Colonization and survival capacities underlying the multifaceted life of Rhodococcus sp. PBTS1 and PBTS2

Little is known about the epidemiology of the newly emerged disease on pistachio rootstock designated pistachio bushy top syndrome (PBTS). Here, we analysed the survival and colonization capacities of the two causative agents, Rhodococcus corynebacterioides-like PBTS1 and R. fascians PBTS2, in the different habitats they occupy. Additionally, we examined their chromosomal sequences for genes that would support their ecology and provide experimental evidence for some of their predicted functions. We demonstrate that the soil competence of PBTS2 is better than that of PBTS1 and that both bacteria behave chemotactically, allowing them to translocate from soil to the rhizoplane. Once on the rhizoplane and the phylloplane, the PBTS bacteria form extensive epiphytic biofilms, probably aided by exopolysaccharide production. From these biofilms, the bacteria colonize the plant's interior tissues through the formation of ingression sites, likely created by the localized activity of cell wall-degrading enzymes. The observed systemic spreading of the PBTS strains throughout the plant tissues is probably mediated by surface motility and agrees with the occurrence of symptoms in both the aerial and root parts of the plant host. Based on all available information, we provide a working model on this pathosystem to be used as a conceptual framework for future research.     

Identification of Equine Herpesvirus 5 in Horses with Lymphoma

Equine multinodular pulmonary fibrosis, equine herpesvirus 5 (EHV-5), and multicentric lymphoma were discovered in one patient. Review of gamma herpesvirus activity in humans revealed a propensity for lymphoproliferative disorders associated with infection. The objective was to determine the frequency of EHV-5 in lymphoma tissues and compare with the frequency found in the lymph nodes of clinically normal horses. Case control investigation of lymphoma-positive tissues and analysis via polymerase chain reaction (PCR) for EHV-5 was performed on 12 horses. Prospective collection and PCR analysis of lymph nodes (mesenteric or submandibular) for EHV-5 was performed on 21 control horses. Thirteen samples of lymphoma-positive tissues and fluid were submitted for PCR analysis for EHV-5. Of these, 67% was positive. In the control horse population, 14% was positive for EHV-5 (P = .004). Neoplastic samples positive for EHV-5 were classified as T-cell rich B-cell lymphoma (three), T-cell lymphoma (one), one was nondifferentiated, and two were not stained. Gamma herpesviruses in humans have been associated with lymphoproliferative diseases such as Kaposi sarcoma and Burkitt lymphoma. This study reveals an increased frequency of EHV-5 (gamma herpesvirus) in horses diagnosed with lymphoma compared with healthy control horses. Although the exact role this virus plays in the initiation or perpetuation of lymphoproliferative neoplasia is unknown, EHV-5 may be an etiologic agent associated with the development of some types of equine lymphoma.     

Equine Multinodular Pulmonary Fibrosis and Lymphoma in a Horse Associated with Equine Herpesvirus-5

A 9-year-old mare presented with a 2-week history of partial anorexia, multiple swellings in the area of the throatlatch, and purulent nasal discharge. On initial presentation, the horse had submandibular and retropharyngeal lymphadenopathy, mild ventral edema, and weight loss. Thoracic radiographs revealed a pulmonary interstitial pattern. Necropsy revealed enlarged lymph nodes throughout the body. The lung parenchyma contained multiple random, well-circumscribed nodules, which, on cut section, were pale, tan colored, and very firm with a distinct line of demarcation from the surrounding normal parenchyma. The subendocardium of the left ventricle, left atrium, and, multifocally, the right ventricle contained white, gritty areas of mineralization. There was marked subintimal mineralization of the aorta and pulmonary artery. Histopathology of the lymph nodes revealed effacement of the parenchyma by a neoplasm composed of large numbers of small mature lymphocytes, fewer large lymphocytes, and scattered moderate numbers of histiocytes. Immunohistochemistry tests for CD3, CD79a, and CD20, confirmed the lymphoma was T-cell-rich B-cell lymphoma. The lungs contained marked interstitial fibrosis with alveolar histiocytosis. Polymerase chain reaction test results of lymph node and lungs were positive for equine herpesvirus-5 (EHV-5), a gammaherpesvirus. Gammaherpesvirus infection has been associated with lymphoma and pulmonary fibrosis in other species. This report describes the association between EHV-5 and both pulmonary fibrosis and lymphoma.     

Treatment of Metastatic Equine Melanoma with a Plasmid DNA Vaccine Encoding Streptococcus Pyogenes EMM55 Protein

A 19-year-old castrated male Arab/Quarter horse presented with an extensive history of cutaneous metastatic melanoma. Over a period of 8 months, a total of 8 doses of plasmid DNA vaccine expressing the Streptococcus pyogenes emm55 gene (pAc/emm55) were administered intratumorally at 300 μg/dose via a needless injector. Upon completion of the vaccination protocol, the size of the injected lesions, on average, were reduced by 40.3% from the initial size measurements. Lesions that were not injected were reduced by 47.6%. The overall reduction in total tumor burden was 42.3%. Tumor regression was also associated with the augmentation of antimelanoma IgG antibody response, thus implying that an induction of an effective antimelanoma response would be of great advantage in the management of equine melanoma.     

