The aim of this study was to evaluate the factors influencing the inactivation effect of intense pulsed light(IPL)on Aeromonas salmonicida grown on chicken meat and skin,and to further develop prediction models of ina...The aim of this study was to evaluate the factors influencing the inactivation effect of intense pulsed light(IPL)on Aeromonas salmonicida grown on chicken meat and skin,and to further develop prediction models of inactivation.In this work,chicken meat and skin inoculated with meat-borne A.salmonicida isolates were subjected to IPL treatments under different conditions.The results showed that IPL had obvious bactericidal effect in the chicken skin and thickness groups when the treatment voltage and time were 7 V combined with 5 s.In addition,the lethality curves of A.salmonicida were fitted under IPL conditions of 3.5-7.5 V.The comparison of statistical parameters revealed that the Weibull model could best fit the mortality curves and could accurately predict the mortality dynamic of A.salmonicida grown on chicken skin.And further a secondary model between the scale factor b and the treatment voltage in Weibull model was established using linear equations,which determined that the secondary model could accurately predict the inactivation of A.salmonicida.This study provides a theoretical basis for future prediction models of Aeromonas,and also provides new ideas for sterilization approaches of meat-borne Aeromonas.展开更多
Vibrio scophthalmi and Aeromonas salmonicida can cause high turbot mortality and huge economic losses.Presently,vaccination is the most promising method for preventing communicable diseases.In this study,we used forma...Vibrio scophthalmi and Aeromonas salmonicida can cause high turbot mortality and huge economic losses.Presently,vaccination is the most promising method for preventing communicable diseases.In this study,we used formalin to kill V.scophthalmi and A.salmonicida cells,and mixed with the mineralized oil adjuvant(Montanide^(TM)ISA 763 AVG)to prepare the bivalent inactivated vaccine.The results showed that turbot inoculated with the bivalent inactivated vaccine exhibited strong tolerance to the infection of V.scophthalmi and A.salmonicida,and no obvious clinical symptoms and pathological changes were observed.The activities of enzymes lysozyme,acid phosphatase and complement C3 had significantly increased after the vaccination.The antibody titer response of vaccinated turbot was greatly boosted,which was positively connected with the immunological impact according to ELISA results.Simultaneously,the expression levels of immune-related genes such as MHC-IIα,MHC-IIβ,CD4,CD8,TNF-αand IL^(-1)βwere up-regulated,demonstrating that it might stimulate humoral and cellular immunological response in turbot.These findings highlight the potential of the bivalent inactivated vaccine for controlling V.scophthalmi and A.salmonicida infections in turbot.展开更多
Aim of this research was to find out the concentration of enrofloxacin residue in tilapia meat for several weeks after antibiotic treatment. Twenty seven tilapia fishes were divided into three groups. The first group ...Aim of this research was to find out the concentration of enrofloxacin residue in tilapia meat for several weeks after antibiotic treatment. Twenty seven tilapia fishes were divided into three groups. The first group was not infected and treated, the second group was infected with A. salmonicida subsp, smithia and the third group was infected with A. salmonicida subsp. achromogenes intramuscularly. Six days after infection, treatment was carried out using Baytril administered orally for