[Objectives]This study was conducted to evaluate the practical efficacy of Haemophilus parasuis(Hps)vaccines and determine whether the continuous use of vaccines leads to changes in Hps serotypes.[Methods]A swine prod...[Objectives]This study was conducted to evaluate the practical efficacy of Haemophilus parasuis(Hps)vaccines and determine whether the continuous use of vaccines leads to changes in Hps serotypes.[Methods]A swine production group that had used a bivalent vaccine of Hps serotypes 4 and 5 for two consecutive years were investigated and retrospectively analyzed.Data on Hps detection rate,nursery production and disease indexes,as well as serotype shifts after continuous vaccination,were collected to assess vaccine effectiveness.[Results]Continuous vaccination reduced the total nursery culling and mortality rate by 1.75%.Hps infection persisted in the farms,and the pathogen could still be isolated.However,serotyping revealed a significant shift in the dominant prevalent serotypes,indicating that the vaccine was indeed effective.[Conclusions]The matched Hps vaccine demonstrated effective results.However,whether replacing serotypes or adding more prevalent serotypes could further improve the control of Hps disease requires further investigation.展开更多
[Objective] The study aimed to obtain attenuated strain of Haemophilus parasuis.[Method] Tn5 transposon technology was used to construct a library of mutants.Positive mutants were screened by kanamycin resistance.Fals...[Objective] The study aimed to obtain attenuated strain of Haemophilus parasuis.[Method] Tn5 transposon technology was used to construct a library of mutants.Positive mutants were screened by kanamycin resistance.False positive was identified by PCR and then removed.Mice were infected to detect the virulence of mutants.The bionomics of attenuated strains were detected,too.[Result] The attenuated mutants showed similar reproductive activity to that of wild strain.The virulence of mutants was still stable after 30 passages.[Conclusion] This study provided foundation for exploring the virulence factors and pathogenic mechanism of HPS.展开更多
[Objective] The aim was to clone and express the pilA gene of outer membrane protein of Haemophilus parasuis.[Method] The published pilA gene sequence of HPS was analyzed for primer synthesis,and the genome of serotyp...[Objective] The aim was to clone and express the pilA gene of outer membrane protein of Haemophilus parasuis.[Method] The published pilA gene sequence of HPS was analyzed for primer synthesis,and the genome of serotype 5-type of HPS was used as template for PCR amplification of the pilA gene of HPS;the recombinant expression plasmid was constructed and transformed into E.coli BL21（DE3）after induced by IPTG.SDS-PAGE and Western blot analysis were then carried out.[Result] The molecular weight of expressed protein was consistent with the expected（43 kD）.[Conclusion] The results provided a foundation for the preparation of subunit vaccine and diagnostic reagents.展开更多
To characterize the β-lactam resistance in veterinary clinical isolates of Haemophilus parasuis, 115 isolates were examined for the β-lactam resistance, the possession of β-lactamase, and the presence of β-lactama...To characterize the β-lactam resistance in veterinary clinical isolates of Haemophilus parasuis, 115 isolates were examined for the β-lactam resistance, the possession of β-lactamase, and the presence of β-lactamase genes. The genetic relationship among isolates was evaluated by pulsed-field gel electrophoresis （PFGE）. Overall, the commonly detected resistance phenotypes were resistant to ampicillin （26.09%）, penicillin （22.61%）, amoxicillin （21.74%）, cefazolin （14.78%）, cefaclor （12.17%）, and cefotaxime （6.96%）. These strains showed high minimal inhibitory concentration （MICs） to oxacillin. 20.87% strains produced β-lactamase, and 4.35% strains showed extended-spectrum b-lactamase （ESBL） phenotype. Moreover, 19 strains harboured bla genes including TEM-1 （n=5）, TEM-116 （n=10）, and ROB-1 （n=5）. Significantly, one strain possessed both TEM-1 and ROB-1, and displayed resistance to cefotaxime （MIC=8 mg L-1）. The epidemiological analysis of PFGE revealed high genetic diversity among bla-positive isolates. This work shows that TEM- and ROB-type β-lactamases are prevalent in H. parasuis isolates in China.展开更多
