[Objectives]To explore the function of hcp gene in Aeromonas hydrophila.[Methods]A pair of specific primers was designed referring to the hcp gene sequence of A.hydrophila.The hcp gene was amplified by PCR,and perform...[Objectives]To explore the function of hcp gene in Aeromonas hydrophila.[Methods]A pair of specific primers was designed referring to the hcp gene sequence of A.hydrophila.The hcp gene was amplified by PCR,and performed bioinformatics analysis.[Results]The hcp gene had a total length of 1650 bp and encoded 549 amino acids.The theoretical molecular weight of the protein predicted was about 59476.44 kDa.After predicting the N-terminal signal peptide structure of the amino acid sequence,neither obvious signal peptide cleavage site nor signal peptide was found,and the protein had no transmembrane region.The amino acid sequence had a N-glycosylation site,4 protein kinase C phosphorylation sites,7 casein kinase II phosphorylation sites,9 N-myristoylation sites,4 isoprene binding sites,10 microbody C-terminal target signal sites,and an ATP/GTP binding site motif A(P-ring).The amino acid sequence of hcp gene of A.hydrophila was performed homology analysis with other Aeromonas strains,and it showed higher homology with A.veronii.In the secondary structure,theα-helix,β-sheet,random coil and extended strand accounted for 45.36%,6.01%,37.52%and 11.11%,respectively.The tertiary structure model consisted of 18α-helix and 22β-sheet.Analysis of protein-protein network interaction demonstrated that the proteins interacting with Hcp protein were AHA_3407,nrfA,nirB-1,nirB-2 and AHA_1112.[Conclusions]Through the bioinformatics prediction results,the basic information of hcp gene of A.hydrophila is preliminarily understood,and the possible function of this protein is predicted,in order to provide guidance for subsequent vaccine research.展开更多
Aeromonas hydrophila is a Gram-negative pathogen that can infect various fish,including largemouth bass(Micropterus salmoides),which have caused huge economic losses.In present study,largemouth bass anti-A.hydrophila ...Aeromonas hydrophila is a Gram-negative pathogen that can infect various fish,including largemouth bass(Micropterus salmoides),which have caused huge economic losses.In present study,largemouth bass anti-A.hydrophila antibodies were produced,then a highly immunogenic outer membrane proteins,AHA_3793,was identified by combined western blotting and mass spectrometry analysis.Moreover,AHA_3793 was expressed,and its immunogenicity was further verified by western blotting.Subsequently,the protective efficacy of AHA_3793 were evaluated in largemouth bass.The results showed that rAHA_3793 could produce a relative percentage survival(RPS)of 61.76% for largemouth bass against A.hydrophila challenge.ELISA analysis showed the specific serum antibodies of largemouth bass against rAHA_3793 and A.hydrophila in vaccinated group in weeks 4 and 5 after immunization were significantly higher than those in control group,which suggested that rAHA_3793 induced production of specific serum antibodies against rAHA_3793 and A.hydrophila.The qRT-PCR analysis showed that expressions of CD4-2 and MHC Ⅱα were also significantly up-regulated after immunization.These results collectively demonstrated that rAHA_3793 could induce a strong humoral immune response of largemouth bass,and then produce high immune protection ef fects against A.hydrophila infection.展开更多
Rationale:Aeromonas hydrophila,a bacterium commonly found in a variety of soils,aquatic habitats,and other natural environments,is known for causing infections and hemorrhagic diseases in both aquatic and terrestrial ...Rationale:Aeromonas hydrophila,a bacterium commonly found in a variety of soils,aquatic habitats,and other natural environments,is known for causing infections and hemorrhagic diseases in both aquatic and terrestrial animals.Patients with leukemia,when infected by Aeromonas hydrophila following chemotherapy,face an increased risk of developing necrotizing fasciitis.Patient concerns:A 48-year-old male with a history of acute myeloid leukemia presented with swelling and pain in his right hand after being stabbed by a grass carp two days ago.The injury occurred on the fifth day after his last chemotherapy session when he was accidentally punctured in the right thumb and ring finger by a carp while fishing.Two days following the injury,the skin isurrounding the wound turned black immediately,while the rest of his hand was reddened,swollen,and bleeding.Diagnosis:Aeromonas hydrophila infection in the soft tissue of the right hand.Interventions:Intravenous antibiotics and local debridement.Outcomes:The patient experienced an alleviation of systemic poisoning symptoms,effective control of the local wound infection.Lessons:It is important for physicians to appreciate the potential for highly unusual and life-threatening infections in patients with acute myeloid leukemia.Early diagnosis and prompt treatment can prevent the development of necrotizing fasciitis and save patients’lives.展开更多
