Development of an aerogenous Escherichia coli infection model in adult broiler breeders.

Escherichia coli constitutes an immense challenge to the poultry industry due to its devastating effect on productivity, mortality, and carcass condemnations. To aid future studies on disease mechanisms and interventions, an aerogenous infection model was established in adult broiler breeders. Hens (n = 120) were randomly allocated into six groups receiving either aerosolised E. coli or vehicle, or intratracheal E. coli or vehicle. Replication of aerosol inoculation was performed on distinct days. Alternating euthanasia time points were predetermined in order to evaluate the progression of the disease. All animals were thoroughly necropsied, and bacteriological samples were collected as well as tissues for histopathology. Birds inoculated with E. coli exhibited clinical signs and developed characteristic gross and histopathological lesions of colibacillosis, including splenic fibrinoid necrosis, folliculitis, polyserositis and impaction of parabronchi with fibrinoheterophilic exudate and necrotic debris, as well as positive in situ localisation of intralesional E. coli by immunohistochemistry. This study presents a successful development of a discriminative colibacillosis model through aerosol inoculation of adult broiler breeders. Gross and histopathological lesions characteristic of colibacillosis were established in two independent experiments.

Comparison and assessment of necropsy lesions in end-of-lay laying hens from different housing systems in Denmark.

Apperantly healthy laying hens at the end of production (60 to 91 wk) were investigated for the occurrence of pathology and bacterial infections. In total, 7,477 hens from 15 flocks representing the following production systems: Enriched cages, barn housed layers, and organic/free range layers were necropsied. Indications of bacterial infection were investigated by bacteriological cultivation. The overall prevalence of lesions was 16.60%, including lesions of both infectious and non-infectious origin. The most prevalent lesions were bursitis presternalis (6.65%), reproductive tract lesions (e.g., salpingitis and/or peritonitis and/or oophoritis) (3.50%), serosal scarification (e.g., fibrotic adhesive peritonitis) 1.55%, and neoplasm 1.73%. Significant differences were observed between different production systems and/or flocks in the prevalence of reproductive tract lesions, bursitis presternalis, serosal scarification, skin infections, juvenile hens, and traumas/fractures. No significant difference was observed between different production systems in the prevalence of neoplasia, infection of septicemic etiology, and pododermatitis. In total, 3.4% of the hens were out of lay, with significantly higher rate in organic flocks. Infections of the reproductive tract were the most prevalent lesions with bacterial etiology in all productions systems. In total, 40% of the hens with lesions associated to the oviduct were out of lay and significant difference between production systems were observed. Escherichia coli was the most commonly isolated bacteria and in 90% of the cases they were isolated from the reproductive tract lesions. The second most prevalent bacteria was Gallibacteruim anatis. Significant difference in the prevalence of E. coli positive hens was observed between production systems (P < 0.05). In conclusion, the prevalence of reproductive tract lesions in apparently healthy end-of-lay laying was higher than indicated in previous reports. These findings support the previous suggestions that E. coli and G. anatis are the major pathogens causing reproductive tract lesions.

Injection Vaccines Formulated with Nucleotide, Liposomal or Mineral Oil Adjuvants Induce Distinct Differences in Immunogenicity in Rainbow Trout.

Protection facilitated by the widespread use of mineral oil adjuvanted injection vaccines in salmonid fish comes with adverse effects of varying severity. In this study, we characterized the immunological profiles of two alternative vaccine formulations, both with proven efficacy and an improved safety profile in rainbow trout. Experimental injection vaccines were prepared on an identical whole-cell Aeromonas salmonicida bacterin platform and were formulated with CpG oligodeoxynucleotides, a liposomal (CAF01) or a benchmark mineral oil adjuvant, respectively. A naïve group, as well as bacterin and saline-injected groups were also included. Following administration, antigen-specific serum antibody titers, the tissue distribution of immune cell markers, and the expression of immune-relevant genes following the in vitro antigenic restimulation of anterior kidney leukocytes was investigated. Immunohistochemical staining suggested prolonged antigen presentation for the particulate formulations and increased mucosal presence of antigen-presenting cells in all immunized fish. Unlike the other immunized groups, the CAF01 group only displayed a transient elevation in specific antibody titers and immunohistochemical observations, and the transcription data suggest an increased role of cell-mediated immunity for this group. Finally, the transcription profile of the CpG formulation approached that of a TH1 profile. When compared to the benchmark formulation, CAF01 and CpG adjuvants induce slight, but distinct differences in the resulting protective immune responses. This is important, as it allows a broader immunological approach for the future development of safer vaccines.

Modified MHC Class II-Associated Invariant Chain Induces Increased Antibody Responses against Plasmodium falciparum Antigens after Adenoviral Vaccination.

