In silico prediction of Gallibacterium anatis pan-immunogens

The Gram-negative bacterium Gallibacterium anatis is a major cause of salpingitis and peritonitis in commercial egg-layers, leading to reduced egg production and increased mortality. Unfortunately, widespread multidrug resistance and antigenic diversity makes it difficult to control infections and novel prevention strategies are urgently needed. In this study, a pan-genomic reverse vaccinology (RV) approach was used to identify potential vaccine candidates. Firstly, the genomes of 10 selected Gallibacterium strains were analyzed and proteins selected on the following criteria; predicted surface-exposure or secretion, none or one transmembrane helix (TMH), and presence in six or more of the 10 genomes. In total, 42 proteins were selected. The genes encoding 27 of these proteins were successfully cloned in Escherichia coli and the proteins expressed and purified. To reduce the number of vaccine candidates for in vivo testing, each of the purified recombinant proteins was screened by ELISA for their ability to elicit a significant serological response with serum from chickens that had been infected with G. anatis. Additionally, an in silico prediction of the protective potential was carried out based on a protein property prediction method. Of the 27 proteins, two novel putative immunogens were identified; Gab_1309 and Gab_2312. Moreover, three previously characterized virulence factors; GtxA, FlfA and Gab_2156, were identified. Thus, by combining the pan-genomic RV approach with subsequent in vitro and in silico screening, we have narrowed down the pan-proteome of G. anatis to five vaccine candidates. Importantly, preliminary immunization trials indicated an in vivo protective potential of GtxA-N, FlfA and Gab_1309.

Electronic supplementary material

The online version of this article (doi:10.1186/s13567-014-0080-0) contains supplementary material, which is available to authorized users.     

Wild ostrich (<Emphasis Type="Italic">Struthio camelus australis</Emphasis>) reproduction in Orbata,a nature reserve in Tunisia

The aim of the current investigation was to determine wild ostrich reproductive behaviour in Orbata Nature Reserve by observing 16 hens and 28 cocks over a seven-year period. Intense laying commenced in January, one month after the cessation of the rainy season, and 92% of the eggs were produced during the dry season (January to May, peaking in March). Over the seven years, 1,322 eggs were laid in 69 nests, which corresponded to an annual average production of 19.2 ± 9.1 eggs/nest and 11.8 eggs/hen. 24 nests (34.78%) were non-brooded, 17 nests (24.64%) were deserted in the course of incubation, and 28 nests (40.58%) possessed hatched eggs. All the non-incubated nests had egg losses equivalent to 46.6 ± 12.6%. Hatchability success of incubated eggs was 41.9 ± 12.0%. Ostriches tended to dig their nests adjacent to the reserve enclosure which had direct access by road and track, the latter subjecting them to human disturbance and predation. The systematic obstruction of these nests stimulated ostriches to build additional nests within the reserve perimeter. The authors discussed the results recorded in an ostrich flock in relation to the environmental factors (climatic factors, food disponibility and predation) and suggested possibilities for improved wildlife management.     

Oral eosinophilic granulomas in tigers (Panthera tigris)--a collection of 16 cases.

Oral eosinophilic granulomas were diagnosed in 16 tigers (Panthera tigris). All lesions were located on the hard or soft palate and typically consisted of flat or slightly raised circular ulcers. Histologic features of these lesions were essentially identical to those seen in oral eosinophilic granulomas of domestic cats and dogs. No clinical signs were noted in eight cases, though various degrees of inappetence, excessive salivation, and dysphagia were noted in the other eight tigers. Six cases were not treated. Treatment for the remaining 10 cases centered on corticosteroids and additional treatments included surgical removal, cryotherapy, antibiotics, and chlorpheniramine. Treatment with corticosteroids did appear to be effective in some cases, though lesions would worsen after cessation of therapy and no cases were cured. In addition, three cases developed complications possibly related to this corticosteroid therapy. The etiology of these lesions remains unknown, though an underlying allergic condition is likely.     