嗜水气单胞菌是水产养殖中一种比较常见的致病病原菌。本研究以嗜水气单胞菌为研究对象,利用SELEX指数富集系统进化技术(Systematic Evolution of Ligands by Exponential Enrichment,SELEX),经过12轮的筛选,通过生物素-抗地高辛碱性磷...嗜水气单胞菌是水产养殖中一种比较常见的致病病原菌。本研究以嗜水气单胞菌为研究对象,利用SELEX指数富集系统进化技术(Systematic Evolution of Ligands by Exponential Enrichment,SELEX),经过12轮的筛选,通过生物素-抗地高辛碱性磷酸酶显色系统检测DNA适配子与嗜水气单胞菌的亲和力,克隆、测序后,用DNAMAN软件分析其一级序列与二级结构。研究结果表明,通过12轮筛选后,亲和力由最初的0.32升到第9轮的0.72,亲和力提高了2.25倍。一级结构主要由GT组成,说明同源性适体有了一定程度的富集。二级结构模拟图以茎环状结构为主,说明这些茎环状结构可能是适体与靶分子进行结合作用的基础,也是其进一步形成三维空间结构的基础。展开更多
[Objective] This study was conducted to develop compound probiotics that can be used as the alternatives of chemical drugs and antibiotics in aquaculture. [Method] Different concentrations of EM (effective microorgan...[Objective] This study was conducted to develop compound probiotics that can be used as the alternatives of chemical drugs and antibiotics in aquaculture. [Method] Different concentrations of EM (effective microorganisms) and Bacillus amyloliquefaciens were mixed at a ratio of 1:1, and sprayed on fish feed. The growth of intestinal villi of the fishes that had been fed with the feed mixed with compound probiotics for three months was observed; meanwhile, the content of in- testinal Aeromonas hydrophila was detected by real-time quantitative PCR. |Result] The compound probiotics promoted the development of intestinal villi, and inhibited the growth of A. hydrophila, and the effects were also concentration dependent. However, the compound probiotics did not increase the thickness of the intestinal serous layer, muscular layer and submucosal layer. [Conclusion] The compound probiotics we prepared can be used and popularized in aquiculture as it can inhibit the growth of A. hydrophila.展开更多
A strain WL-11 with high laccase activity was isolated from activated sludge collected from the effluent treatment plant of a textile and dyeing industry. It was identified as Aeromonas hydrophila by physiological tes...A strain WL-11 with high laccase activity was isolated from activated sludge collected from the effluent treatment plant of a textile and dyeing industry. It was identified as Aeromonas hydrophila by physiological test and 16S rDNA sequence analysis. A gene encoding of laccase from a newly isolated Aeromonas hydrophila WL-11 was cloned and characterized. Nucleotide sequence analysis showed an open reading frame of 1605 bp encoding a polypeptide comprised of 534 amino acids. The primary structure of the enzyme predicted the structural features characteristic of other laccases, including the conserved regions of four histidine-rich copper-binding sites. The predicted amino acid sequence showed a high homology (more than 60%) with bacterial laccases in the genome and protein databases and the highest degree of similarity (61% identity) was observed with the multicopper oxidase of KlebsieUa sp. 601. When expressed in Escherichia coli, the recombinant enzyme was overproduced in the cytoplasm as soluble and active form. The purified enzyme had an optimum pH of 2.6 and 8.0 for ABTS (2,2'-azino-bis(3-ethylbenzthiazolinesulfonic acid) and DMP (2,6-dimethoxyphenol), respectively. The kinetic study on ABTS revealed a higher affinity of this enzyme to this substrate than DMP.展开更多
Specific immunoglobulin (IgY) from egg yolk against Aeromonas hydrophila was produced by immunization of White Leghorn hens with formalin-killed whole cells of A. hydrophila. ELISA test using A. hydrophila as the coat...Specific immunoglobulin (IgY) from egg yolk against Aeromonas hydrophila was produced by immunization of White Leghorn hens with formalin-killed whole cells of A. hydrophila. ELISA test using A. hydrophila as the coating antigen revealed that the specific antibody titer started to increase in the egg yolk at the 13th day post-immunization (P/N=2.18), reached the peak at the 56th day (P/N=13.82), and remained at high level until day 133 (P/N=7.03). The antibody was purified by saturated ammonium sulphate with a recovery rate of 63.5%. The specific IgY inhibited the growth of A. hydrophila at a concentration of 1.0 mg/ml during the 18 h incubation. Pre-treatment of polyploid gibel carps Carassius auratus Gibelio with specific IgY had a protection rate of 60% (6/10) against challenge with A. hydrophila, while none of the fishes in the control groups receiving sterile phosphate buffered saline (PBS) or non-specific IgY survived the challenge. Treatment of fishes with the specific IgY 4 h after the challenge also had lower mortality (70%, 7/10), a 30% reduction against the control PBS or non-specific IgY groups (10/10). These results indicate that specific IgY antibodies could be obtained easily from hens immunized with an inactivated A. hydrophila and could provide a novel alternative approach to control of diseases in fishes caused by this organism.展开更多