Haemophilus parasuis (H. parasuis) is one of the bacterial pathogens of great concern as it causes huge economic losses to the swine industry worldwide. One of the reasons why the control of H. parasuis has failed is ...Haemophilus parasuis (H. parasuis) is one of the bacterial pathogens of great concern as it causes huge economic losses to the swine industry worldwide. One of the reasons why the control of H. parasuis has failed is the increase in antimicrobial resistance (AMR). The country of Vietnam has the second-largest pig production in Asia. However, there is still a lack of data about the AMR prevalence of H. parasuis in Vietnam.The purpose of this study is to investigate the prevalence of AMR and analyze the association between AMR and AMR genes (ARGs). The H. parasuis strains used in this research were isolated from swine in the Quang Binh and Thua Thien Hue Provinces, Central Vietnam, as reported in our previous study. All of the strains were tested for AMR against 25 antibacterial agents using the broth microdilution method and for the presence of ARGs using the polymerase chain reaction (PCR) method. The tested strains were shown to have a high frequency of resistance to trimethoprim/sulfamethoxazole (94.6%), followed by resistance to colistin, chloramphenicol, gentamicin, penicillin, lincomycin, and amoxicillin. The most prevalent ARGs in these strains were blaTEM-1 (94.6%), int (76.8%), gyrA (58.9%), and rmtD (50.0%). Cefuroxime, chloramphenicol, and tobramycin resistances were strongly correlated with the presence of the ARGs blaROB-1 (odds ratio (OR) = 26.3, 95% confidence interval (CI) 2.7–255.7, p = 0.002), catl (OR = 25.1, 95% CI 2.4–258.9, p = 0.004), and strB (OR = 23.5, 95% CI 2.6–212.6, p = 0.001), respectively.This study reveals for the first time the current situation of H. parasuis AMR in Central Vietnam, which is helpful for the clinical control of this disease, as well as for the development of policies and clinical practice guidelines to reduce AMR in swine production in Central Vietnam.展开更多
[ Objective] To develop multiplex-PCR for diagnosis of mixed infection caused by Pasteurella multocida （ PM）, Haomophilus parasuis （HPS） and Actinbaci/lus pleuropneumoniae （App）. [ Method ] PCR method was develo...[ Objective] To develop multiplex-PCR for diagnosis of mixed infection caused by Pasteurella multocida （ PM）, Haomophilus parasuis （HPS） and Actinbaci/lus pleuropneumoniae （App）. [ Method ] PCR method was developed to detect single infection caused by PM, HPS or App. The conditions of amplification and primers were optimized, and the multiple-PCR was developed to detect mixed infection of PM, HPS and App. [ Result] A 457-bp band, a 821-bp band and a 342-bp band were simultaneously amplified in the one PCR reaction system. The method had high sensitivity and specificity. [ Conduslon] The multiple-PCR is successfully developed and can be used for differential diagnosis of PM, HPS and App.展开更多
Respiratory diseases are one of the most important health problems in pig herds. The porcine respiratory disease complex (PRDC) is the term used to describe pneumonic diseases caused by multiple infectious agents that...Respiratory diseases are one of the most important health problems in pig herds. The porcine respiratory disease complex (PRDC) is the term used to describe pneumonic diseases caused by multiple infectious agents that provoke weight loss in animals or death. In the PRDC multiple pathogens (bacteria and/or viruses) work in combination to induce this respiratory disease. Within this complex, Actinobacillus pleuropneumoniae, Streptococcus suis, Pasteurella multocida, Bordetella bronchiseptica, Haemophilus parasuis and Mycoplasma hyopneumoniae are the main bacterial pathogens involved in great economic losses to the swine industry. The aim of this work was to estimate the presence of A. pleuropneumoniae, S. suis, P. multocida, B. bronchiseptica, H. parasuis and M. hyopneumoniae in the upper respiratory tract of pigs in representative swine farms inAguascalientes,Mexico, using PCR technique. The study was performed in 14 swine farms. We obtained a total of 212 nasal swabs. Near 20% of samples were positive for A. pleuropneumoniae (located in the 79% of farms);17% were positive for S. suis (in 86% of farms), of these, 3% were S. suis serovar 2;30% were positive for H. parasuis (93% of farms);23% of the samples to P. multocida (in 79% of farms);and 19% to M. hyopneumoniae (in 64% of farms). B. bronchiseptica was not detected in this study. The results obtained show that bacterial pathogens of PRDC were present in the upper respiratory tract of pigs in all farms studied;therefore, these pathogens are widely disseminated in pig farms of Aguascalientes, Mexico.展开更多
