黄芪多糖和低浓度嗜水气单胞菌对中华绒螯蟹免疫相关因子基因表达的影响

采用逆转录-聚合酶链反应(RT-PCR)方法研究黄芪多糖和低浓度嗜水气单胞菌对中华绒螯蟹免疫相关因子基因表达的影响。结果表明:注射低浓度嗜水气单胞菌后,河蟹血细胞抗脂多糖因子(ALF)、热休克蛋白(HSP)、抗菌肽(Crus1)、脂肪酸结合蛋白酶(EsFABP3)、JAK通路细胞受体因子(EsDOME)、JAK通路信号转导与转录激活因子(EsSTAT)、河蟹肝胰腺HSP、EsFABP3和EsSTAT及河蟹鳃组织HSP的基因表达水平显著上调(P<0.05),黄芪多糖能够显著提高河蟹血细胞中ALF、酚氧化酶(PO)、HSP、Crus1、Toll样受体基因(TLR)、EsFABP3、EsJAK、EsDOME、EsSTAT、河蟹肝胰腺中ALF、PO、谷胱甘肽过氧化物酶(GPX)、HSP、TLR、EsFABP3、EsJAK、EsSTAT及河蟹鳃组织中ALF、GPX、HSP、Crus1、masquerade样蛋白(MasL)、TLR、EsJAK、EsDOME和EsSTAT的基因表达水平(P<0.05)。除了HSP和MasL外,注射黄芪多糖的河蟹免疫相关因子的基因表达水平普遍高于嗜水气单胞菌组河蟹。     

嗜水气单胞菌弹性蛋白酶基因的克隆表达

为了研究嗜水气单胞菌重组弹性蛋白酶的酶学性质,试验根据GenBank中的嗜水气单胞菌弹性蛋白酶基因ahyB设计1对含酶切位点的特异引物,以嗜水气单胞菌J-1(AhJ-1)株为模板,经PCR扩增得到不含信号肽的成熟弹性蛋白酶基因片段(787 bp),并与pMD18-T载体连接、测序,再用DNAStar软件分析。结果表明:该基因片段与豚鼠气单胞菌胞外蛋白酶同源性高达95%,与嗜水气单胞菌AG2株弹性蛋白酶ahyB基因同源性为92%,与铜绿假单胞菌LasB基因同源性为82%;将PCR产物连入表达载体pET-32a,转化至大肠杆菌BL21菌株中进行诱导表达,出现50 ku的融合表达蛋白,该表达产物纯化复性后表现出酶的活性。     

Role of microfilaments and microtubules in the invasion of EPC cells by Aeromonas hydrophila

The influence of the cytoskeleton on the invasion of Aeromonas hydrophila strain AhJ-1, isolated from diseased fish, in the monolayer cell of epithelioma papillosum cells of carp (EPC) was evaluated by the recovery of gentamicin-resistant bacteria from Triton X-100 cell lysates. The depolymerization of microfilaments (MF) by cytochalasin B and D inhibited the uptake of A. hydrophila in a dose-dependent manner and that of microtubules (MT) by colchicines and nocodazole did not affect the invasion of A. hydrophila in EPC cells significantly. The invasion frequency decreased approximately 62% with the addition of 0.1 microg/ml cytochalasin D and nearly 86% by the addition of 5.0 microg/ml. Invasion decreased approximately 49% and 83% by addition of cytochalasin B in a concentration of 2.5 microg/ml and 10.0 microg/ml. Colchicine and nocodazole, inhibitors of MT formation appears to have little effect on the invasion of EPC cells by strain Ah J-1. Thus MF formation, but not MT formation seems to play an important role in the internalization of A. hydrophila J-1.     

Clinical, haematological and blood biochemical changes in goats after experimental infection with tick-borne fever

Tick-borne fever in goats caused by Ehrlichia (Cytoecetes) phagocytophila was characterised by high fever, dullness, anorexia, tachycardia and a slight to moderate inhibition of rumen motility. The animals developed a gradual decline in the total number of circulating white blood cells. There was a decrease in lymphocytes over a short period, followed by an increase. The number of neutrophils was higher on the 3rd day, causing considerable change in the lymphocyte:neutrophil ratios. The number of eosinophils increased slightly. Serum alkaline phosphatase (ALP) decreased during the febrile episodes, and a marked decline was observed in both plasma zinc and iron concentrations. Furthermore, there was a small but progressive decrease of haemoglobin and haematocrit values. Circulating endogenous pyrogen/leucocyte endogenous mediator could not be detected in plasma from febrile goats. Tick-borne fever was passively transmitted to kids with plasma obtained from these febrile animals.     