Consequences for the bovine embryo of being derived from a spermatozoon subjected to post-ejaculatory aging and heat shock: development to the blastocyst stage and sex ratio

The objective was to determine whether aging of sperm caused by incubation at normothermic (38.5 C) or heat shock (40 C) temperatures for 4 h prior to oocyte insemination affects sperm motility, fertilizing ability, competence of the resultant embryo to develop to the blastocyst stage and blastocyst sex ratio. In the first experiment, the percent of sperm that were motile was reduced by aging (P<0.001) and the reduction in motility was greater for sperm at 40 C compared to sperm at 38.5 C (P<0.01). In the second experiment, oocytes were inseminated with aged sperm. A smaller percent of oocytes fertilized with sperm aged at either temperature cleaved by Day 3 after insemination than oocytes fertilized with fresh sperm (P<0.05). There was no effect of sperm aging on the percent of oocytes or cleaved embryos that developed to the blastocyst stage. Aging of sperm before fertilization at 38.5 C reduced the percent of blastocysts that were male (P=0.08). In the third experiment, incubation of sperm at 38.5 C or 40 C for 4 h did not reduce fertilizing ability of sperm as determined by pronuclear formation at 18 h post insemination. In conclusion, aging of sperm reduced cleavage rate and the percent of blastocysts that were males but had no effect on the developmental capacity of the embryo. The effect of aging on cleavage rate may represent reduced motility and errors occurring after fertilization and pronuclear formation. Aging at a temperature characteristic of maternal hyperthermia had little additional effect except that polyspermy was reduced. Results indicate that embryo competence for development to the blastocyst stage is independent of sperm damage as a result of aging for 4 h at normothermic or hyperthermic temperatures.     

Modulation of the immunogenicity of the Trypanosoma congolense cysteine protease, congopain, through complexation with ��2-macroglobulin

The protozoan parasite Trypanosoma congolense is the main causative agent of livestock trypanosomosis. Congopain, the major lysosomal cysteine proteinase of T. congolense, contributes to disease pathogenesis, and antibody-mediated inhibition of this enzyme may contribute to mechanisms of trypanotolerance. The potential of different adjuvants to facilitate the production of antibodies that would inhibit congopain activity was evaluated in the present study. Rabbits were immunised with the recombinant catalytic domain of congopain (C2), either without adjuvant, with Freund’s adjuvant or complexed with bovine or rabbit α₂-macroglobulin (α₂M). The antibodies were assessed for inhibition of congopain activity. Rabbits immunised with C2 alone produced barely detectable anti-C2 antibody levels and these antibodies had no effect on recombinant C2 or native congopain activity. Rabbits immunised with C2 and Freund’s adjuvant produced the highest levels of anti-C2 antibodies. These antibodies either inhibited C2 and native congopain activity to a small degree, or enhanced their activity, depending on time of production after initial immunisation. Rabbits receiving C2-α₂M complexes produced moderate levels of anti-C2 antibodies and these antibodies consistently showed the best inhibition of C2 and native congopain activity of all the antibodies, with maximum inhibition of 65%. Results of this study suggest that antibodies inhibiting congopain activity could be raised in livestock with a congopain catalytic domain-α₂M complex. This approach improves the effectiveness of the antigen as an anti-disease vaccine candidate for African trypanosomosis.     

Functional characterization of bovine TIRAP and MyD88 in mediating bacterial lipopolysaccharide-induced endothelial NF-κB activation and apoptosis

Mastitis is a prevalent disease in dairy cows. Gram-negative bacteria, which express the pro-inflammatory molecule lipopolysaccharide (LPS), are responsible for the majority of acute clinical cases of mastitis. Previous studies have identified differential susceptibility of human and bovine endothelial cells (EC) to the pro-inflammatory and injury-inducing effects of LPS. The Toll-like receptor (TLR)-4 signaling pathway, which is activated by LPS, has been well studied in humans, but not in ruminants. Human myeloid differentiation-factor 88 (MyD88) and TIR-domain containing adaptor protein (TIRAP) are critical proteins in the LPS-induced NF-κB and apoptotic signaling pathways. To assess the role of the bovine orthologs of these proteins in bovine TLR-4 signaling, dominant-negative constructs were expressed in bovine EC, and LPS-induced NF-κB activation and apoptosis evaluated. The results from this study indicate that bovine MyD88 and TIRAP play functional roles in transducing LPS signaling from TLR-4 to downstream effector molecules involved in NF-κB activation, and that TIRAP promotes apoptotic signaling.