To investigate the infection of Haemophilus parasuislin some regions of Sichuan Province, indirect hemagglutination from 2006 to 2008 was tested by Haemophilus parasuis antibody, and 1 062 serum samples from 10 differ...To investigate the infection of Haemophilus parasuislin some regions of Sichuan Province, indirect hemagglutination from 2006 to 2008 was tested by Haemophilus parasuis antibody, and 1 062 serum samples from 10 different regions and sources in Sichuan Province were also examined. The results showed that the positive rate ranged from 2% to 30% with average of 10%. The positive rate varied with pigs at different ages, and especially that of 30-day-old to 70-day-old weaned piglets reached 25%. Thus, there were various infections of HaemophUus parasuis in some regions of Sichuan Province, and the statistic data provided a reference for effective prevention and control of Haemophilus parasuis in Sichuan Province.展开更多
[ Objective] To develop a real-time fluorescent PCR assay for rapid detection of Haempohlius parasuis （HPS）. [ Method] According to the conservative sequences of 16 S rRNA genes of HPS published in GenBank, a pair o...[ Objective] To develop a real-time fluorescent PCR assay for rapid detection of Haempohlius parasuis （HPS）. [ Method] According to the conservative sequences of 16 S rRNA genes of HPS published in GenBank, a pair of specific primers was designed. The real-time fluorescent PCR was developed by optimizing primer concentration and annealing temperature. And its specificity and reproducibility were evaluated. Ten HPS- suspected samples were detected by the developed method. [ Result] The lowest detection limit of the developed real-time fluorescent PCR was 50 copies/μl. This method had good reproducibility, and its coefficient of variation was lower than 2%. Only HPS rather than Streptococcus suis type 2, Staphylococcus aureus, E. coli DH5 alpha, and swine Salmonella typhi could be detected by the developed real-time fluorescent PCR. The HPS-pesitive samples detected by this method were also positive when they were detected by isolation of bacteria or conventional PCR. [ Conclusion] The developed real-time fluorescent PCR is rapid, sensitive, specific and highly reproducible; thus, it can be used for rapid detection of HPS.展开更多
[Objective]The aim was to establish an indirect ELISA for detection on antibody of Hps. [Method] The optimal conditions of indirect ELISA were selected and determined based on heat-resistant serotype 4 and 5 of Hps; s...[Objective]The aim was to establish an indirect ELISA for detection on antibody of Hps. [Method] The optimal conditions of indirect ELISA were selected and determined based on heat-resistant serotype 4 and 5 of Hps; specific,repeating and sensitive tests were conducted and 200 serums were detected. [Result]The optimal conditions were as follows： coating concentration of antigen at 10 μg /ml,and coating for 2 h at 37 °C; PBST containing 20 g /L of skim milk powder as blocking fluid for 30 min; serum dilution at 1∶ 80; reaction time of antigen for 45 min; dilution of secondary antibody at 1∶ 12 000 and effecting for 30 min; color development reaction for 15 min. [Conclusion] The established indirect ELISA is good in specificity and repetitiveness with higher sensitivity than that of indirect hemagglutination test; the results of clinic samples （ negative /positive serums） were in consistent with those detected with foreign ELISA kits. The established method can be made use of in serum antibody of Hps de- tection and seroepidemiology study.展开更多
Haemophilus parasuis is one kind of constant bacteria in porcine upper respiratory tract, and it can cause multiple serositis, arthritis and other diseases under certain conditions. Due to lack of efficient genetic op...Haemophilus parasuis is one kind of constant bacteria in porcine upper respiratory tract, and it can cause multiple serositis, arthritis and other diseases under certain conditions. Due to lack of efficient genetic operating system, its pathogenic mechanism is not very clear. Ligation with DNA ligase and fusion PCR were used to construct targeting hhdA gene of Haemophilus parasuis, respectively. The fidelity, application scope, operation and conditions of the constructed fusion fragments were compared. The results showed that construction with DNA ligase was more mature technology as manifested by more stable conditions and more extensive application. The fusion PCR method had high fidelity and simple operation, and the transformation rate was 9.5 times as high as that of ligation with DNA ligase. For this reason, this method was more suitable for construction of multi-fragment targeting genes. The study lays a foundation for establishing an efficient operating system of targeting gene of Haemophilus parasuis in the future.展开更多