the second group and intramuscularly for the third group during five days. At the 1 st, 4th and 8th week after the treatment, Three fish were taken from each group to be analyzed for its concentration of enrofloxacin residue by diffusion on Mueller Hinton Agar （MHA） method and quantitatively using high performance liquid chromatography （HPLC） method. The MHA test showed the formation of inhibition zone, at the 1 st week and 4th week after the treatment, while at 8th week after treatment did not show inhibition zone. The HPLC test on enrofloxacin residual concentration in tilapia infected with A. salmonicida subsp, smithia （second group） at the 1st, 4th and 8th week after treatment showed the average of 33.0, 6.10 and 0.0021 μg/g of enrofloxacin residue level. While in tilapia infected with A. salmonicida subsp, achromogenes and treated with enrofloxacin intramuscularly （third group） showed the average of residue level 35.79, 2.18 and 0.00065 μg/g. In conclusion, the residue of enrofloxacin was still high concentration until the fourth week after treatment in the second and third groups. Based on Indonesian National Standards and Rules, the maximum limit of enrofloxacin residue is 0.01 μg/g. The concentration of enrofloxacine residue was very low and the concentration of enrofloxacin residue collected from tilapia using orally and intramuscularly method of treatment was not different.展开更多
Aeromonas salmonicida is a common pathogen of salmonid fishes that poses a significant threat to the fresh water and marine culture industry,potentially resulting in huge economic losses.To prevent and control fish di...Aeromonas salmonicida is a common pathogen of salmonid fishes that poses a significant threat to the fresh water and marine culture industry,potentially resulting in huge economic losses.To prevent and control fish diseases caused by A.salmonicida,rapid and effective diagnostic approaches must be developed,and which are important for routine monitoring and clinical care.By combining recombinase polymerase amplification(RPA)technology with a visible lateral flow strip(RPA-LF),we have enhanced both the precision of RPA detection and the convenience of real-time monitoring.In this study,we introduce a robust method for detecting A.salmonicida using RPA-LF.This assay specifically targets the ASA_1441 gene of A.salmonicida,ensuring high specificity,without cross-reactivity with other prevalent fresh water or marine pathogens.The optimal amplification temperature of the RPA assay was 39℃.Its sensitivity extends to as low as 100 fg of purified DNA,representing more than 1000-fold higher sensitivity than conventional PCR methods.Furthermore,to enhance the usability of the RPA-LF assay,we developed a rapid sample preparation method using cellulose dipsticks for nucleic acid extraction.This method achieves a limit of detection(LOD)as low as 1.67 CFU/μL and completes the entire process within 20 min.In conclusion,our findings present a rapid and precise tool for monitoring A.salmonicida infection in aquaculture and marine culture.This advancement offers valuable insights for effective disease prevention and control strategies.展开更多