Aeromonas hydrophila isolates from clinical cases (n=43) were tested against 8 antimicrobial agents and typed by outer membrane protein (OMP) pattern by using sodium dodecyl sulfate gel electrophoresis. All isolat...Aeromonas hydrophila isolates from clinical cases (n=43) were tested against 8 antimicrobial agents and typed by outer membrane protein (OMP) pattern by using sodium dodecyl sulfate gel electrophoresis. All isolates were resistant to ampicillin (MICs, ≥16 μg mL-1) and sulfamonomethoxine (MICs≥64 μg mLl), but susceptible to norfloxacin (MICs,≤0.5 μg mL-1). There was a high incidence of resistance to erythromycin (90.70%) and tylosin (93.02%), while a low incidences of resistance to ciprofloxacin (2.33%), enrofloxacin (2.33%) and florfenicol (4.65%). Six different outer membrane protein patterns were found among 34 isolates by analyzing proteins in the range of 22 to 50 kDa, other than 9 isolates with their respective profiles. The strains with the similar OMP profiles had similar resistances. Compared with the other strains from the same OMP patterns, NB-1, A.Pun and MR-1 had lacked the proteins in the range of 30 to 45 kDa and their resistance to florfenicol substantially increased. It is speculated that the outer membrane protein changes might correlate with decreased susceptibility to florfenicol in the three strains. Some strains which showed completely identical OMP types had a little difference in their resistance to fluoroquinolones, indicating that there might be other factors that were involved in the antimicrobial resistance of A. hydrophila.展开更多
The Aeromonas species, belonging to the family Aeromonadaceae, are opportunistic pathogens found in humans with an incidence rate of 76 cases per million inhabitants in Southern Taiwan.The incidence of Aeromonas septi...The Aeromonas species, belonging to the family Aeromonadaceae, are opportunistic pathogens found in humans with an incidence rate of 76 cases per million inhabitants in Southern Taiwan.The incidence of Aeromonas septicemia is relatively low, accounting for less than 15% of cases.Patients diagnosed with Aeromonas hydrophila bacteremia who were presented with skin blisters and septic shock have been reported to have a mortality rate of 100%.Aeromonas infection must be considered in the differential diagnosis of gangrene-like tissue damage or skin lesions in patients with end-stage renal disease, due to the potential sources of infections.A 49-year-old Taiwan Residents diabetic woman with end-stage renal disease had underwent regular hemodialysis.She was referred to our hospital due to a one-day course of fever, dyspnea, hypotension, and fulminant hemorrhagic blisters covering her whole body.A physical examination uncovered multiple hemorrhagic blisters, along with a ruptured blister over the lower left leg.Laboratory tests revealed an elevation of liver enzymes, impaired renal function, lactatemia, and high anion-gap metabolic acidosis.Cultures of both blood and hemorrhagic blister fluid grew Aeromonas hydrophila.However, she experienced persistent shock despite aggressive intravenous fluid, empiric antibiotics, and inotropic agents with norepinephrine and dopamine.Early diagnosis and prompt management using intravenous fluids, antibiotics and surgical debridement is recommended in order to improve a patient's survival rate.展开更多
The ubiquitous Aeromonas hydrophila is responsible for several pathological conditions in fish and human. Like most gram negative bacteria, its virulence relies on outer membrane lipopolysachharide (LPS). The Leloir p...The ubiquitous Aeromonas hydrophila is responsible for several pathological conditions in fish and human. Like most gram negative bacteria, its virulence relies on outer membrane lipopolysachharide (LPS). The Leloir pathway enzyme UDP-galactose 4-epimerase (GalE), plays an important role in the LPS biosynthesis, and therefore is a potential drug target. We have earlier carried out extensive biochemical and biophysical studies with histidine-tagged recombinant GalE. However, for effective drug design it is desirable to understand the structure-function relation of a protein in its native form without any additional sequences or tags. In the present study, we report the high level expression, purification and characterization of recombinant GalE (rGalE) of Aeromonas hydrophila in its native form in E coli. The rGalE expressed as a soluble protein was purified to near homogeneity. From 1 L of shake flask culture ~15 mg of purified rGalE was obtained. The purified protein was biologically active with Km and Kcat values of 0.7 mM and 28.8 s–1, respectively. The enzyme exhibited a temperature and pH optima of 37。C and 7 - 9, respectively. Thus, the present study employed for soluble expression and purification of functionally active rGalE without any tag bypasses the need for cumbersome strategies associated with removal of tag from purified protein.展开更多
The Chinese giant salamander(Andrias davidianus) is the largest living amphibian in the world. Aeromonas hydrophila strain L602 was isolated from A. davidianus. The 16S rDNA gene of this isolate was amplified using PC...The Chinese giant salamander(Andrias davidianus) is the largest living amphibian in the world. Aeromonas hydrophila strain L602 was isolated from A. davidianus. The 16S rDNA gene of this isolate was amplified using PCR,and the phylogenetic tree was constructed by the neighbor-joining method. Four virulence genes(aerA,aha1,hly and alt) of A. hydrophila were amplified by PCR and drug resistances were tested using Kirby-Bauer disk diffusion method. The results showed that the length of this 16S rDNA sequence was 1453 bp,which showed 99% homology with A. hydrophila. The GenBank accession number was JX155398. Phylogenetic analysis indicated it grouped together with A. hydrophila. Four virulence genes were all detected,indicating that strain L602 was highly virulent. This stain was resistant to four antibiotics(vibramycin,furazolidone,ampicillin and erythromycin),while it was insensitive to streptomycin. Furthermore,this strain was susceptible to six antibiotics(sulfafurazole,ciprofloxacin,penbritin,norfloxacin,florfenicol and enrofloxacin). This study will help to validate the classification and virulence of pathogenic bacteria in amphibians.展开更多
