牡丹红斑病的研究

牡丹红斑病在山东菏泽牡丹栽培区的发病率达50％左右，病重的牡丹园可达90％以上。从8批次384个样品中分离获得512个分离株，经筛选、纯化、回接试验，依其培养性状和形态特征，病原菌鉴定为牡丹枝孢Cladosporium paeoniae Pass。病害发生与牡丹不同品种、初侵染源等因子密切相关。筛选出了50％多菌灵、70％甲基托布津等防治红斑病的有效药剂，应用800倍液从3月中旬牡丹展叶前至7月中旬防治4次，效果达90％以上，感病指数可控制在5以下。     

Evaluation of dietary natural mineral materials as an antibiotic replacer on growth performance,non‐specific immune responses and disease resistance in rainbow trout,Oncorhynchus mykiss

We evaluated the effects of some dietary natural mineral materials as an antibiotic replacer based on growth performance, non‐specific immune responses and disease resistance in juvenile and subadult rainbow trout, Oncorhynchus mykiss. First experiment, juvenile rainbow trout averaging 2.7 ± 0.02 g (mean ± SD) were fed one of the six experimental diets; a basal commercial diet as a control (CON), CON with oxytetracycline (OTC), with yellow loess (YL), with Macsumsuk^® (MS), with Song‐Gang^® stone (SG) and with barley stone (BS) at 0.4% of each diet. At the end of 8‐week feeding trial, weight gain (WG), specific growth rate (SGR), feed efficiency (FE) and protein efficiency ratio (PER) of fish fed YL diet were significantly higher than those of fish fed CON diet. Non‐specific immune responses such as superoxide dismutase (SOD), myeloperoxidase (MPO), lysozyme (LYS) activity and oxidative radical production of fish fed YL diet were higher than those of fish fed CON diet. At the end of 15 days of challenge test with Aeromonas salmonicida, average cumulative survival rate of fish fed YL diet was significantly higher than that of fish fed BS and CON diets. However, there were no significant differences among fish fed YL, SG and OTC diets. Second experiment, subadult rainbow trout averaging 261.5 ± 3.5 g (mean ± SD) were fed one of the four experimental diets for 22 weeks: CON, and CON with OTC, YL or SG at 0.4% of each diet. At the end of feeding, growth performance of fish fed SG and YL diets was significantly higher than that of fish fed CON diet. Non‐specific immune responses in terms of SOD, MPO, LYS and NBT of fish fed SG and YL diets were significantly higher than those of fish fed CON diet. However, there were no significant differences among the fish fed YL, SG and OTC diets. The results indicate that dietary yellow loess or Song‐gang^® stone at 0.4% of diet could replace oxytetracycline in juvenile and subadult rainbow trout.     

Transgenic expression of a sorghum gene (SbLRR2) encoding a simple extracellular leucine-rich protein enhances resistance against necrotrophic pathogens in Arabidopsis

Plant leucine-rich repeat (LRR) domain-containing proteins are known to play important roles in signaling transduction and defense responses. In sorghum, SbLRR2 is pathogen-inducible gene encoding a simple extracellular LRR protein. Here, we demonstrated an earlier and stronger expression of SbLRR2 in a sorghum resistant genotype in comparison to a susceptible genotype following inoculation with the anthracnose pathogen (Colletotrichum sublineolum). In addition, SbLRR2 expression was found to be induced strongly by methyl-jasmonate treatment. Functional analysis was performed in SbLRR2 over-expression (OE) Arabidopsis plants, which showed enhanced resistance against the necrotrophic pathogens Botrytis cinerea and Alternaria brassicicola. In addition, the OE lines were found to have elevated expression of several jasmonate acid (JA)-associated genes and higher endogenous JA contents. Hence, the SbLRR2-mediated defense responses in transgenic Arabidopsis are likely to be dependent on JA-signaling through increased JA production. On the other hand, the OE lines remained susceptible to Pseudomonas syringae pv. tomato like the wild type plants. Consistently, there was no up-regulation of salicylic acid (SA) defense marker gene expression or SA levels in the OE lines. Our results suggested that SbLRR2 is potentially useful for enhancing resistance against necrotrophic pathogens in transgenic dicot crops.     