In vitro and in vivo studies of the use of some medicinal herbals against the pathogen Aeromonas hydrophila in goldfish

Aeromonas hydrophila is a ubiquitous and opportunistic bacterial pathogen that produces ulcerative dermatitis under stress conditions and inflicts severe losses on global fisheries and fish culture. This study evaluates the antimicrobial potency of aqueous and ethanolic decoction (individual extract) and concoction (mixed extract) of three common medicinal herbs, turmeric Curcuma longa, Tulsi plant Ocimum sanctum, and neem Azadirachta indica, against the in vitro growth of A. hydrophila. Among the decoctions, A. indica exhibited the most potent antibacterial property (P < 0.05) against A. hydrophila. Among the concoctions, both the aqueous and ethanolic triherbal extracts mixed in the ratio of 1:1:1 had higher antibacterial activity (P < 0.05) than the other concoctions and decoctions. Goldfish Carassius auratus (10 +/- 2 g) were challenged with A. hydrophila intramuscularly in the caudal region with two separate doses (days 1 and 3) of 50 microL/fish (1.8 x 10(3) colony-forming units per milliliter). On days 9 (early) and 15 (late) of infection, fish were held in a net and dip treated for 5 min daily in a 1-L solution of 1% aqueous triherbal concoction. Red blood cell (RBC) count, hemoglobin, and hematocrit levels of the infected group were significantly higher (P < 0.05) than those of the control group. In the early treated group, all of the affected profile values returned to near normal, while the late-treated group registered a partial recovery, such as improved RBC count. The derived hematological values, such as mean corpuscular volume, mean corpuscular hemoglobin, and mean corpuscular hemoglobin concentration, of the early and late-treated groups also significantly declined (P < 0.05) but were restored to near normal (P > 0.05) only in the early treated group. The results suggest that dip treatment of A. hydrophila-infected goldfish in an aqueous triherbal concoction had a synergistic restorative effect on the hematological variables.     

Histopathology of the early phase during experimental Corynebacterium pseudotuberculosis infection in lambs.

Histological responses during experimental Corynebacterium pseudotuberculosis infection in lambs were investigated in parotid lymph nodes for ten days following inoculation. Lambs were infected by the subcutaneous route into the right eyelid with a virulent strain of C. pseudotuberculosis. Multiple microscopic acute abscesses, predominantly infiltrated with polymorphonuclear (PMN) leucocytes, were seen in the right parotid lymph node on the 1st day post-inoculation (PI). This massive PMN infiltration coincided with a peripheral blood granulocytosis. On day 3 PI, an influx of histiocytes was observed, while the microabscesses became confluent. From day 3 to day 10 PI, these lesions became enlarged and transformed into typical pyogranulomas with a central necrosis and a peripheral mantle of mononuclear cells composed of macrophages, epithelioid cells and lymphocytes; these histological changes were associated with a bacterial dissemination limited to the superficial lymph nodes. A lymphoid hyperplasia with prominent germinal centers was observed in the draining lymph nodes from day 3 PI. These results illustrate the dual role of granulomatous lesions in chronic bacterial infections: although they limit bacterial dissemination, the granulomas do not impair the persistence of infectious organisms in the host, leading to focal tissue damage.     

维氏气单胞菌TH0426株ompW基因的克隆、生物信息学分析及其原核表达

为研究维氏气单胞菌(Aeromonas veronii)ompW基因的生物信息学特性及相关功能,试验克隆了维氏气单胞菌ompW基因片段,通过生物信息学软件分析其编码蛋白的理化性质、疏水性、信号肽、跨膜区、二级及三级结构,并对其进行原核表达及反应原性检测。结果显示,ompW基因全长594 bp,编码197个氨基酸,TH0426株ompW基因与A.veronii B5B6株属于同一分支;该蛋白属于外膜蛋白,在19～29位氨基酸之间有一个典型的疏水区域,不存在跨膜区、信号肽,二级结构以β折叠与无规则卷曲为主。Western blot结果显示OmpW重组蛋白能与鼠抗维氏气单胞菌(A.veronii)TH0426株血清发生特异性结合。综上,OmpW重组蛋白具有一定的免疫原性,为进一步研究A.veronii疫苗奠定基础。     

Is there a relationship between haemoglobin genotype and the innate resistance to experimental Haemonchus contortus infection in Merino lambs?