A total of 59 isolates of Aeromonas hydrophila were collected from common carp suffering from freshwater fish hemorrhage disease in 13 fishing grounds in the northeast China, and their phenotypic and genetic character...A total of 59 isolates of Aeromonas hydrophila were collected from common carp suffering from freshwater fish hemorrhage disease in 13 fishing grounds in the northeast China, and their phenotypic and genetic characteristics were investigated. All of the isolates were identified as A. hydrophila by traditional biochemical method and yielded a 686-bp DNA fragment of the 16S rDNA gene in the PCR experiments. Collected strains were also evaluated for their susceptibility to 17 different antibiotics. The isolates showed an even trend of the resistance and sensitivity to drugs, highly sensitive to antibiotics, such as Levofloxacin, PolymyxinB, Ofloxacin and resistant to antibiotics, such as Bristopen, Lincomycin, Ampicillin, Teicoplanin. Evaluation of genetic diversity was performed on all isolates by molecular typing with enterobacterial repetitive intergenic consensus (ERIC-PCR) method. The results showed that three different types, i.e. type I, ]I, and type ~I, were found in 59 isolates and type III accounted for a large proportion of 54.84%. There was no dominant difference between the tendency of the isolates of Heilongjiang Province and Jilin Province in these three types, which showed DI 〉 I 〉 I, while the isolates of Liaoning Province showed III 〉 II 〉 I. The percentage of different types in different provinces varied in each other; however, they didn't show any obvious regional or cluster-specific branches. In conclusion, the ability to distinguish Aeromonas hydrophila strains from diseased common carp with ERIC-PCR would be useful for epidemiological investigation and population genetic analysis of this pathogen in China.展开更多
In order to understand in vitro antibacterial effect of antibiotics and EM on Aeromonas hydrophila,14 isolates of A. hydrophila were isolated from diseased fish and aquaculture in the study. PCR amplification results ...In order to understand in vitro antibacterial effect of antibiotics and EM on Aeromonas hydrophila,14 isolates of A. hydrophila were isolated from diseased fish and aquaculture in the study. PCR amplification results of aerolysin gene( Aero gene) of isolates indicated that 7 out of 14 isolates were pathogenic A.hydrophila. In vitro antibacterial test indicated that 14 isolates were resistance to penicillin G,ampicillin,cephradine,cefazolin,tetracycline,terimethoprim,lincomycin and cephalexin,but showed high sensitivity to cefotaxime and furazolidone. In vitro antibacterial effect of EM on 14 isolates of A. hydrophila was positively correlated with concentration,and EM with the concentration greater than 60% had antibacterial effect on 14 isolates.展开更多
Non-specific immune response of bullfrog Rana catesbeiana to pathogenic Aeromonas hydrophila was studied to 60 individuals in two groups.Each bullfrog in bacterium-injected group was injected intraperitoneally(i.p.) w...Non-specific immune response of bullfrog Rana catesbeiana to pathogenic Aeromonas hydrophila was studied to 60 individuals in two groups.Each bullfrog in bacterium-injected group was injected intraperitoneally(i.p.) with 0.2 ml bacterial suspension at a density of 5.2 × 106 CFU/ml,while each one in control group injected i.p.with 0.2 ml sterile saline solution(0.85%,w/v).Three bullfrogs in both groups were sampled at 0,1,3,7,11,15 and 20 days post-injection(dpi) for the evaluation of non-specific immune parameters.It was observed that intraperitoneal injection of A.hydrophila significantly increased the number of leucocytes and that of NBT-positive cells in peripheral blood.Significant increases in serum bactericidal activity and serum acid phosphatase activity were also observed in the bacterium-injected frogs when compared with those in the control group.However,a significant reduction was detected in vitro in phagocytosis activity of peripheral blood phagocytes.No significant difference in changes in the number of peripheral erythrocytes,serum superoxide dismutase(SOD) activity,and lysozyme activity was detected between the two groups.It is suggested that bullfrogs may produce a series of non-specific immune reactions in response to the A.hydrophila infection.展开更多
基金Supported by Outstanding Graduate Entering Laboratory Project of College of Fisheries,Guangdong Ocean UniversityNational Natural Science Foundation of China(32073015)+1 种基金Undergraduate Innovation and Entrepreneurship Training Program of Guangdong Ocean University(CXXL2023008)Undergraduate Innovation Team of Guangdong Ocean University(CCTD201802).