This paper introduced the characteristics of Glasserella parasuis from the aspects of pathogen,epidemiology,clinical symptoms and anatomical symptoms,and put forward its clinical and laboratory diagnosis methods.Moreo...This paper introduced the characteristics of Glasserella parasuis from the aspects of pathogen,epidemiology,clinical symptoms and anatomical symptoms,and put forward its clinical and laboratory diagnosis methods.Moreover,the disease was differentiated from similar diseases such as mycoplasmal pneumonia of swine,swine flu infectious pleuropneumonia of swine,swine streptococcosis and porcine reproductive and respiratory syndrome.Finally,the prevention and treatment measures of the disease are proposed.展开更多
To construct the suicide vector of hhd B gene marker-free mutant in Haemophilus parasuis( HPS),two pairs of specific primers were designed and synthesized according to the hhd B gene upstream and downstream sequences ...To construct the suicide vector of hhd B gene marker-free mutant in Haemophilus parasuis( HPS),two pairs of specific primers were designed and synthesized according to the hhd B gene upstream and downstream sequences of HPS published in Gen Bank. The hhd B gene upstream and downstream sequences were amplified by PCR,which were further ligated( hhd B-up + down) through overlapping PCR method. NotⅠand SalⅠrestriction enzyme sites were introduced on both ends of the ligated sequence. After the corresponding digestion,the hhd B-up + down sequence was directionally cloned to the suicide plasmid vector p EMOC2. Results showed that the suicide vector of hhd B gene marker-free deleted( p EMOC2Δhhd B) with stable inheritance in E. coli β2155 strain was successfully obtained,thereby laying the foundation for construction of HPS-hhd B gene marker-free mutant strain.展开更多
A bacterial strain was isolated from the sick pigs suspiciously infected by polyserositis and arthritis in a pig farm in Shandong Province, and identified through morphological observation, culture traits, bioehemical...A bacterial strain was isolated from the sick pigs suspiciously infected by polyserositis and arthritis in a pig farm in Shandong Province, and identified through morphological observation, culture traits, bioehemical characteristics and PCR amplifieation. Additionally, primers were de- signed according to the 16S rRNA sequence of Haemophihzs parasuis, and the bacterial strain was amplified by PCR. The amplified fragments of approximately 1 400 bp was sequenced, and aligned with the sequence in GenBank. The results showed that it shared the homology of 97%-99% with the 16S rRNA sequence of foreign H.parasuis, and confirmed as H.parasuis （HPS）. The strain was determined as serotype 4 through serotype identification. The strain was named SD02.展开更多
针对副猪嗜血杆菌(Haemophilus parasuis,HPS)16 S rRNA序列设计1对特异性引物,能扩增出821 bp的特异性DNA片段,并据此建立了快速准确鉴定副猪嗜血杆菌PCR方法,临床试验证明该方法具有很好的特异性和敏感性。13份疑似病料检测结果表明,...针对副猪嗜血杆菌(Haemophilus parasuis,HPS)16 S rRNA序列设计1对特异性引物,能扩增出821 bp的特异性DNA片段,并据此建立了快速准确鉴定副猪嗜血杆菌PCR方法,临床试验证明该方法具有很好的特异性和敏感性。13份疑似病料检测结果表明,PCR检测结果与传统生化鉴定结果符合率为100%。结果表明已成功建立了副猪嗜血杆菌PCR检测方法,并可应用于临床副猪嗜血杆菌的检测。展开更多
Haemophilus parasuis（HPS） is one of the most important bacterial infectious diseases harming China＇s pig industry for the last few years.To understand the prevalence status of glasser＇s disease in China, the disea...Haemophilus parasuis（HPS） is one of the most important bacterial infectious diseases harming China＇s pig industry for the last few years.To understand the prevalence status of glasser＇s disease in China, the disease samples submitted by Shandong, Henan, Heilongjiang, Jiangxi, Liaon-ing, Guangdong, Shaanxi and Sichuan provinces were isolated and identified. The synovial fluid, lung, brain tissue, heart blood and nasal cavity ofsick pigs in 194 pig farms collected from 2006 to 2009 were detected, and 47 strains of HPS were eventually isolated, with the separation rate of24.2%. Serotypes of the isolates were identified through KRG （Kieletein-Rapp-Gabriedson） agar diffusion method. There were 13 strains of serotype4 （27.7%）, 10 strains of serotype 5 （21.3%）, 8 strains of serotype 12 （17%） and7 strains of serotype 13 （14.9%）, indicating the major serotypes ofHPS in China were serotype 4, serotype 5, serotype 12 and serotype 13.展开更多
基金Supported by New Era Vocational School famous teachers and Principals Training Program of Hubei ProvinceAction Plan for Science and Technology Support in Rural Revitalization of Higher Education Institutions of Department of Education of Hubei Province(BXLBX1376)Horizontal Project of Animal Epidemic Diagnosis Technology Research and Product Development.