Adenoviral vectors can induce T and B cell immune responses to Ags encoded in the recombinant vector. The MHC class II invariant chain (Ii) has been used as an adjuvant to enhance T cell responses to tethered Ag encoded in adenoviral vectors. In this study, we modified the Ii adjuvant by insertion of a furin recognition site (Ii-fur) to obtain a secreted version of the Ii. To test the capacity of this adjuvant to enhance immune responses, we recombined vectors to encode Plasmodium falciparum virulence factors: two cysteine-rich interdomain regions (CIDR) α1 (IT4var19 and PFCLINvar30 var genes), expressed as a dimeric Ag. These domains are members of a highly polymorphic protein family involved in the vascular sequestration and immune evasion of parasites in malaria. The Ii-fur molecule directed secretion of both Ags in African green monkey cells and functioned as an adjuvant for MHC class I and II presentation in T cell hybridomas. In mice, the Ii-fur adjuvant induced a similar T cell response, as previously demonstrated with Ii, accelerated and enhanced the specific Ab response against both CIDR Ags, with an increased binding capacity to the cognate endothelial protein C receptor, and enhanced the breadth of the response toward different CIDRs. We also demonstrate that the endosomal sorting signal, secretion, and the C-terminal part of Ii were needed for the full adjuvant effect for Ab responses. We conclude that engineered secretion of Ii adjuvant-tethered Ags establishes a single adjuvant and delivery vehicle platform for potent T and B cell-dependent immunity.

Alternatives to mineral oil adjuvants in vaccines against Aeromonas salmonicida subsp. salmonicida in rainbow trout offer reductions in adverse effects.

In an effort to reduce the frequency and severity of adverse reactions seen from the use of mineral oil adjuvants in salmonid fish, the effects of two alternative adjuvants were assessed, focusing on the induction of adverse effects as well as protection. Using rainbow trout (Oncorhynchus mykiss) as recipients, injection vaccines based on formalin-inactivated Aeromonas salmonicida subspecies salmonicida were formulated with CpG oligodeoxynucleotides, the liposomal cationic adjuvant formulation 01 (CAF01) or with Freund's incomplete adjuvant and administered intraperitoneally. Control groups of unvaccinated, Tris-buffered saline-injected or bacterin-injected individuals were included, and each group included in the study held a total number of 240 individuals. Subsequently, individuals from each group were examined for differences in Fulton's condition factor, macro- and microscopic pathological changes, as well as protection against experimental infection with A. salmonicida. While adverse effects were not eliminated, reductions in microscopic and macroscopic adverse effects, in particular, were seen for both the nucleotide- and liposome-based vaccine formulations. Furthermore, the induced protection appears similar to that of the benchmark formulation, thus introducing viable, potential alternative types of adjuvants for use in future fish vaccines.

Pasteurization Procedures for Donor Human Milk Affect Body Growth, Intestinal Structure, and Resistance against Bacterial Infections in Preterm Pigs.

Background: Holder pasteurization (HP) destroys multiple bioactive factors in donor human milk (DM), and UV-C irradiation (UVC) is potentially a gentler method for pasteurizing DM for preterm infants.Objective: We investigated whether UVC-treated DM improves gut maturation and resistance toward bacterial infections relative to HP-treated DM.Methods: Bacteria, selected bioactive components, and markers of antioxidant capacity were measured in unpasteurized donor milk (UP), HP-treated milk, and UVC-treated milk (all from the same DM pool). Fifty-seven cesarean-delivered preterm pigs (91% gestation; ratio of males to females, 30:27) received decreasing volumes of parental nutrition (average 69 mL · kg-1 · d-1) and increasing volumes of the 3 DM diets (n = 19 each, average 89 mL · kg-1 · d-1) for 8-9 d. Body growth, gut structure and function, and systemic bacterial infection were evaluated.Results: A high bacterial load in the UP (6×105 colony forming units/mL) was eliminated similarly by HP and UVC treatments. Relative to HP-treated milk, both UVC-treated milk and UP showed greater activities of lipase and alkaline phosphatase and concentrations of lactoferrin, secretory immunoglobulin A, xanthine dehydrogenase, and some antioxidant markers (all P < 0.05). The pigs fed UVC-treated milk and pigs fed UP showed higher relative weight gain than pigs fed HP-treated milk (5.4% and 3.5%), and fewer pigs fed UVC-treated milk had positive bacterial cultures in the bone marrow (28%) than pigs fed HP-treated milk (68%) (P < 0.05). Intestinal health was also improved in pigs fed UVC-treated milk compared with those fed HP-treated milk as indicated by a higher plasma citrulline concentration (36%) and villus height (38%) (P < 0.05) and a tendency for higher aminopeptidase N (48%) and claudin-4 (26%) concentrations in the distal intestine (P < 0.08). The gut microbiota composition was similar among groups except for greater proportions of Enterococcus in pigs fed UVC-treated milk than in pigs fed UP and those fed HP-treated milk in both cecum contents (20% and 10%) and distal intestinal mucosa (24% and 20%) (all P < 0.05).Conclusions: UVC is better than HP treatment in preserving bioactive factors in DM. UVC-treated milk may induce better weight gain, intestinal health, and resistance against bacterial infections as shown in preterm pigs as a model for DM-fed preterm infants.

Further evidence of Chelonid herpesvirus 5 (ChHV5) latency: high levels of ChHV5 DNA detected in clinically healthy marine turtles.

