Comparative Aspects of the Endotoxin- and Cytokine-Induced Endocrine Cascade Influencing Neuroendocrine Control of Growth and Reproduction in Farm Animals

Disease in animals is a well-known inhibitor of growth and reproduction. Earlier studies were initiated to determine the effects of endotoxin on pituitary hormone secretion. These studies found that in sheep, growth hormone (GH) concentration was elevated, whereas insulin-like growth factor-I (IGF-I) was inhibited, as was luteinizing hormone (LH). Examination of the site of action of endotoxin in sheep determined that somatotropes expressed the endotoxin receptor (CD14) and that both endotoxin and interleukin-Iβ activated GH secretion directly from the pituitary. In the face of elevated GH, there is a reduction of IGF-I in all species examined. As GH cannot activate IGF-I release during disease, there appears to be a downregulation of GH signalling at the liver, perhaps related to altered nitration of Janus kinase (JAK). In contrast to GH downregulation, LH release is inhibited at the level of the hypothalamus. New insights have been gained in determining the mechanisms by which disease perturbs growth and reproduction, particularly with regard to nitration of critical control pathways, with this perhaps serving as a novel mechanism central to lipopolysaccharide suppression of all signalling pathways. This pathway-based analysis is critical to the developing novel strategies to reverse the detrimental effect of disease on animal production.     

Identification of putative orthologs of clinically relevant antimicrobial peptides in the equine ocular surface and amniotic membrane

Objectives

This study aimed to define the antimicrobial peptide (AMP) expression pattern of the equine ocular surface and amniotic membrane using a targeted qPCR approach and 3'Tag-sequencing. It will serve as a reference for future studies of ocular surface innate immunity and amniotic membrane therapies.

Procedures

A targeted qPCR approach was used to investigate the presence of orthologs for three of the most highly expressed beta-defensins (DEFB1, DEFB4B, and DEFB103A) of the human ocular surface and amniotic membrane in equine corneal epithelium, conjunctiva, and amniotic membrane. 3'Tag-sequencing was performed on RNA from one sample of corneal epithelium, conjunctiva, and amniotic membrane to further characterize their AMP expression.

Results

Equine corneal epithelium, conjunctiva, and amniotic membrane expressed DEFB1, DEFB4B, and DEFB103A. DEFB103A was expressed at the highest amounts in corneal epithelium, while DEFB4B was most highly expressed in conjunctiva and amniotic membrane. 3'Tag-sequencing from all three tissues confirmed these findings and identified expression of five additional beta-defensins, 11 alpha-defensins and two cathelicidins, with the alpha-defensins showing higher normalized read counts than the beta-defensins.

Conclusions

This study identified AMP expression in the equine cornea and conjunctiva, suggesting that they play a key role in the protection of the equine eye, similar to the human ocular surface. We also determined that equine amniotic membrane expresses a substantial number of AMPs suggesting it could potentiate an antimicrobial effect as a corneal graft material. Future studies will focus on defining the antimicrobial activity of these AMPs and determining their role in microbial keratitis.     

Antimicrobial resistance in commensal faecal Escherichia coli of hospitalised horses

The objective of this study was to examine the impact of hospitalisation and antimicrobial drug administration on the prevalence of resistance in commensal faecal E. coli of horses. Faecal samples were collected from ten hospitalised horses treated with antimicrobials, ten hospitalised horses not treated with antimicrobials and nine non-hospitalised horses over a consecutive five day period and susceptibility testing was performed on isolated E. coli. Results revealed that hospitalisation alone was associated with increased prevalence of antimicrobial resistance and multidrug resistance in commensal E. coli of horses. Due to the risk of transfer of resistance between commensal and pathogenic bacteria, veterinarians need to be aware of possible resistance in commensal bacteria when treating hospitalised horses.     