文摘[Objectives]To explore the function of hcp gene in Aeromonas hydrophila.[Methods]A pair of specific primers was designed referring to the hcp gene sequence of A.hydrophila.The hcp gene was amplified by PCR,and performed bioinformatics analysis.[Results]The hcp gene had a total length of 1650 bp and encoded 549 amino acids.The theoretical molecular weight of the protein predicted was about 59476.44 kDa.After predicting the N-terminal signal peptide structure of the amino acid sequence,neither obvious signal peptide cleavage site nor signal peptide was found,and the protein had no transmembrane region.The amino acid sequence had a N-glycosylation site,4 protein kinase C phosphorylation sites,7 casein kinase II phosphorylation sites,9 N-myristoylation sites,4 isoprene binding sites,10 microbody C-terminal target signal sites,and an ATP/GTP binding site motif A(P-ring).The amino acid sequence of hcp gene of A.hydrophila was performed homology analysis with other Aeromonas strains,and it showed higher homology with A.veronii.In the secondary structure,theα-helix,β-sheet,random coil and extended strand accounted for 45.36%,6.01%,37.52%and 11.11%,respectively.The tertiary structure model consisted of 18α-helix and 22β-sheet.Analysis of protein-protein network interaction demonstrated that the proteins interacting with Hcp protein were AHA_3407,nrfA,nirB-1,nirB-2 and AHA_1112.[Conclusions]Through the bioinformatics prediction results,the basic information of hcp gene of A.hydrophila is preliminarily understood,and the possible function of this protein is predicted,in order to provide guidance for subsequent vaccine research.
基金Supported by the National Natural Science Foundation of China(No.32102824)the Zhejiang Provincial Natural Science Foundation of China(No.LQ18B060005)the Zhejiang Sci-Tech University Foundation(Nos.11612932611702,11612832611909)。
文摘Aeromonas hydrophila is a Gram-negative pathogen that can infect various fish,including largemouth bass(Micropterus salmoides),which have caused huge economic losses.In present study,largemouth bass anti-A.hydrophila antibodies were produced,then a highly immunogenic outer membrane proteins,AHA_3793,was identified by combined western blotting and mass spectrometry analysis.Moreover,AHA_3793 was expressed,and its immunogenicity was further verified by western blotting.Subsequently,the protective efficacy of AHA_3793 were evaluated in largemouth bass.The results showed that rAHA_3793 could produce a relative percentage survival(RPS)of 61.76% for largemouth bass against A.hydrophila challenge.ELISA analysis showed the specific serum antibodies of largemouth bass against rAHA_3793 and A.hydrophila in vaccinated group in weeks 4 and 5 after immunization were significantly higher than those in control group,which suggested that rAHA_3793 induced production of specific serum antibodies against rAHA_3793 and A.hydrophila.The qRT-PCR analysis showed that expressions of CD4-2 and MHC Ⅱα were also significantly up-regulated after immunization.These results collectively demonstrated that rAHA_3793 could induce a strong humoral immune response of largemouth bass,and then produce high immune protection ef fects against A.hydrophila infection.
文摘Rationale:Aeromonas hydrophila,a bacterium commonly found in a variety of soils,aquatic habitats,and other natural environments,is known for causing infections and hemorrhagic diseases in both aquatic and terrestrial animals.Patients with leukemia,when infected by Aeromonas hydrophila following chemotherapy,face an increased risk of developing necrotizing fasciitis.Patient concerns:A 48-year-old male with a history of acute myeloid leukemia presented with swelling and pain in his right hand after being stabbed by a grass carp two days ago.The injury occurred on the fifth day after his last chemotherapy session when he was accidentally punctured in the right thumb and ring finger by a carp while fishing.Two days following the injury,the skin isurrounding the wound turned black immediately,while the rest of his hand was reddened,swollen,and bleeding.Diagnosis:Aeromonas hydrophila infection in the soft tissue of the right hand.Interventions:Intravenous antibiotics and local debridement.Outcomes:The patient experienced an alleviation of systemic poisoning symptoms,effective control of the local wound infection.Lessons:It is important for physicians to appreciate the potential for highly unusual and life-threatening infections in patients with acute myeloid leukemia.Early diagnosis and prompt treatment can prevent the development of necrotizing fasciitis and save patients’lives.