Responses to a single or repeated infection with 7000 infective larvae of Haemonchus contortus were studied in an experiment using a total of 106 3-month-old lambs with AA, AB or BB haemoglobin (Hb) genotypes. Results were assessed by faecal egg counts, adult worm counts, haematocrit values, haemoglobin concentrations, total serum protein and serum antibody IgG1 and IgA ELISA titres. None of these parameters showed a strong relationship to the Hb type. The prevalence of low responder (greater than 500 worms) and of high responder (less than 50 worms) animals in groups AA, AB and BB Hb types was 3.8 and 34.6, 20.6 and 35.2, 28.1 and 43.7%, respectively, suggesting that the responsiveness to nematode infection is under the control of gene(s) not closely linked with those determining the Hb genotype. Worm counts of a primary infection are more subject to variation than those of a secondary infection. There is a strong relationship between adult worm counts and faecal egg counts taken close to the time of slaughter. In living animals low and high responder discrimination can be based on individual faecal egg counts around 50 days after a secondary infection. Haematocrit values proved to be of little value in the low and high responder selection. In this regard neither Hb concentration nor total serum protein values are of practical significance. In 3-month-old lambs primary infection induced partial immunity which could prevent the establishment of a part of the secondary infection, irrespective of the presence or absence of the primary worm population. The development of immunity was not associated with an increase of serum IgG1 and IgA antibody levels. Specific antibody production was not influenced by Hb types. Mean antibody levels of low responder lambs showed no difference from those of high responders. Thus, serum IgG1 and IgA levels are of no predictive value in identifying lambs which are genetically resistant to Haemonchus infection.     

Immunological and gene expression responses to a Salmonella infection in the chicken intestine

Besides infection in humans, Salmonella enteritidis can also cause serious illness in young chickens. However, the genetic and immunological parameters important for the disease in chickens are not well characterized. In this study, processes in the chicken intestine in response to a Salmonella infection were investigated in two different chicken lines. One-day-old chickens were orally infected with Salmonella. T-cell subpopulations, phagocytic properties of intestinal mononuclear cells and RNA expression levels of the jejunum were investigated. The two chicken lines differed in the amount of cfu in the liver and growth retardation after the infection. Differences in phagocytic activity of intestinal mononuclear cells were found between control and Salmonella infected chickens. The number of CD4+ T-cells of the intestine decreased after the Salmonella infection in one chicken line, while the number of CD8+ T-cells increased in both chicken lines, but the time post infection of this increase differed between the lines. In one chicken line the expression levels of the genes carboxypeptidase M and similar to ORF2 decreased after the Salmonella infection, which might be related to a decrease in the amount of macrophages. With the microarray, ten genes were found that were regulated in only one of the chicken lines, while we found six genes regulated in response to the infection in both chicken lines. So differences in genetic background of the chickens influence the intestinal host response of the Salmonella infection as observed by phagocytic activity, gene expression and changes in the number of T-cell subpopulations and macrophages.     

Temporal changes in the populations of immune cells at the site of experimental Dermatophilus congolensis infection in mice and sheep