In this study,bacterial pathogens were isolated from the liver,spleen,kidney,heart,and muscle of diseased goldfish(Carassius auratus)after a massive outbreak of disease in an aquaculture farm in Anshan,Liaoning Provin...In this study,bacterial pathogens were isolated from the liver,spleen,kidney,heart,and muscle of diseased goldfish(Carassius auratus)after a massive outbreak of disease in an aquaculture farm in Anshan,Liaoning Province,North China(N41°05′58.42″E122°53′31.89″).Based on physiological and biochemical characterization of the bacteria and 16S rDNA sequence analysis,the bacterial isolates were identified as Aeromonas salmonicida salmonicida.Virulence gene multilocus sequence typing analysis showed that the bacterial isolates from all the organs collected were of the same type,and clustered in the same phylogenetic group as A.salmonicida salmonicida.The pathogen,was named AS.17 and was resistant to rifampicin,bacitracin,vancomycin,penicillin and compound sulfamethoxazole,but was highly sensitive to nalidixic acid,lomefloxacin,spectinomycin,ofloxacin,and ciprofloxacin.Goldfish experimentally infected with AS.17 had similar symptoms to naturally infected fish,including gill filament anemia,intestinal bleeding,intra-abdominal fluid,and jaw bleeding.Histological analyses showed that the liver,the spleen,the kidney and the intestines of fish infected with AS.17 had severe pathological alterations.In summary,this study reports the isolation of A.salmonicida salmonicida from C.auratus in North China and provides a reference strain and a research foundation for further studies on A.salmonicida disease control and epidemiology.展开更多
利用 A 值法与活菌平板计数法,测定了杀鲑气单胞菌在不同条件下的生长曲线,研究了不同温度及pH对杀鲑气单胞菌（Aeromonas salmonicha）生长的影响。结果表明,活菌计数法更能真实的反映细菌数量的动态变化。在28℃,15℃,10℃,5℃条件下...利用 A 值法与活菌平板计数法,测定了杀鲑气单胞菌在不同条件下的生长曲线,研究了不同温度及pH对杀鲑气单胞菌（Aeromonas salmonicha）生长的影响。结果表明,活菌计数法更能真实的反映细菌数量的动态变化。在28℃,15℃,10℃,5℃条件下,随着温度降低,杀鲑气单胞菌生长速率明显下降；在pH6.0-8.0时,酸性条件能明显抑制杀鲑气单胞菌的生长；在28℃、pH7.5条件下,杀鲑气单胞菌生长速率最快,达到稳定期时细菌数量也最多。在养殖生产过程中,为了抑制杀鲑气单胞菌的生长繁殖,建议在不影响鱼类正常生长的条件下,尽量降低养殖水体温度并合理控制水体pH,以降低养殖鱼类发病率。展开更多
为实现杀鲑气单胞菌早期快速准确定量检测,研究旨在建立杀鲑气单胞菌的SYBR Green Ⅰ实时荧光定量PCR(Real-time PCR)检测方法。根据GenBank中杀鲑气单胞菌毒力阵列蛋白基因(vapA)保守序列设计并合成一对特异性引物,对其特异性、灵敏度...为实现杀鲑气单胞菌早期快速准确定量检测,研究旨在建立杀鲑气单胞菌的SYBR Green Ⅰ实时荧光定量PCR(Real-time PCR)检测方法。根据GenBank中杀鲑气单胞菌毒力阵列蛋白基因(vapA)保守序列设计并合成一对特异性引物,对其特异性、灵敏度、可重复性和应用性进行评价。结果显示,研究设计的引物具有良好的种间特异性,仅对杀鲑气单胞菌及其亚种有阳性扩增,与其他细菌不发生交叉反应。构建的Real-time PCR标准曲线质粒拷贝数与循环阈值呈良好的线性关系,扩增所得标准曲线分别为y=–4.8345x+42.535,相关系数R^2为0.998,最低检测限为34拷贝/μL,较常规PCR的灵敏度高出约1000倍。应用建立的方法检测人工感染的虹鳟病样,15个被检样品呈阳性反应,与细菌常规鉴定方法结果一致。研究表明,所建立的基于实时荧光定量PCR技术的杀鲑气单胞菌检测方法快速、特异、灵敏,可用于临床诊断和疫病监测。展开更多
基金supported by projects funded by grants from the Natural Science Foundation of Jiangsu Province in China(BK20221515)the National Natural Science Foundation of China(32172266)the Changzhou Science and Technology Support Program(CE20222002)。
文摘The aim of this study was to evaluate the factors influencing the inactivation effect of intense pulsed light(IPL)on Aeromonas salmonicida grown on chicken meat and skin,and to further develop prediction models of inactivation.In this work,chicken meat and skin inoculated with meat-borne A.salmonicida isolates were subjected to IPL treatments under different conditions.The results showed that IPL had obvious bactericidal effect in the chicken skin and thickness groups when the treatment voltage and time were 7 V combined with 5 s.In addition,the lethality curves of A.salmonicida were fitted under IPL conditions of 3.5-7.5 V.The comparison of statistical parameters revealed that the Weibull model could best fit the mortality curves and could accurately predict the mortality dynamic of A.salmonicida grown on chicken skin.And further a secondary model between the scale factor b and the treatment voltage in Weibull model was established using linear equations,which determined that the secondary model could accurately predict the inactivation of A.salmonicida.This study provides a theoretical basis for future prediction models of Aeromonas,and also provides new ideas for sterilization approaches of meat-borne Aeromonas.