文摘嗜水气单胞菌是水产养殖中一种比较常见的致病病原菌。本研究以嗜水气单胞菌为研究对象,利用SELEX指数富集系统进化技术(Systematic Evolution of Ligands by Exponential Enrichment,SELEX),经过12轮的筛选,通过生物素-抗地高辛碱性磷酸酶显色系统检测DNA适配子与嗜水气单胞菌的亲和力,克隆、测序后,用DNAMAN软件分析其一级序列与二级结构。研究结果表明,通过12轮筛选后,亲和力由最初的0.32升到第9轮的0.72,亲和力提高了2.25倍。一级结构主要由GT组成,说明同源性适体有了一定程度的富集。二级结构模拟图以茎环状结构为主,说明这些茎环状结构可能是适体与靶分子进行结合作用的基础,也是其进一步形成三维空间结构的基础。
基金Supported by the Corporation Project(09003699)Foundation of the Education Department of Jiangxi Province(GJJ150416)Foundation of the Science and Technology Department of Jiangxi Province(20122BBF60082)~~
文摘[Objective] This study was conducted to develop compound probiotics that can be used as the alternatives of chemical drugs and antibiotics in aquaculture. [Method] Different concentrations of EM (effective microorganisms) and Bacillus amyloliquefaciens were mixed at a ratio of 1:1, and sprayed on fish feed. The growth of intestinal villi of the fishes that had been fed with the feed mixed with compound probiotics for three months was observed; meanwhile, the content of in- testinal Aeromonas hydrophila was detected by real-time quantitative PCR. |Result] The compound probiotics promoted the development of intestinal villi, and inhibited the growth of A. hydrophila, and the effects were also concentration dependent. However, the compound probiotics did not increase the thickness of the intestinal serous layer, muscular layer and submucosal layer. [Conclusion] The compound probiotics we prepared can be used and popularized in aquiculture as it can inhibit the growth of A. hydrophila.
基金supported by the Korea Research Foundation Grant funded by the Korean Government(MOEHRD,Basic Research Promotion Fund) (No.KRF-2007-313-D00402)
文摘A strain WL-11 with high laccase activity was isolated from activated sludge collected from the effluent treatment plant of a textile and dyeing industry. It was identified as Aeromonas hydrophila by physiological test and 16S rDNA sequence analysis. A gene encoding of laccase from a newly isolated Aeromonas hydrophila WL-11 was cloned and characterized. Nucleotide sequence analysis showed an open reading frame of 1605 bp encoding a polypeptide comprised of 534 amino acids. The primary structure of the enzyme predicted the structural features characteristic of other laccases, including the conserved regions of four histidine-rich copper-binding sites. The predicted amino acid sequence showed a high homology (more than 60%) with bacterial laccases in the genome and protein databases and the highest degree of similarity (61% identity) was observed with the multicopper oxidase of KlebsieUa sp. 601. When expressed in Escherichia coli, the recombinant enzyme was overproduced in the cytoplasm as soluble and active form. The purified enzyme had an optimum pH of 2.6 and 8.0 for ABTS (2,2'-azino-bis(3-ethylbenzthiazolinesulfonic acid) and DMP (2,6-dimethoxyphenol), respectively. The kinetic study on ABTS revealed a higher affinity of this enzyme to this substrate than DMP.
基金Project (No. 2004C26026) supported by the Science and Technology Department of Zhejiang Province, China
文摘Specific immunoglobulin (IgY) from egg yolk against Aeromonas hydrophila was produced by immunization of White Leghorn hens with formalin-killed whole cells of A. hydrophila. ELISA test using A. hydrophila as the coating antigen revealed that the specific antibody titer started to increase in the egg yolk at the 13th day post-immunization (P/N=2.18), reached the peak at the 56th day (P/N=13.82), and remained at high level until day 133 (P/N=7.03). The antibody was purified by saturated ammonium sulphate with a recovery rate of 63.5%. The specific IgY inhibited the growth of A. hydrophila at a concentration of 1.0 mg/ml during the 18 h incubation. Pre-treatment of polyploid gibel carps Carassius auratus Gibelio with specific IgY had a protection rate of 60% (6/10) against challenge with A. hydrophila, while none of the fishes in the control groups receiving sterile phosphate buffered saline (PBS) or non-specific IgY survived the challenge. Treatment of fishes with the specific IgY 4 h after the challenge also had lower mortality (70%, 7/10), a 30% reduction against the control PBS or non-specific IgY groups (10/10). These results indicate that specific IgY antibodies could be obtained easily from hens immunized with an inactivated A. hydrophila and could provide a novel alternative approach to control of diseases in fishes caused by this organism.