Abstract The temporal patterns of dermal immune cell influx were compared in mice and sheep, species reputedly resistant and susceptible, respectively, to infection with Dermatophilus congolensis. In both species, the response involved early mast cell degranulation, vasodilatation and an influx of dendritic cells which accumulated and apparently differentiated beneath the infected epidermis. A concomitant dermal invasion by neutrophils and T and B lymphocytes led to epidermal infiltration, particularly by neutrophils and thence to the formation of the surface scab. Hypertrophy of the epidermis also indicates keratinocyte involvement in the host response. The duration of the response, however, was considerably shorter in the mouse (about 5 days) and B cells' were the predominant lymphocyte under and adjacent to the lesion. During the more protracted response in the sheep (> 21 days), T cells, including T19 antigen +γδ T cells, outnumbered B cells. Résumé— Des modifications dans les populations des cellules immunes dans le derme sont comparées chez la souris et le mouton, espèces respectivement résistantes et sensibles à Dermatophilus congolensis. Dans ces deux espèces, la réponse fait intervenir d'abord la dégranulation des mastocytes, la vasodilatation et l'arrivée de cellules dendritiques qui s'accumulent et se différencient apparemment sous l'épiderme infecté. Une invasion concomittente dermique par des neutrophiles, des lymphocytes T et B est observée et aboutit à l'infiltration épidermique, particulièrement par les neutrophiles et pour cette raison, à la formation de croûtes superficielles. L'hypertrophie de l'épiderme indique aussi l'implication du kératinocyte dans la réponse de l'hôte. Cependant, la durée de cette réponse, est bien plus courte chez la souris (environ 5 jours) et les lymphocytes B sont les lymphocytes prédominants, sous-jacents à la lésion. Chez le mouton, la réponse est prolongée (plus de 21 jours) et les lymphocytes T, incluant les lymphocytes CD 19 et γδ surpassent en nombre les lymphocytes B. [Sasiak, A.B., Lloyd, D.H., McEwan Jenkinson, D., Kitson, S., Elder, H. Y. Temporal changes in the populations of immune cells at the site of experimental Dermatophilus congolensis infection in mice and sheep (Particularités de la population cellulaire immunitaire aux sites d'infections expérimentales à Dermatophilus congolensis chez la souris et le mouton). Veterinary Dermatology 1996; 7 : 59–66.] ResumeAn Se compararon los patrones temporales de llegada de células inmunitarias en el ratón y en la oveja, especies consideradas resistentes y susceptibles, respectivamente a la infección por Dermatophilus congolensis. En ambas especies la respuesta implicó degranulación temprana de mastocitos, vasodilatación y llegada de células dendríticas que se acumulaban y aparentemente diferenciaban bajo la epidermis infectada. La invasión dérmica concomitante por netrófilos y linfocitos T y B llevó a una infiltración epidérmica, especialmente por neutrófilos y consecuentemente a la formación de una costra superficial. La hipertrofi de la epidermis también indica la implicación de queratinocitos en la respuesta del huésped. Sin embargo, la duración de la respuesta fue considerablemente más corta en el ratón (unos 5 dias) y los linfocitos B fueron la célula predominante en la zona adyacente a la lesión y debajo de ella. Durante la respuesta más prolongada en la oveja (> 21 días), las células T, incluyendo el antigeno T19, células T γδ, superaban en número las células B. [Sasiak, A.B., Lloyd, D.H., McEwan Jenkinson, D., Kitson, S., Elder, H. Y. Temporal changes in the populations of immune cells at the site of experimental Dermatophilus congolensis infection in mice and sheep (Alteraciones temporales en las poblaciones de celulas inmunitarias en zonas cutaneas de infeccion experimental por Dermatophilus congolensis en raton y en la oveja). Veterinary Dermatology 1996; 7 : 59–66.] Zusammenfassung— Die temporären Muster des Influx dermaler Immunzellen wurden bei Maus und Schaf verglichen, Tierarten, die angeblich resistent und empfänglich speziell für die Infektion mit Dermatophilus congolensis sind. Bei beiden Spezies bezog die Reaktion eine frühe Mastzelldegranulation ein, weiterhin Vasodilatation und einen Influx dendritischer Zellen, die sich unter der infizierten Epidermis anhäuften und offensichtlich differenzierten. Eine begleitende Invasion der Dermis durch Neutrophile, T- und B-Lymphozyten führte zu einer epidermalen Infiltration, besonders durch Neutrophile und von dort an zur Bildung von Oberflächenschorf. Die Hypertrophic der Epidermis zeigt außerdem eine Beteiligung der Keratinozyten bei der Antwort des Wirtstieres. Die Dauer der Reaktion jedoch war bei der Maus beträchtlich kürzer (etwa 5 Tage) und die B-Zellen waren die vorherrschenden Lymphozyten unter und angrenzend an die Läsion. Während der mehr protrahierten Reaktion beim Schaf (> 21 Tage) übertrafen die T-Zellen einschließlich T19-Antigen und gamma-delta-T-Zellen die Anzahl der B-Zellen. [Sasiak, A. B., Lloyd, D. H., McEwan Jenkinson, Kitson, S., Elder, H. Y. Temporal chnges in the populations of immune cells at the site of experimental Dermatophilus congolensis infection in mice and sheep (Temporäre Veränderungen in den Populationen von Immunzellen an der Stelle experimenteller Infektionen mit Dermatophilus congolensis bei Maus und Schaf). Veterinary Dermatology 1996; 7 : 59–66.]     