红树植物—白骨壤黑斑病病原菌的分离与鉴定

<正>红树林是生长在热带、亚热带海岸潮间带,可海陆两栖的特殊木本植物群落,具有防风御浪、净化环境、维护沿海湿地生物多样性等生态功能,为珍稀植物群落。有报道认为红树植物生长在海边又富含抗菌等活性物质,不利于病害发生,但近年来本实验室在广东雷州半岛等地红树林发现有多种病害发生~([1,2]),仅白骨壤(Avicennia marina)上就发现有黑茎病、灰斑病等病害。2014年我们在该区白骨壤小苗     

Pathogen Analysis of Goat's Respiratory Diseases

To investigate the pathogens of goat's respiratory disease from a goat farm in Guangxi province, the epidemiological investigation, clinic observation, pathological examination, bacterium isolation and identification, biochemical test, PCR and animal experiment test were conducted.The preventive and control measures were taken on the basis of pathogen epidemiological characteristics and drug sensitive test results.The results showed that Mycoplasma, influenza virus and parasite were negative by isolation and culture or PCR, however, two strains of gram-negative bacteria named MS1 and MS2 were isolated from lung.MS1 was identified as Serratia marcescens by biochemical test and 16S rRNA sequencing, which shared 99% nucleotide homology with other Serratia marcescens in GenBank.And MS2 was identified as E.coil by the same methods that shared 99% nucleotide homology with other E.coli in GenBank.Animal experiment showed that two strains could cause death in mice.The drug sensitive tests showed two strains were highly sensitive to spectinomycin, amikacin, kanamicin and neomycin.Kanamycin and dexamethasone were used to treat the sick goats, and achieved good results.     

Identification and pathogenicity study of emerging fish pathogens Acinetobacter junii and Acinetobacter pittii recovered from a disease outbreak in Labeo catla (Hamilton, 1822) and Hypophthalmichthys molitrix (Valenciennes, 1844) of freshwater wetland in West Bengal,India

The disease outbreaks in aquaculture system of wetlands are the major cause of fish mortality. Among various bacterial septicaemic diseases, fish mortality caused by Acinetobacter spp. is recently reported in different fish species. Fish disease outbreak was investigated in a wetland of West Bengal, India to identify the aetiological factors involved. The moribund fish were examined and subjected to bacterial isolation. Two bacterial causative agents were identified as Acinetobacter junii and Acinetobacter pittii by biochemical characterization and 16S rRNA gene amplification. Both the isolates were oxidase‐negative, nitrate‐negative, catalase‐positive and indole‐negative. The molecular identification using 16S rRNA gene sequencing and phylogenetic tree analysis further confirmed the two Acinetobacter spp. with 97%–99% similarity. The antibiotic resistance patterns of these two bacteria revealed that both of them were resistant to β‐lactam, cefalexin, cephalothin, amoxyclav, cefuroxime, cefadroxil, clindamycin, vancomycin and penicillin. In addition, A. pittii was also resistant to other antibiotics of cephams group such as ceftazidime and cefotaxime. In the challenge experiment, both A. junii and A. pittii were found to be pathogenic with LD₅₀ of 1.24 × 10⁵ and 1.88 × 10⁷ cfu/fish respectively. Histopathological examination of gill, liver and kidney revealed prominent changes supporting bacterial septicaemia. The investigation reports for the first time on concurrent infection by A. junii and multidrug‐resistant (MDR)‐A. pittii as emerging fish pathogens to cause severe mortality in Labeo catla and Hypophthalmichthys molitrix in a freshwater wetland.     

Stability comparison of three candidate internal reference genes in Chinese giant salamander (Andrias davidianus)

The Chinese giant salamander (Andrias davidianus) as food and medicinal product has been an important aquaculture object in China. Study of gene function in the Chinese giant salamander requires accurate normalization though the use of appropriate reference genes. In this study, the expression levels of three candidate reference genes including β‐actin, GAPDH and cytb of different tissues, different developmental stages and different challenges in Chinese giant salamander were evaluated by qPCR. The stabilities of these three reference genes were analysed by geNorm, NormFinder and BestKeeper software. The results showed that the expression of GAPDH was more stable than that of β‐actin and cytb in four tissues and at two developmental stages of Chinese giant salamander. Compared with GAPDH and cytb, β‐actin was the most stable in spleen of Chinese giant salamander treated with LPS or GSIV. Therefore, the result showed that GAPDH was the suitable reference gene in different tissues and at different developmental stages of Chinese giant salamander. The β‐actin could be used as a reference gene in spleen of Chinese giant salamander challenged with LPS and GSIV. This study provides convincing information for the GAPDH and β‐actin as suitable reference gene in Chinese giant salamander of different tissues, different developmental stages and different challenges respectively.     