基金supported by the Fish Innovation Team of Shandong Agriculture Research System (No. SDAIT-1206)the Aquatic Animal Immunologic Agents Engineering Research Center of Shandong Province, the Qingdao Agricultural University Doctoral Start-Up Fund (6631122030)+5 种基金the National Natural Science Foundation of China (No. 32002421)the Advanced Talents Foundation of QAU (No. 6651118016)the Natural Science Foundation of Shandong Province (No. ZR2019BC009)the ‘First-Class Fishery Discipline’ program of Shandong Province, the special top talent plan ‘One Thing One Decision (Yi Shi Yi Yi)’the Key Research and Development Program in Shandong Province (No. 2018YFJH0703)Breeding Plan of Shandong Provincial Qingchuang Research Team (2019)
文摘Vibrio scophthalmi and Aeromonas salmonicida can cause high turbot mortality and huge economic losses.Presently,vaccination is the most promising method for preventing communicable diseases.In this study,we used formalin to kill V.scophthalmi and A.salmonicida cells,and mixed with the mineralized oil adjuvant(Montanide^(TM)ISA 763 AVG)to prepare the bivalent inactivated vaccine.The results showed that turbot inoculated with the bivalent inactivated vaccine exhibited strong tolerance to the infection of V.scophthalmi and A.salmonicida,and no obvious clinical symptoms and pathological changes were observed.The activities of enzymes lysozyme,acid phosphatase and complement C3 had significantly increased after the vaccination.The antibody titer response of vaccinated turbot was greatly boosted,which was positively connected with the immunological impact according to ELISA results.Simultaneously,the expression levels of immune-related genes such as MHC-IIα,MHC-IIβ,CD4,CD8,TNF-αand IL^(-1)βwere up-regulated,demonstrating that it might stimulate humoral and cellular immunological response in turbot.These findings highlight the potential of the bivalent inactivated vaccine for controlling V.scophthalmi and A.salmonicida infections in turbot.
文摘Aim of this research was to find out the concentration of enrofloxacin residue in tilapia meat for several weeks after antibiotic treatment. Twenty seven tilapia fishes were divided into three groups. The first group was not infected and treated, the second group was infected with A. salmonicida subsp, smithia and the third group was infected with A. salmonicida subsp. achromogenes intramuscularly. Six days after infection, treatment was carried out using Baytril administered orally for the second group and intramuscularly for the third group during five days. At the 1 st, 4th and 8th week after the treatment, Three fish were taken from each group to be analyzed for its concentration of enrofloxacin residue by diffusion on Mueller Hinton Agar （MHA） method and quantitatively using high performance liquid chromatography （HPLC） method. The MHA test showed the formation of inhibition zone, at the 1 st week and 4th week after the treatment, while at 8th week after treatment did not show inhibition zone. The HPLC test on enrofloxacin residual concentration in tilapia infected with A. salmonicida subsp, smithia （second group） at the 1st, 4th and 8th week after treatment showed the average of 33.0, 6.10 and 0.0021 μg/g of enrofloxacin residue level. While in tilapia infected with A. salmonicida subsp, achromogenes and treated with enrofloxacin intramuscularly （third group） showed the average of residue level 35.79, 2.18 and 0.00065 μg/g. In conclusion, the residue of enrofloxacin was still high concentration until the fourth week after treatment in the second and third groups. Based on Indonesian National Standards and Rules, the maximum limit of enrofloxacin residue is 0.01 μg/g. The concentration of enrofloxacine residue was very low and the concentration of enrofloxacin residue collected from tilapia using orally and intramuscularly method of treatment was not different.
基金financially supported by National Key Research and Development Program of China(2022YFE0101200)National Natural Science Foundation of China(32102850)Shanghai Agricultural Science and Technology Innovation Project(T2023328).
文摘Aeromonas salmonicida is a common pathogen of salmonid fishes that poses a significant threat to the fresh water and marine culture industry,potentially resulting in huge economic losses.To prevent and control fish diseases caused by A.salmonicida,rapid and effective diagnostic approaches must be developed,and which are important for routine monitoring and clinical care.By combining recombinase polymerase amplification(RPA)technology with a visible lateral flow strip(RPA-LF),we have enhanced both the precision of RPA detection and the convenience of real-time monitoring.In this study,we introduce a robust method for detecting A.salmonicida using RPA-LF.This assay specifically targets the ASA_1441 gene of A.salmonicida,ensuring high specificity,without cross-reactivity with other prevalent fresh water or marine pathogens.The optimal amplification temperature of the RPA assay was 39℃.Its sensitivity extends to as low as 100 fg of purified DNA,representing more than 1000-fold higher sensitivity than conventional PCR methods.Furthermore,to enhance the usability of the RPA-LF assay,we developed a rapid sample preparation method using cellulose dipsticks for nucleic acid extraction.This method achieves a limit of detection(LOD)as low as 1.67 CFU/μL and completes the entire process within 20 min.In conclusion,our findings present a rapid and precise tool for monitoring A.salmonicida infection in aquaculture and marine culture.This advancement offers valuable insights for effective disease prevention and control strategies.