Serum Starvation and Full Confluency for Cell Cycle Synchronization of Domestic Cat (Felis catus) Foetal Fibroblasts

Nuclear transfer of domestic cat can be used as a tool to develop reproductive biotechnologies in wild felids. The importance of cell cycle phase during the nuclear transfer has been a matter of debate since the first mammalian clone was produced. The cell cycle phase of donor cells interferes on maintenance of correct ploidy and genetic reprogramming of the reconstructed embryo. The use of G0/G1 arrested donor cells has been shown to improve nuclear transfer efficiency. The present study was conducted to test the hypothesis that domestic cat foetal fibroblasts cultured up to the fifth passage and submitted to full confluency provide a higher percentage of cells at G0/G1 stage than fibroblasts cultured in serum starved media. Results demonstrated that serum starvation increased (p ≤ 0.05) the percentage of G0/G1 fibroblasts when compared with control. Moreover, the combined protocol using confluency and serum starvation was more efficient (p ≤ 0.05) synchronizing cells at G0/G1 stage than serum starvation or confluency alone for the first 3 days of treatment. In conclusion, serum starvation and full confluency act in a synergistic manner to improve domestic cat foetal fibroblast cell cycle synchronization at the G0/G1 stage.     

Significance of Non‐conventional Parameters in the Evaluation of Cooling‐induced Damage to Ram Spermatozoa Diluted in Three Different Media

The objectives of this study were two. First, to compare three base media with different sugar composition as an initial step to achieve a good chemically‐defined extender for ram sperm refrigeration. The second one, to determine which sperm quality parameters may be more useful for revealing differences between sperm samples. One medium contained 200 mm sucrose and 2.8 mm glucose (SM), another only disaccharides (D) such as sucrose, trehalose, maltose and lactose (75 mm each); and the third one (D+M) included a mix of monosaccharides (50 mm glucose, 20 mm fructose and 20 mm galactose,) and the same disaccharides as in D (50 mm each). Ram semen samples diluted in the mentioned media were refrigerated at 5°C for 1 h, and rewarmed upto 37°C in order to mimic the temperature in the female reproductive tract. Addition of monosaccharides to the extender did not produce a better preservation of motility or viability after cooling. The supplementation with other disaccharides apart from sucrose did not enhance the viability either. Thus, after cooling and rewarming, there were no significant differences in sperm viability (membrane integrity evaluated by CFDA/PI staining) or the percentage of progressive motile and rapid sperm (evaluated by CASA) between the three media. However, the percentage of viable non‐capacitated sperm evaluated by the chlortetracycline (CTC) assay was higher and sperm oxygen consumption was lower in SM than in D and in D+M. Although the apoptosis‐like markers [phosphatidylserine exposure assessed by Annexin V/CFDA staining and DNA‐damage evaluated by TUNEL assay] showed a continuous increment throughout the process with all diluents, the percentage of sperm with damaged DNA at the end of the process was significantly lower in SM than in the other two media (p < 0.01). On the basis of these results, we would make two recommendations: the use of an extender supplemented only with sucrose and glucose for ram sperm refrigeration; the inclusion of non‐conventional methods such as oxygen consumption measure, evaluation of capacitation state and apoptosis‐like markers for revealing differences between sperm samples.     