文摘[Objectives]This study was conducted to evaluate the practical efficacy of Haemophilus parasuis(Hps)vaccines and determine whether the continuous use of vaccines leads to changes in Hps serotypes.[Methods]A swine production group that had used a bivalent vaccine of Hps serotypes 4 and 5 for two consecutive years were investigated and retrospectively analyzed.Data on Hps detection rate,nursery production and disease indexes,as well as serotype shifts after continuous vaccination,were collected to assess vaccine effectiveness.[Results]Continuous vaccination reduced the total nursery culling and mortality rate by 1.75%.Hps infection persisted in the farms,and the pathogen could still be isolated.However,serotyping revealed a significant shift in the dominant prevalent serotypes,indicating that the vaccine was indeed effective.[Conclusions]The matched Hps vaccine demonstrated effective results.However,whether replacing serotypes or adding more prevalent serotypes could further improve the control of Hps disease requires further investigation.
基金Supported by National High Technology Research and Development Program of China(2006AA10A206)National Natural Science Foundation of China(31001072)the Fund of Beijing Academy of Agriculture and Forestry Sciences for Young Scholars(QNJJ201012)~~
文摘[Objective] The study aimed to obtain attenuated strain of Haemophilus parasuis.[Method] Tn5 transposon technology was used to construct a library of mutants.Positive mutants were screened by kanamycin resistance.False positive was identified by PCR and then removed.Mice were infected to detect the virulence of mutants.The bionomics of attenuated strains were detected,too.[Result] The attenuated mutants showed similar reproductive activity to that of wild strain.The virulence of mutants was still stable after 30 passages.[Conclusion] This study provided foundation for exploring the virulence factors and pathogenic mechanism of HPS.
基金Supported by National Natural Science Foundation of China(31001072)National High Technology Research and Development Program of China(2006AA10A206)+1 种基金Youth Foundation of Beijing Academy of Agriculture and Forestry(QNJJ201012)Program of Beijing Academy of Agriculture and Forestry(2010A008)~~
文摘[Objective] The aim was to clone and express the pilA gene of outer membrane protein of Haemophilus parasuis.[Method] The published pilA gene sequence of HPS was analyzed for primer synthesis,and the genome of serotype 5-type of HPS was used as template for PCR amplification of the pilA gene of HPS;the recombinant expression plasmid was constructed and transformed into E.coli BL21（DE3）after induced by IPTG.SDS-PAGE and Western blot analysis were then carried out.[Result] The molecular weight of expressed protein was consistent with the expected（43 kD）.[Conclusion] The results provided a foundation for the preparation of subunit vaccine and diagnostic reagents.
基金supported by the Program for New Century Excellent Talents in University,China (NCET-06-0752)the Guangdong Technology Planning Committee,China (2006B0152 and 2009A0201006)
文摘To characterize the β-lactam resistance in veterinary clinical isolates of Haemophilus parasuis, 115 isolates were examined for the β-lactam resistance, the possession of β-lactamase, and the presence of β-lactamase genes. The genetic relationship among isolates was evaluated by pulsed-field gel electrophoresis （PFGE）. Overall, the commonly detected resistance phenotypes were resistant to ampicillin （26.09%）, penicillin （22.61%）, amoxicillin （21.74%）, cefazolin （14.78%）, cefaclor （12.17%）, and cefotaxime （6.96%）. These strains showed high minimal inhibitory concentration （MICs） to oxacillin. 20.87% strains produced β-lactamase, and 4.35% strains showed extended-spectrum b-lactamase （ESBL） phenotype. Moreover, 19 strains harboured bla genes including TEM-1 （n=5）, TEM-116 （n=10）, and ROB-1 （n=5）. Significantly, one strain possessed both TEM-1 and ROB-1, and displayed resistance to cefotaxime （MIC=8 mg L-1）. The epidemiological analysis of PFGE revealed high genetic diversity among bla-positive isolates. This work shows that TEM- and ROB-type β-lactamases are prevalent in H. parasuis isolates in China.