The Chelonid herpesvirus 5 (ChHV5) has been consistently associated with fibropapillomatosis (FP), a transmissible neoplastic disease of marine turtles. Whether ChHV5 plays a causal role remains debated, partly because while FP tumours have been clearly documented to contain high concentrations of ChHV5 DNA, recent PCR-based studies have demonstrated that large proportions of asymptomatic marine turtles are also carriers of ChHV5. We used a real-time PCR assay to quantify the levels of ChHV5 Glycoprotein B (gB) DNA in both tumour and non-tumour skin tissues, from clinically affected and healthy turtles drawn from distant ocean basins across four species. In agreement with previous studies, higher ratios of viral to host DNA were consistently observed in tumour versus non-tumour tissues in turtles with FP. Unexpectedly however, the levels of ChHV5 gB DNA in clinically healthy turtles were significantly higher than in non-tumour tissues from FP positive turtles. Thus, a large proportion of clinically healthy sea turtle populations worldwide across species carry ChHV5 gB DNA presumably through persistent latent infections. ChHV5 appears to be ubiquitous regardless of the animals' clinical conditions. Hence, these results support the theory that ChHV5 is a near ubiquitous virus with latency characteristics requiring co-factors, possibly environmental or immune related, to induce FP.

Global distribution of Chelonid fibropapilloma-associated herpesvirus among clinically healthy sea turtles.

BACKGROUND: Fibropapillomatosis (FP) is a neoplastic disease characterized by cutaneous tumours that has been documented to infect all sea turtle species. Chelonid fibropapilloma-associated herpesvirus (CFPHV) is believed to be the aetiological agent of FP, based principally on consistent PCR-based detection of herpesvirus DNA sequences from FP tumours. We used a recently described PCR-based assay that targets 3 conserved CFPHV genes, to survey 208 green turtles (Chelonia mydas). This included both FP tumour exhibiting and clinically healthy individuals. An additional 129 globally distributed clinically healthy individual sea turtles; representing four other species were also screened. RESULTS: CFPHV DNA sequences were obtained from 37/37 (100%) FP exhibiting green turtles, and 45/300 (15%) clinically healthy animals spanning all five species. Although the frequency of infected individuals per turtle population varied considerably, most global populations contained at least one CFPHV positive individual, with the exception of various turtle species from the Arabian Gulf, Northern Indian Ocean and Puerto Rico. Haplotype analysis of the different gene markers clustered the CFPHV DNA sequences for two of the markers (UL18 and UL22) in turtles from Turks and Caicos separate to all others, regardless of host species or geographic origin. CONCLUSION: Presence of CFPHV DNA within globally distributed samples for all five species of sea turtle was confirmed. While 100% of the FP exhibiting green turtles yielded CFPHV sequences, surprisingly, so did 15% of the clinically healthy turtles. We hypothesize that turtle populations with zero (0%) CFPHV frequency may be attributed to possible environmental differences, diet and/or genetic resistance in these individuals. Our results provide first data on the prevalence of CFPHV among seemingly healthy turtles; a factor that may not be directly correlated to the disease incidence, but may suggest of a long-term co-evolutionary latent infection interaction between CFPHV and its turtle-host across species. Finally, computational analysis of amino acid variants within the Turks and Caicos samples suggest potential functional importance in a substitution for marker UL18 that encodes the major capsid protein gene, which potentially could explain differences in pathogenicity. Nevertheless, such a theory remains to be validated by further research.

Streptococcus equi subsp. zooepidemicus isolates from equine infectious endometritis belong to a distinct genetic group.

Streptococcus equi subsp. zooepidemicus is the pathogen most commonly isolated from the uterus of mares. S. zooepidemicus is an opportunistic pathogen and part of the resident flora in the caudal reproductive tract. The aim of this study was to investigate whether a genotypically distinct subpopulation of S. zooepidemicus is associated with endometritis in the mare, by genotyping and comparing uterine S. zooepidemicus strains with isolates from the vagina and clitoral fossa. Mares with (n=18) or without (n=11) clinical symptoms of endometritis were included. Uterine samples were obtained using a guarded endometrial biopsy punch, whereas a swab was used to recover samples from the cranial vagina and the clitoral fossa. If S. zooepidemicus was present, up to three colonies were selected from each anatomical location (max. 9 isolates per mare). Bacterial isolates were characterized by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). S. zooepidemicus was isolated from the endometrium of 12 mares. A total of 88 isolates were analyzed by PFGE: 31 from the endometrium, 26 from the cranial vagina and 31 isolates from the clitoral fossa. For MLST 21 isolates were chosen. Results demonstrated a higher genetic similarity of the isolates obtained from infectious endometritis compared to isolates obtained from the caudal reproductive tract. In conclusion, we demonstrate for the first time that a genetically distinct group of S. zooepidemicus is associated with infectious endometritis in the mare.

Bovine subclinical mastitis caused by Mannheimia granulomatis.

Mannheimia granulomatis was isolated for 10 months from the milk of a cow with elevated somatic cell counts. The infection was self-limiting. Phenotypic and molecular characteristics of the isolate were determined.