Withdrawn: Effects of OmniGen‐AF on blood neutrophil gene expression,milk leucocyte populations and production performance of early lactating dairy cows

The following article from Journal of Animal Physiology and Animal Nutrition, ‘Effects of OmniGen‐AF on blood neutrophil gene expression, milk leucocyte populations and production performance of early lactating dairy cows’ by R. D. Schramm, S. L. Shields, P. Rezamand, K. S. Seo, M. A. McGuire, published online on 6th July 2011 in Wiley Online Library ( wileyonlinelibrary.com ), has been withdrawn by agreement between the journal Editors and Blackwell Verlag GmbH. The article has been withdrawn for legal reasons.     

Interferon-gamma expression associated with suppression of bovine leukemia virus at the early phase of infection in sheep

Immunological control of bovine leukemia virus (BLV)-infection has been reported as dependent on the expression balance of types 1 and 2 cytokines. In this report, mRNA expression of interferon (IFN)-gamma and interleukin (IL)-2 (type 1 cytokines), and of IL-4 and IL-10 (type 2 cytokines) were evaluated in concanavalin A-stimulated peripheral blood mononuclear cells (PBMC) from BLV-infected sheep. Despite the same dose of BLV-infection, the extent of viral propagation was markedly different between eight individual sheep by 12 weeks post infection. The virus did not propagate well in three sheep, which showed augmented mRNA expression of IFN-gamma, a strong indicator of cell-mediated immunity, immediately after BLV-infection. Among the other five sheep having more than 2% of BLV-infected cells among PBMC at 12 weeks post infection, four sheep developed B-cell leukemia or lymphoma within 2 years after infection. These observations indicate IFN-gamma expression may play an important role in the protective mechanism against BLV propagation at the early phase of the infection.     

金属硫蛋白对奶牛血液抗氧化酶GSH-Px和CAT基因表达的影响

为探讨金属硫蛋白(MT)对奶牛抗氧化应激机能的调控效果及其机理,选取24头中国荷斯坦奶牛经产泌乳母牛,随机均分为I、II、III、IV 4组,分别按每头0(对照组),4.0,8.0和12.0 mg静脉注射Zn-MT.每隔15 d逐头采取血样,测定不同剂量的外源性MT和MT不同作用时间对奶牛抗氧化酶GSH-Px和CAT基因表达水平的影响.结果表明,补给外源性MT后,3个试验组GSH-Px和CAT基因表达水平都有不同幅度的提高,其中III组和IV组GSH-Px基因表达量分别较对照组有显著(P<0.05)提高.III组CAT基因表达水平显著(P<0.05)高于对照组;IV组CAT基因表达水平又比对照组、II组和III组分别提高了22.88%(P<0.01),17.71%(P<0.05)和7.10%(P>0.05).而II组GSH-Px和CAT基因表达量与对照组差异均不显著(P>0.05).从正试期间不同时间看,补给外源性MT后,各组GSH-Px和CAT基因表达水平逐步升高,至30 d时达最高,45 d时又有下降.尤其是30 d时的GSH-Px基因表达水平及15和30 d 时的CAT基因表达水平均显著(P<0.05)高于第1天.说明外源性MT能有效调控奶牛GSH-Px和CAT基因表达水平,并表现出较为明显的剂量效应和时间效应.     

The occurrence of Escherichia coli, Aeromonas hydrophila, Plesiomonas shigelloides and Clostridium perfringens in the intestinal flora of gray herons (Ardea cinerea)

The flora of the large intestine of 92 grey herons was examined for the frequency of aerobic and microaerobic growing bacteria. Clostridium perfringens, Aeromonas hydrophila, Plesiomonas shigelloides and E. coli were isolated from 55%, 48%, 14% and 35% of the birds, respectively. It could be demonstrated that the findings of these bacteria in the intestinal flora are depending on the age of the birds. The percentage of carriers of Clostridium perfringens, Aeromonas hydrophila and Plesiomonas shigelloides was highest in nestlings younger than 18 days, less high in older nestlings and lowest in adult grey herons. Contrary to those bacteria, E. coli was found more often in the intestinal flora at increasing age of the birds. Salmonella spp. were isolated from 6 birds. Two birds yielded positive for Yersinia enterocolitica and Campylobacter spp., respectively. Other aerobic and microaerobic bacteria play a less significant role as part of the intestinal flora.