文摘Haemophilus parasuis (H. parasuis) is one of the bacterial pathogens of great concern as it causes huge economic losses to the swine industry worldwide. One of the reasons why the control of H. parasuis has failed is the increase in antimicrobial resistance (AMR). The country of Vietnam has the second-largest pig production in Asia. However, there is still a lack of data about the AMR prevalence of H. parasuis in Vietnam.The purpose of this study is to investigate the prevalence of AMR and analyze the association between AMR and AMR genes (ARGs). The H. parasuis strains used in this research were isolated from swine in the Quang Binh and Thua Thien Hue Provinces, Central Vietnam, as reported in our previous study. All of the strains were tested for AMR against 25 antibacterial agents using the broth microdilution method and for the presence of ARGs using the polymerase chain reaction (PCR) method. The tested strains were shown to have a high frequency of resistance to trimethoprim/sulfamethoxazole (94.6%), followed by resistance to colistin, chloramphenicol, gentamicin, penicillin, lincomycin, and amoxicillin. The most prevalent ARGs in these strains were blaTEM-1 (94.6%), int (76.8%), gyrA (58.9%), and rmtD (50.0%). Cefuroxime, chloramphenicol, and tobramycin resistances were strongly correlated with the presence of the ARGs blaROB-1 (odds ratio (OR) = 26.3, 95% confidence interval (CI) 2.7–255.7, p = 0.002), catl (OR = 25.1, 95% CI 2.4–258.9, p = 0.004), and strB (OR = 23.5, 95% CI 2.6–212.6, p = 0.001), respectively.This study reveals for the first time the current situation of H. parasuis AMR in Central Vietnam, which is helpful for the clinical control of this disease, as well as for the development of policies and clinical practice guidelines to reduce AMR in swine production in Central Vietnam.
基金funded by the subproject of National Key Technology R&D Program (2006BAD06A01 and 2006BAD06A12)Basic Work of National Science and Technology Special Plan (2008FY210200)
文摘[ Objective] To develop multiplex-PCR for diagnosis of mixed infection caused by Pasteurella multocida （ PM）, Haomophilus parasuis （HPS） and Actinbaci/lus pleuropneumoniae （App）. [ Method ] PCR method was developed to detect single infection caused by PM, HPS or App. The conditions of amplification and primers were optimized, and the multiple-PCR was developed to detect mixed infection of PM, HPS and App. [ Result] A 457-bp band, a 821-bp band and a 342-bp band were simultaneously amplified in the one PCR reaction system. The method had high sensitivity and specificity. [ Conduslon] The multiple-PCR is successfully developed and can be used for differential diagnosis of PM, HPS and App.
文摘Respiratory diseases are one of the most important health problems in pig herds. The porcine respiratory disease complex (PRDC) is the term used to describe pneumonic diseases caused by multiple infectious agents that provoke weight loss in animals or death. In the PRDC multiple pathogens (bacteria and/or viruses) work in combination to induce this respiratory disease. Within this complex, Actinobacillus pleuropneumoniae, Streptococcus suis, Pasteurella multocida, Bordetella bronchiseptica, Haemophilus parasuis and Mycoplasma hyopneumoniae are the main bacterial pathogens involved in great economic losses to the swine industry. The aim of this work was to estimate the presence of A. pleuropneumoniae, S. suis, P. multocida, B. bronchiseptica, H. parasuis and M. hyopneumoniae in the upper respiratory tract of pigs in representative swine farms inAguascalientes,Mexico, using PCR technique. The study was performed in 14 swine farms. We obtained a total of 212 nasal swabs. Near 20% of samples were positive for A. pleuropneumoniae (located in the 79% of farms);17% were positive for S. suis (in 86% of farms), of these, 3% were S. suis serovar 2;30% were positive for H. parasuis (93% of farms);23% of the samples to P. multocida (in 79% of farms);and 19% to M. hyopneumoniae (in 64% of farms). B. bronchiseptica was not detected in this study. The results obtained show that bacterial pathogens of PRDC were present in the upper respiratory tract of pigs in all farms studied;therefore, these pathogens are widely disseminated in pig farms of Aguascalientes, Mexico.
