All antigen-specific mucosal IgA in group A were also significantly greater than those of group B. In addition, the significant splenic lymphocyte proliferative responses, the elevations of CD3⁺CD4⁺, CD3⁺CD8⁺ and B-cell populations, and the induction of IFN-γ expression in group A were observed. In conclusion, the mixture of five delivery strains expressing specific antigen for these diseases was found to be capable of inducing significant humoral and cellular immune responses.     

2019-2020年酒泉市市售羊肉多杀性巴氏杆菌污染情况调查及分析

目的 调查酒泉市市售羊肉多杀性巴氏杆菌污染情况,为该病的防控提供科学依据。方法 于2019年3-9月和2020年5-9月从酒泉市不同地区随机采集市售羊肉样品共257份,对样品中多杀性巴氏杆菌进行分离鉴定及耐药性分析。结果 257份被检测样品中有16份样品(6.22%)为多杀性巴氏杆菌阳性,其中2019年和2020年调查样品检出率分别为9.17%和3.65%;且农贸市场来源样品检出率高于超市和屠宰场,分别为8.49%、4.55%和3.13%。PCR法鉴定结果共发现3种血清型,其中A型、B型和D型所占比例分别为56.25%、31.25%和12.50%;分离获得菌株对14种常见抗生素耐药率为0.0%~100.0%,对林可霉素和甲氧嘧啶耐药率最高,分别为100.0%和81.25%。结论 酒泉市市售羊肉多杀性巴氏杆菌污染情况较为严重,且分离菌株对部分抗生素耐药性严重,相关部门应引起重视。     

Frontal osteomyelitis (Pott��s puffy tumour) associated with Pasteurella multocida�CA case report and review of the literature

A 58-year-old woman presented with progressive midforehead swelling and erythema with frontal headache. Investigations revealed erosion of the anterior wall of the frontal sinus with subgaleal abscess formation, establishing a diagnosis of Pott’s puffy tumour. Pasteurella multocida was isolated in pure growth from an aspirate of the abscess. P multocida is a rare cause of sinusitis. It is isolated from the respiratory tract of asymptomatic individuals and, more commonly, patients with chronic respiratory conditions. Although a cause of osteomyelitis associated with animal bites or scratches, P multocida has not previously been implicated as a cause of frontal osteomyelitis or Pott’s puffy tumour. A review of reported cases of Pott’s puffy tumour, including clinical presentation, microbiology, treatment and outcome, is provided.     

Development of immunization trials against Pasteurella multocida

Pasteurellosis is one of the most important respiratory diseases facing economically valuable farm animals such as poultry, rabbit, cattle, goats and pigs. It causes severe economic loss due to its symptoms that range from primary local infection to fatal septicemia. Pasteurella multocida is the responsible pathogen for this contagious disease. Chemotherapeutic treatment of Pasteurella is expensive, lengthy, and ineffective due to the increasing antibiotics resistance of the bacterium, as well as its toxicity to human consumers. Though, biosecurity measures played a role in diminishing the spread of the pathogen, the immunization methods were always the most potent preventive measures. Since the early 1950s, several trials for constructing and formulating effective vaccines were followed. This up-to-date review classifies and documents such trials. A section is devoted to discussing each group benefits and defects.     

禽巴氏杆菌OmpH外膜蛋白DNA疫苗的免疫效果

目的:研究禽多杀性巴氏杆菌omph DNA疫苗在小鼠体内诱导的免疫保护效果。方法:利用PCR技术扩增出禽多杀性巴氏杆菌的omph基因片段,克隆到pMD18-T上,再亚克隆到真核载体pcDNA3.1(+)上,构建重组质粒pOMPH,体外转染SP2/0细胞,RT-PCR、间接免疫荧光试验、Western blot检测其转录及表达情况。然后进行动物免疫,实验动物分为三组即:pOMPH组、pCDNA3.1(+)空载体组和PBS对照组,每组16只BALB/c小鼠,pOMPH组和pCDNA3.1(+)组以100μg/只的剂量肌注免疫,PBS组每只小鼠肌注100μl1×PBS,各组均进行了三次免疫,每次间隔两周。间接ELISA检测免疫后小鼠血清特异性抗体水平,MTT法检测免疫小鼠脾淋巴细胞增殖情况,三免两周后检测脾淋巴细胞IFN-γ分泌情况。强毒攻击,计算小鼠存活数目及保护率。结果:间接免疫荧光试验、Western blot和RT-PCR检测结果均表明pOMPH可在体外培养的SP2/0细胞中表达目的蛋白。动物免疫后,pOMPH组免疫小鼠血清抗体水平持续上升,与pCDNA3.1(+)组和PBS组相比差异极显著(P0.01)。经提取的外膜蛋白(Omps)刺激后,pOMPH组的刺激值(SI值)与pCDNA3.1(+)组及PBS免疫组相比均差异显著(P0.05)。免疫小鼠脾细胞产生的IFN-γ极显著高于两对照组产生的IFN-γ(P0.01)。攻毒后pOMPH组保护率明显高于两对照组,可达70%。结论:成功构建了禽多杀性巴氏杆菌omph DNA疫苗,该疫苗可诱导免疫小鼠产生较强的体液和细胞免疫应答及较好的保护效果。