Outbreak of viral haemorrhagic septicaemia (VHS) in lumpfish (Cyclopterus lumpus) in Iceland caused by VHS virus genotype IV

A novel viral haemorrhagic septicaemia virus (VHSV) of genotype IV was isolated from wild lumpfish (Cyclopterus lumpus), brought to a land‐based farm in Iceland, to serve as broodfish. Two groups of lumpfish juveniles, kept in tanks in the same facility, got infected. The virus isolated was identified as VHSV by ELISA and real‐time RT‐PCR. Phylogenetic analysis, based on the glycoprotein (G) gene sequences, may indicate a novel subgroup of VHSV genotype IV. In controlled laboratory exposure studies with this new isolate, there was 3% survival in the I.P. injection challenged group while there was 90% survival in the immersion group. VHSV was not re‐isolated from fish challenged by immersion. In a cohabitation trial, lumpfish infected I.P. (shedders) were placed in tanks with naïve lumpfish as well as naïve Atlantic salmon (Salmo salar L.). 10% of the lumpfish shedders and 43%–50% of the cohabiting lumpfish survived after 4 weeks. 80%–92% of the Atlantic salmon survived, but no viral RNA was detected by real‐time RT‐PCR nor VHSV was isolated from Atlantic salmon. This is the first isolation of a notifiable virus in Iceland and the first report of VHSV of genotype IV in European waters.     

Multilocus sequence typing detects new Piscirickettsia salmonis hybrid genogroup in Chilean fish farms: Evidence for genetic diversity and population structure

Piscirickettsia salmonisis the causative bacterial pathogen of piscirickettsiosis, a salmonid disease that causes notable mortalities in the worldwide aquaculture industry. Published research describes the phenotypic traits, virulence factors, pathogenicity and antibiotic‐resistance potential for various P. salmonisstrains. However, evolutionary and genetic information is scarce for P. salmonis. The present study used multilocus sequence typing (MLST) to gain insight into the population structure and evolution of P. salmonis. Forty‐two Chilean P. salmonisisolates, as well as the type strain LF‐89^T, were recovered from diseased Salmo salar, Oncorhynchus kisutchand Oncorhynchus mykissfrom two Chilean Regions. MLST assessed the loci sequences of dnaK, efp, fumC, glyA, murG, rpoD and trpB. Bioinformatics analyses established the genetic diversity among P. salmonis isolates (H = 0.5810). A total of 23 sequence types (ST) were identified, 53.48% of which were represented by ST1, ST5 and ST2. Population structure analysis through polymorphism patterns showed few polymorphic sites (218 nucleotides from 4,010 bp), while dN/dS ratio analysis indicated purifying selection for dnaK, epf, fumC, murG, and rpoD but neutral selection for the trpB loci. The standardized index of association indicated strong linkage disequilibrium, suggesting clonal population structure. However, recombination events were detected in a group of seven isolates. Findings included genogroups homologous to the LF‐89^T and EM‐90 strains, as well as a seven‐isolate hybrid genogroup recovered from both assessed regions (three O. mykiss and four S. salar isolates). The presented MLST scheme has comparative potential, with promising applications in studying distinct P. salmonis isolates (e.g., from different hosts, farms, geographical areas) and in understanding the epidemiology of this pathogen.     

Development of piscirickettsiosis in Atlantic salmon (Salmo salar L.) smolts after intraperitoneal and cohabitant challenge using an EM90‐like isolate: A comparative study

Piscirickettsiosis, caused by the intracellular Gram‐negative bacteria Piscirickettsia salmonis, is at present the most devastating disease in the Chilean salmon industry. The aim of this study was to analyse disease development after challenge with a P. salmonis strain (EM90‐like) under a controlled environment by comparing intraperitoneal challenge with cohabitation challenge. The P. salmonis EM90‐like isolate was cultured in a liquid medium for the challenge of 400 Atlantic salmon (Salmo salar) smolts. Cumulative mortality was registered, necropsy was performed, and bacterial distribution in the tissues and histopathological changes were analysed. The results revealed a similar progression of the disease for the two different challenge models. Pathological and histopathological changes became more visible during the development of the clinical phase of the disease. Bacterial DNA was identified in all the analysed tissues indicating a systemic infection. Bacterial tropism to visceral organs was demonstrated by real‐time quantitative PCR and immunohistochemistry. Better knowledge of disease development during P. salmonis infection may contribute to further development of challenge models that mimic the field situation during piscirickettsiosis outbreaks. The models can be used to develop and test future preventive measures against the disease.