基金funded in part by National Natural Science Foundation of China(41706177)Key Laboratory of Mariculture and Stock Enhancement in North China's Sea Open Project(2018-KF-17)Doctoral Research Project of Dalian Ocean University(HDYJ201617)to Dr.R.J.Li.
文摘In this study,bacterial pathogens were isolated from the liver,spleen,kidney,heart,and muscle of diseased goldfish(Carassius auratus)after a massive outbreak of disease in an aquaculture farm in Anshan,Liaoning Province,North China(N41°05′58.42″E122°53′31.89″).Based on physiological and biochemical characterization of the bacteria and 16S rDNA sequence analysis,the bacterial isolates were identified as Aeromonas salmonicida salmonicida.Virulence gene multilocus sequence typing analysis showed that the bacterial isolates from all the organs collected were of the same type,and clustered in the same phylogenetic group as A.salmonicida salmonicida.The pathogen,was named AS.17 and was resistant to rifampicin,bacitracin,vancomycin,penicillin and compound sulfamethoxazole,but was highly sensitive to nalidixic acid,lomefloxacin,spectinomycin,ofloxacin,and ciprofloxacin.Goldfish experimentally infected with AS.17 had similar symptoms to naturally infected fish,including gill filament anemia,intestinal bleeding,intra-abdominal fluid,and jaw bleeding.Histological analyses showed that the liver,the spleen,the kidney and the intestines of fish infected with AS.17 had severe pathological alterations.In summary,this study reports the isolation of A.salmonicida salmonicida from C.auratus in North China and provides a reference strain and a research foundation for further studies on A.salmonicida disease control and epidemiology.
文摘利用 A 值法与活菌平板计数法,测定了杀鲑气单胞菌在不同条件下的生长曲线,研究了不同温度及pH对杀鲑气单胞菌（Aeromonas salmonicha）生长的影响。结果表明,活菌计数法更能真实的反映细菌数量的动态变化。在28℃,15℃,10℃,5℃条件下,随着温度降低,杀鲑气单胞菌生长速率明显下降；在pH6.0-8.0时,酸性条件能明显抑制杀鲑气单胞菌的生长；在28℃、pH7.5条件下,杀鲑气单胞菌生长速率最快,达到稳定期时细菌数量也最多。在养殖生产过程中,为了抑制杀鲑气单胞菌的生长繁殖,建议在不影响鱼类正常生长的条件下,尽量降低养殖水体温度并合理控制水体pH,以降低养殖鱼类发病率。
文摘为实现杀鲑气单胞菌早期快速准确定量检测,研究旨在建立杀鲑气单胞菌的SYBR Green Ⅰ实时荧光定量PCR(Real-time PCR)检测方法。根据GenBank中杀鲑气单胞菌毒力阵列蛋白基因(vapA)保守序列设计并合成一对特异性引物,对其特异性、灵敏度、可重复性和应用性进行评价。结果显示,研究设计的引物具有良好的种间特异性,仅对杀鲑气单胞菌及其亚种有阳性扩增,与其他细菌不发生交叉反应。构建的Real-time PCR标准曲线质粒拷贝数与循环阈值呈良好的线性关系,扩增所得标准曲线分别为y=–4.8345x+42.535,相关系数R^2为0.998,最低检测限为34拷贝/μL,较常规PCR的灵敏度高出约1000倍。应用建立的方法检测人工感染的虹鳟病样,15个被检样品呈阳性反应,与细菌常规鉴定方法结果一致。研究表明,所建立的基于实时荧光定量PCR技术的杀鲑气单胞菌检测方法快速、特异、灵敏,可用于临床诊断和疫病监测。