基金the National Natural Science Foundation of China(31072151)the Specialized Research Fund for the Doctoral Program of Higher Education,China(20090097110007)the Priority Academic Program Development of Jiangsu Higher Education Institutions,China(PAPD)
文摘Aeromonas hydrophila isolates from clinical cases (n=43) were tested against 8 antimicrobial agents and typed by outer membrane protein (OMP) pattern by using sodium dodecyl sulfate gel electrophoresis. All isolates were resistant to ampicillin (MICs, ≥16 μg mL-1) and sulfamonomethoxine (MICs≥64 μg mLl), but susceptible to norfloxacin (MICs,≤0.5 μg mL-1). There was a high incidence of resistance to erythromycin (90.70%) and tylosin (93.02%), while a low incidences of resistance to ciprofloxacin (2.33%), enrofloxacin (2.33%) and florfenicol (4.65%). Six different outer membrane protein patterns were found among 34 isolates by analyzing proteins in the range of 22 to 50 kDa, other than 9 isolates with their respective profiles. The strains with the similar OMP profiles had similar resistances. Compared with the other strains from the same OMP patterns, NB-1, A.Pun and MR-1 had lacked the proteins in the range of 30 to 45 kDa and their resistance to florfenicol substantially increased. It is speculated that the outer membrane protein changes might correlate with decreased susceptibility to florfenicol in the three strains. Some strains which showed completely identical OMP types had a little difference in their resistance to fluoroquinolones, indicating that there might be other factors that were involved in the antimicrobial resistance of A. hydrophila.
文摘The Aeromonas species, belonging to the family Aeromonadaceae, are opportunistic pathogens found in humans with an incidence rate of 76 cases per million inhabitants in Southern Taiwan.The incidence of Aeromonas septicemia is relatively low, accounting for less than 15% of cases.Patients diagnosed with Aeromonas hydrophila bacteremia who were presented with skin blisters and septic shock have been reported to have a mortality rate of 100%.Aeromonas infection must be considered in the differential diagnosis of gangrene-like tissue damage or skin lesions in patients with end-stage renal disease, due to the potential sources of infections.A 49-year-old Taiwan Residents diabetic woman with end-stage renal disease had underwent regular hemodialysis.She was referred to our hospital due to a one-day course of fever, dyspnea, hypotension, and fulminant hemorrhagic blisters covering her whole body.A physical examination uncovered multiple hemorrhagic blisters, along with a ruptured blister over the lower left leg.Laboratory tests revealed an elevation of liver enzymes, impaired renal function, lactatemia, and high anion-gap metabolic acidosis.Cultures of both blood and hemorrhagic blister fluid grew Aeromonas hydrophila.However, she experienced persistent shock despite aggressive intravenous fluid, empiric antibiotics, and inotropic agents with norepinephrine and dopamine.Early diagnosis and prompt management using intravenous fluids, antibiotics and surgical debridement is recommended in order to improve a patient's survival rate.
文摘The ubiquitous Aeromonas hydrophila is responsible for several pathological conditions in fish and human. Like most gram negative bacteria, its virulence relies on outer membrane lipopolysachharide (LPS). The Leloir pathway enzyme UDP-galactose 4-epimerase (GalE), plays an important role in the LPS biosynthesis, and therefore is a potential drug target. We have earlier carried out extensive biochemical and biophysical studies with histidine-tagged recombinant GalE. However, for effective drug design it is desirable to understand the structure-function relation of a protein in its native form without any additional sequences or tags. In the present study, we report the high level expression, purification and characterization of recombinant GalE (rGalE) of Aeromonas hydrophila in its native form in E coli. The rGalE expressed as a soluble protein was purified to near homogeneity. From 1 L of shake flask culture ~15 mg of purified rGalE was obtained. The purified protein was biologically active with Km and Kcat values of 0.7 mM and 28.8 s–1, respectively. The enzyme exhibited a temperature and pH optima of 37。C and 7 - 9, respectively. Thus, the present study employed for soluble expression and purification of functionally active rGalE without any tag bypasses the need for cumbersome strategies associated with removal of tag from purified protein.