基金supported by National Science and Technology Support Program " Epidemiological Studies on Newly-emerged Animal Diseases such as Highly Pathogenic Avian Influenza" (2008BAD06A01,2006BAD06A12)973 Project "Important Pathogenic Molecular Mechanism of Animal Pathogens" (2006CB504403)
文摘To investigate the infection of Haemophilus parasuislin some regions of Sichuan Province, indirect hemagglutination from 2006 to 2008 was tested by Haemophilus parasuis antibody, and 1 062 serum samples from 10 different regions and sources in Sichuan Province were also examined. The results showed that the positive rate ranged from 2% to 30% with average of 10%. The positive rate varied with pigs at different ages, and especially that of 30-day-old to 70-day-old weaned piglets reached 25%. Thus, there were various infections of HaemophUus parasuis in some regions of Sichuan Province, and the statistic data provided a reference for effective prevention and control of Haemophilus parasuis in Sichuan Province.
基金funded by the Key Technologies R&D Program of Guangxi of China (0993009-1)
文摘[ Objective] To develop a real-time fluorescent PCR assay for rapid detection of Haempohlius parasuis （HPS）. [ Method] According to the conservative sequences of 16 S rRNA genes of HPS published in GenBank, a pair of specific primers was designed. The real-time fluorescent PCR was developed by optimizing primer concentration and annealing temperature. And its specificity and reproducibility were evaluated. Ten HPS- suspected samples were detected by the developed method. [ Result] The lowest detection limit of the developed real-time fluorescent PCR was 50 copies/μl. This method had good reproducibility, and its coefficient of variation was lower than 2%. Only HPS rather than Streptococcus suis type 2, Staphylococcus aureus, E. coli DH5 alpha, and swine Salmonella typhi could be detected by the developed real-time fluorescent PCR. The HPS-pesitive samples detected by this method were also positive when they were detected by isolation of bacteria or conventional PCR. [ Conclusion] The developed real-time fluorescent PCR is rapid, sensitive, specific and highly reproducible; thus, it can be used for rapid detection of HPS.
基金funded by Scientific Research Foundation of Guangdong Dahuanong Animal Health Products CO. LTD.(033A201055-2)
文摘[Objective]The aim was to establish an indirect ELISA for detection on antibody of Hps. [Method] The optimal conditions of indirect ELISA were selected and determined based on heat-resistant serotype 4 and 5 of Hps; specific,repeating and sensitive tests were conducted and 200 serums were detected. [Result]The optimal conditions were as follows： coating concentration of antigen at 10 μg /ml,and coating for 2 h at 37 °C; PBST containing 20 g /L of skim milk powder as blocking fluid for 30 min; serum dilution at 1∶ 80; reaction time of antigen for 45 min; dilution of secondary antibody at 1∶ 12 000 and effecting for 30 min; color development reaction for 15 min. [Conclusion] The established indirect ELISA is good in specificity and repetitiveness with higher sensitivity than that of indirect hemagglutination test; the results of clinic samples （ negative /positive serums） were in consistent with those detected with foreign ELISA kits. The established method can be made use of in serum antibody of Hps de- tection and seroepidemiology study.
基金supported by the grants of the Major State Basic Research Development Program (2006CB504403)the Supporting Program of the"Eleventh Five-year Plan"for Sci & Tech Research of China (2006BAD06A01 and 2006BAD06A12 )+1 种基金the Foundation Project of State key Laboratory of Veterinary Etiological Biology (SKLVEB2008ZZKT009)the Open Foundation Project of Guangdong Common Lab of Veterinary Public Health (GSKJ090202)
文摘Haemophilus parasuis is one kind of constant bacteria in porcine upper respiratory tract, and it can cause multiple serositis, arthritis and other diseases under certain conditions. Due to lack of efficient genetic operating system, its pathogenic mechanism is not very clear. Ligation with DNA ligase and fusion PCR were used to construct targeting hhdA gene of Haemophilus parasuis, respectively. The fidelity, application scope, operation and conditions of the constructed fusion fragments were compared. The results showed that construction with DNA ligase was more mature technology as manifested by more stable conditions and more extensive application. The fusion PCR method had high fidelity and simple operation, and the transformation rate was 9.5 times as high as that of ligation with DNA ligase. For this reason, this method was more suitable for construction of multi-fragment targeting genes. The study lays a foundation for establishing an efficient operating system of targeting gene of Haemophilus parasuis in the future.