Epidemic of blindness in kangaroos - evidence of a viral aetiology

OBJECTIVE: To determine the cause of an epidemic of blindness in kangaroos. DESIGN AND PROCEDURES: Laboratory examinations were made of eyes and brains of a large number of kangaroos using serological, virological, histopathological, electron microscopical, immunohistochemical methods, and PCR with cDNA sequencing. In addition, potential insect viral vectors identified during the disease outbreak were examined for specific viral genomic sequences. SAMPLE POPULATION: For histopathological analysis, 55 apparently blind and 18 apparently normal wild kangaroos and wallabies were obtained from New South Wales, Victoria, South Australia, and Western Australia. A total of 437 wild kangaroos and wallabies (including 23 animals with apparent blindness) were examined serologically. RESULTS: Orbiviruses of the Wallal and Warrego serogroups were isolated from kangaroos affected with blindness in a major epidemic in south-eastern Australia in 1994 and 1995 and extending to Western Australia in 1995/96. Histopathological examinations showed severe degeneration and inflammation in the eyes, and mild inflammation in the brains. In affected retinas, Wallal virus antigen was detected by immunohistochemical analysis and orbiviruses were seen in electron microscopy. There was serological variation in the newly isolated Wallal virus from archival Wallal virus that had been isolated in northern Australia. There were also variations of up to 20% in genotype sequence from the reference archival virus. Polymerase chain reactions showed that Wallal virus was present during the epidemic in three species of midges, Culicoides austropalpalis, C dycei and C marksi. Wallal virus nucleic acid was also detected by PCR in a paraffin-embedded retina taken from a blind kangaroo in 1975. CONCLUSION: Wallal virus and perhaps also Warrego virus are the cause of the outbreak of blindness in kangaroos. Other viruses may also be involved, but the evidence in this paper indicates a variant of Wallal virus, an orbivirus transmitted by midges, has the strongest aetiological association, and immunohistochemical analysis implicates it as the most damaging factor in the affected eyes.     

Foraging under uniform risk from different types of predators

Background  

Many animals live in environments where different types of predators pose a permanent threat and call for predator specific strategies. When foraging, animals have to balance the competing needs of food and safety in order to survive. While animals sometimes can choose between microhabitats that differ in their risk of predation, many habitats are uniform in their risk distribution. So far, little is known about adaptive antipredator behavior under uniform risk. We simulated two predator types, avian and mammalian, each representing a spatially uniform risk in the artificial resource landscapes. Voles served as experimental foragers.     

Resistance risk evaluation, “a European regulatory perspective”

The European Union directive 91/414 on the inclusion of new and existing active substances requires an assessment of the potential risk of resistance being developed by target organisms and for management strategies to be introduced to address such risks. The European and Mediterranean Plant Protection Organisation (EPPO) worked with regulatory authorities and industry's Global Crop Protection Federation's (GCPF) Resistance Action Committees to develop a guideline that would satisfy these requirements. The new resistance risk analysis guideline was published in April 2000. Implementation of this guideline will force significant changes in the way crop protection products are developed. Applicants for registration of crop protection products will need to evaluate resistance risk and develop acceptable use patterns early in the development process. Registration applications for uses that are associated with an unacceptable resistance risk may be refused. In such cases, restriction or modification of supported use patterns to achieve acceptable levels of resistance risk, could result in use patterns which are no longer consistent with other elements of the registration dossier. It is therefore essential that resistance risk is evaluated early in development so that registration dossiers may be developed with this criteria in mind.     

Digestibility and transit time of Azolla filiculoides Lamarck in Oreochromis aureus (Steindachner)

Azolla, an aquatic fern that is able to fix nitrogen directly from the atmosphere, could represent an interesting low-cost food for tropical fish. In the present study, the authors evaluated the apparent digestibility of Azolla filiculoides Lamarck by Oreochromis aureus (Steindachner), an omnivorous-phytoplankton-ophagous species of tilapia. Two digestibility tests (5 days) were performed in digestibility tanks on adult O. aureus fed only with dried A. filiculoides given ad libitum (2% biomass day^?1). Experiment 1 was made on fish weighing 28-49 g (duplicate) at a water temperature of 29 ± 0.5°C; experiment 2 was made on fish weighing 36-81 g (duplicate) at a water temperature of 27 ± 0.5°C. The transit time in O. aureus (weight: 30-50 g; water temperature: 27 ± 0.5°C) fed only and ad libitum with A. filiculoides was determined with methylene blue as a vital dye. The apparent faecal digestibility coefficients (%) obtained for dry matter and crude protein were 49.0 and 55.4%, respectively, in experiment 1, and 53.5 and 60.9% in experiment 2. The transit time values of 2.5-3.0 h and 3.0-3.5 h were obtained, respectively, for fresh and dried A. filiculoides. Thus, A. filiculoides shows relatively good digestibility values in O. aureus, which are comparable to those obtained with other aquatic plants in tropical fish. However, in order to meet the nutrient requirements of tilapias, Azolla has to be mixed with other foods.