基金supported by the Research Grant from the Science and Technology Department of Sichuan, China (2011JY0095)the Construction Project of Southwest University for Nationalities (2011XWD-S071007)
文摘The Chinese giant salamander(Andrias davidianus) is the largest living amphibian in the world. Aeromonas hydrophila strain L602 was isolated from A. davidianus. The 16S rDNA gene of this isolate was amplified using PCR,and the phylogenetic tree was constructed by the neighbor-joining method. Four virulence genes(aerA,aha1,hly and alt) of A. hydrophila were amplified by PCR and drug resistances were tested using Kirby-Bauer disk diffusion method. The results showed that the length of this 16S rDNA sequence was 1453 bp,which showed 99% homology with A. hydrophila. The GenBank accession number was JX155398. Phylogenetic analysis indicated it grouped together with A. hydrophila. Four virulence genes were all detected,indicating that strain L602 was highly virulent. This stain was resistant to four antibiotics(vibramycin,furazolidone,ampicillin and erythromycin),while it was insensitive to streptomycin. Furthermore,this strain was susceptible to six antibiotics(sulfafurazole,ciprofloxacin,penbritin,norfloxacin,florfenicol and enrofloxacin). This study will help to validate the classification and virulence of pathogenic bacteria in amphibians.
基金Supported by the Central-Level Non-profit Scientific Research Institutes Special Funds (201104)the Natural Science Foundation of Heilongjiang Province of China (C201140)
文摘A total of 59 isolates of Aeromonas hydrophila were collected from common carp suffering from freshwater fish hemorrhage disease in 13 fishing grounds in the northeast China, and their phenotypic and genetic characteristics were investigated. All of the isolates were identified as A. hydrophila by traditional biochemical method and yielded a 686-bp DNA fragment of the 16S rDNA gene in the PCR experiments. Collected strains were also evaluated for their susceptibility to 17 different antibiotics. The isolates showed an even trend of the resistance and sensitivity to drugs, highly sensitive to antibiotics, such as Levofloxacin, PolymyxinB, Ofloxacin and resistant to antibiotics, such as Bristopen, Lincomycin, Ampicillin, Teicoplanin. Evaluation of genetic diversity was performed on all isolates by molecular typing with enterobacterial repetitive intergenic consensus (ERIC-PCR) method. The results showed that three different types, i.e. type I, ]I, and type ~I, were found in 59 isolates and type III accounted for a large proportion of 54.84%. There was no dominant difference between the tendency of the isolates of Heilongjiang Province and Jilin Province in these three types, which showed DI 〉 I 〉 I, while the isolates of Liaoning Province showed III 〉 II 〉 I. The percentage of different types in different provinces varied in each other; however, they didn't show any obvious regional or cluster-specific branches. In conclusion, the ability to distinguish Aeromonas hydrophila strains from diseased common carp with ERIC-PCR would be useful for epidemiological investigation and population genetic analysis of this pathogen in China.
基金Supported by Cooperation Project(09003699)Project of Jiangxi Department of Education(GJJ12237)Project of Jiangxi Department of Science and Technology(20122BBF60082)
文摘In order to understand in vitro antibacterial effect of antibiotics and EM on Aeromonas hydrophila,14 isolates of A. hydrophila were isolated from diseased fish and aquaculture in the study. PCR amplification results of aerolysin gene( Aero gene) of isolates indicated that 7 out of 14 isolates were pathogenic A.hydrophila. In vitro antibacterial test indicated that 14 isolates were resistance to penicillin G,ampicillin,cephradine,cefazolin,tetracycline,terimethoprim,lincomycin and cephalexin,but showed high sensitivity to cefotaxime and furazolidone. In vitro antibacterial effect of EM on 14 isolates of A. hydrophila was positively correlated with concentration,and EM with the concentration greater than 60% had antibacterial effect on 14 isolates.
基金Supported by National High-Tech Research and Development Program of China (863 Program, No.2001AA5070, 2002AA639600)Natural Science Foundation of Fujian Province of China (No.B0410022, 2006F5066)
文摘Non-specific immune response of bullfrog Rana catesbeiana to pathogenic Aeromonas hydrophila was studied to 60 individuals in two groups.Each bullfrog in bacterium-injected group was injected intraperitoneally(i.p.) with 0.2 ml bacterial suspension at a density of 5.2 × 106 CFU/ml,while each one in control group injected i.p.with 0.2 ml sterile saline solution(0.85%,w/v).Three bullfrogs in both groups were sampled at 0,1,3,7,11,15 and 20 days post-injection(dpi) for the evaluation of non-specific immune parameters.It was observed that intraperitoneal injection of A.hydrophila significantly increased the number of leucocytes and that of NBT-positive cells in peripheral blood.Significant increases in serum bactericidal activity and serum acid phosphatase activity were also observed in the bacterium-injected frogs when compared with those in the control group.However,a significant reduction was detected in vitro in phagocytosis activity of peripheral blood phagocytes.No significant difference in changes in the number of peripheral erythrocytes,serum superoxide dismutase(SOD) activity,and lysozyme activity was detected between the two groups.It is suggested that bullfrogs may produce a series of non-specific immune reactions in response to the A.hydrophila infection.