基金Supported by Major Special Project of Industry University Research Cooperation in Lu'an City(2020-7)Key Research and Development Program of Anhui Province(202204c06020009)+1 种基金The Natural Science Foundation of Anhui Province(2308085MC102)The Anhui Academy of Agricultural Sciences Platform Project(2024YL016).
文摘This paper introduced the characteristics of Glasserella parasuis from the aspects of pathogen,epidemiology,clinical symptoms and anatomical symptoms,and put forward its clinical and laboratory diagnosis methods.Moreover,the disease was differentiated from similar diseases such as mycoplasmal pneumonia of swine,swine flu infectious pleuropneumonia of swine,swine streptococcosis and porcine reproductive and respiratory syndrome.Finally,the prevention and treatment measures of the disease are proposed.
基金Supported by Science and Technology Project of Guangdong Province(2013B020307002)
文摘To construct the suicide vector of hhd B gene marker-free mutant in Haemophilus parasuis( HPS),two pairs of specific primers were designed and synthesized according to the hhd B gene upstream and downstream sequences of HPS published in Gen Bank. The hhd B gene upstream and downstream sequences were amplified by PCR,which were further ligated( hhd B-up + down) through overlapping PCR method. NotⅠand SalⅠrestriction enzyme sites were introduced on both ends of the ligated sequence. After the corresponding digestion,the hhd B-up + down sequence was directionally cloned to the suicide plasmid vector p EMOC2. Results showed that the suicide vector of hhd B gene marker-free deleted( p EMOC2Δhhd B) with stable inheritance in E. coli β2155 strain was successfully obtained,thereby laying the foundation for construction of HPS-hhd B gene marker-free mutant strain.
文摘A bacterial strain was isolated from the sick pigs suspiciously infected by polyserositis and arthritis in a pig farm in Shandong Province, and identified through morphological observation, culture traits, bioehemical characteristics and PCR amplifieation. Additionally, primers were de- signed according to the 16S rRNA sequence of Haemophihzs parasuis, and the bacterial strain was amplified by PCR. The amplified fragments of approximately 1 400 bp was sequenced, and aligned with the sequence in GenBank. The results showed that it shared the homology of 97%-99% with the 16S rRNA sequence of foreign H.parasuis, and confirmed as H.parasuis （HPS）. The strain was determined as serotype 4 through serotype identification. The strain was named SD02.
文摘针对副猪嗜血杆菌(Haemophilus parasuis,HPS)16 S rRNA序列设计1对特异性引物,能扩增出821 bp的特异性DNA片段,并据此建立了快速准确鉴定副猪嗜血杆菌PCR方法,临床试验证明该方法具有很好的特异性和敏感性。13份疑似病料检测结果表明,PCR检测结果与传统生化鉴定结果符合率为100%。结果表明已成功建立了副猪嗜血杆菌PCR检测方法,并可应用于临床副猪嗜血杆菌的检测。
文摘Haemophilus parasuis（HPS） is one of the most important bacterial infectious diseases harming China＇s pig industry for the last few years.To understand the prevalence status of glasser＇s disease in China, the disease samples submitted by Shandong, Henan, Heilongjiang, Jiangxi, Liaon-ing, Guangdong, Shaanxi and Sichuan provinces were isolated and identified. The synovial fluid, lung, brain tissue, heart blood and nasal cavity ofsick pigs in 194 pig farms collected from 2006 to 2009 were detected, and 47 strains of HPS were eventually isolated, with the separation rate of24.2%. Serotypes of the isolates were identified through KRG （Kieletein-Rapp-Gabriedson） agar diffusion method. There were 13 strains of serotype4 （27.7%）, 10 strains of serotype 5 （21.3%）, 8 strains of serotype 12 （17%） and7 strains of serotype 13 （14.9%）, indicating the major serotypes ofHPS in China were serotype 4, serotype 5, serotype 12 and serotype 13.