不同细胞因子对水牛原始生殖细胞传代培养的影响

为探讨不同细胞因子对水牛原始生殖细胞（PGCs）传代培养的影响，将PGCs分别采用A、B、C、D、E、F和G共7种培养基进行培养，即A组：基础液＋10ng／mL白血病抑制因子（LIF）＋10ng／mL碱性成纤维细胞生长因子（bFGF）；B组：基础液＋20ng／mL LIF＋20ng／mL bFGF；C组：基础液＋20ng／mL LIF＋10ng／mL bFGF；D组：基础液＋20ng／mL LIF；E组：基础液＋20ng／mL LIF＋20ng／mL bFGF＋20ng／mL干细胞因子（SCF）；F组：基础液＋20ng／mL LIF＋40ng／mL bFGF＋40ng／mL SCF；G组（对照组）：基础液。将机械法分离的PGCs小集落接种到水牛胎儿成纤维细胞（BEF）饲养层上进行传代培养。结果，原代时，A、B、C、D、E和F组的克隆数目都显著高于对照组（P％0．05），其中E和F组的克隆数目显著高于A组（P〈0．05），而与B、C、D组差异均不显著（P〉0．05）；1～8代时，B、C、D、E、F组的克隆数目显著高于A组（P〈0．05）；对照组传代数仅为2代，A组为5代，而B、C、D、E和F组均为8代以上。结果表明，在传代过程中，LIF起主要作用，20ng／mL的LIF浓度可以满足水牛PGCs传代培养的需要。     

Identification and localization of the structural proteins of anguillid herpesvirus 1

ABSTRACT: Many of the known fish herpesviruses have important aquaculture species as their natural host, and may cause serious disease and mortality. Anguillid herpesvirus 1 (AngHV-1) causes a hemorrhagic disease in European eel, Anguilla anguilla. Despite their importance, fundamental molecular knowledge on fish herpesviruses is still limited. In this study we describe the identification and localization of the structural proteins of AngHV-1. Purified virions were fractionated into a capsid-tegument and an envelope fraction, and premature capsids were isolated from infected cells. Proteins were extracted by different methods and identified by mass spectrometry. A total of 40 structural proteins were identified, of which 7 could be assigned to the capsid, 11 to the envelope, and 22 to the tegument. The identification and localization of these proteins allowed functional predictions. Our findings include the identification of the putative capsid triplex protein 1, the predominant tegument protein, and the major antigenic envelope proteins. Eighteen of the 40 AngHV-1 structural proteins had sequence homologues in related Cyprinid herpesvirus 3 (CyHV-3). Conservation of fish herpesvirus structural genes seemed to be high for the capsid proteins, limited for the tegument proteins, and low for the envelope proteins. The identification and localization of the structural proteins of AngHV-1 in this study adds to the fundamental knowledge of members of the Alloherpesviridae family, especially of the Cyprinivirus genus.     

Can the protozoan parasite Bonamia ostreae infect larvae of flat oysters Ostrea edulis?

Bonamia ostreae is an intracellular protistan parasite affecting flat oysters Ostrea edulis. It can be detected in juveniles but mortalities mainly affect oysters which are more than 2 years old. The parasite is usually observed inside haemocytes and sometimes free, notably in gill epithelia suggesting a parasite release through this organ. However, the infective form and ways of entry and release remain undetermined. Flat oysters incubate their larvae in their pallial cavity for 8-10 days before releasing them into the water column. Flat oysters in Bay of Quiberon in South Brittany (France) are known to be infected with B. ostreae since 1979 and is the most important area in France for O. edulis spat collection. Flat oysters incubating larvae were sampled in this area during summertime between 2007 and 2009. Both adults and larvae were preserved and assayed by PCR and in situ hybridisation (ISH). PCR tests revealed the presence of parasite DNA in some adults and larvae. Specific labelling could be detected by ISH in gills, digestive system, gonad and mantle in adults and in the epithelium surrounding the visceral cavity of some larvae. Our results demonstrate that larvae can be infected with B. ostreae. Larvae might thus contribute to the spread of the parasite during their planktonic life. In addition, their transfer for aquaculture purpose should be controlled especially when they are exported from infected zones.     

A Novel Approach to the Treatment and Prevention of Laminitis: Botulinum Toxin Type A for the Treatment of Laminitis

Laminitis is an inflammation of the lamina of hoofed animals. According to the United States Department of Agriculture (USDA), laminitis impacts approximately 2% of the horse population each year. Because of the severity of the inflammation and chronic pain, it is frequently necessary to euthanize these horses. Surviving horses may be left useless, with resultant economic and social impact on the industry. Current interventions for laminitis are based on supportive care and alteration of biomechanical forces on the digit. The transition from laminitis to laminar failure (founder) is believed to be caused by the “weight of the horse and the forces of locomotion driving the bone down into the hoof capsule.” We hypothesize that an important factor is the torsional forces placed on the distal phalanx (coffin bone) by the deep digital flexor muscle and tendon. The paralyzing effect of botulinum toxin will result in a decrease of these torsional forces and therefore will aid in the prevention or treatment of the sequelae of laminitis. Seven horses with varying degrees of laminitis were evaluated. Each underwent pretreatment and posttreatment radiographs as well as pretreatment and posttreatment Obel grading. Each horse received botulinum toxin type A injected into the belly of the deep digital flexor muscle. In all cases there was radiographic stabilization of pedal displacement from the dorsal hoof wall. Obel scores showed improvement of 1 to 2 grades during the time monitored.