Effect of Temporary Meiotic Attenuation of Oocytes with Butyrolactone I and Roscovitine in Resistance to Bovine Embryos on Vitrification

This study aimed to produce in vitro bovine embryos by the addition of two drugs, which is responsible for oocyte meiosis inhibition: roscovitine (ROS) and butyrolactone I (BL‐I). Oocytes were recovered from slaughtered cows and matured in a commercial medium and maintained in a 5% CO₂ atmosphere. Oocytes were maintained for 6 h in an in vitro maturation (IVM) medium containing ROS (12.5 μm ), BL‐I (50 μm ) and association of drugs (ROS 6.25 μm and BL‐I 25 μm ). Oocytes were cultured for 18 h in an agent‐free medium for the resumption of meiosis. After 24 h of maturation, oocytes were inseminated in the commercial in vitro fertilization (IVF) medium. Presumptive zygotes were cultured in SOFaa medium in a 5% CO₂ atmosphere. On day 3, rate of cleavage was evaluated and on days 6 and 7, rate of blastocyst formation. BL‐I and its association with the ROS increased the rates of cleavage and blastocyst formation (p < 0.05). The ROS alone was inefficient, impairing embryonic development, with low rates of blastocyst formation when compared to the control group and other treatments (p < 0.05). The embryos from BL‐I and ROS+BL‐I groups presented higher number of cells and lower rates of cellular apoptosis compared to other groups, either for the fresh or for post‐thawing embryos. Embryos from ROS+BL‐I group showed to be more resistant to the vitrification process, presenting a higher rate of embryonic re‐expansion (p < 0.05). In conclusion, block of meiosis using BL‐I or its association with ROS increased the rate of blastocyst formation, and the association of ROS+BL‐I resulted in a better resistance to the embryo cryopreservation process.     

Herd and within-herd BoHV-1 prevalence among Irish beef herds submitting bulls for entry to a performance testing station

Infectious bovine rhinotracheitis (IBR), caused by bovine herpes virus 1 (BoHV-1), may result in various clinical consequences, including severe respiratory disease and conjunctivitis, venereal disease and reduced reproductive performance and abortion. This paper presents the serosurveillance findings from an intake of bulls into a performance testing station in Ireland during November 2007. The herd and within-herd BoHV-1 prevalence in 53 Irish beef herds and the risk factors for infection in these herds were determined, among bulls entering a beef performance testing station in Ireland. BoHV-1 status was determined for 41 herds, of which 30 (73.2%) herds were infected and the mean within-herd BoHV-1 prevalence was 28 (± 20)%. Multivariate exact logistic modelling revealed increasing numbers of contiguous herds and decreasing percentage of males within the herd as significant risk factors associated with infected herds. These findings highlight the high prevalence of BoHV-1 infection in those Irish beef herds that submitted bulls to this performance testing station, and raise concerns regarding IBR control nationally.     

Emergence of pathogenic betanodaviruses belonging to the SJNNV genogroup in farmed fish species from the Iberian Peninsula

Betanodaviruses are the causative agents of viral nervous necrosis (VNN) or viral encephalopathy and retinopathy (VER) in cultured marine fish. Based on the RNA2 gene fish nodaviruses have been traditionally classified into four different genotypes and recently a fifth genotype has been proposed. This study presents sequencing data of 24 new nodaviruses obtained from three different fish species: sea bass, Dicentrarchux labrax (L.), sea bream, Sparus aurata L., and Senegalese sole, Solea senegalensis Kaup, cultured in the Iberian Peninsula (Spain and Portugal). Sequence analysis was performed on the T4 region (388 nt) of the coat protein gene. In addition, phylogenetic analysis, according to maximum parsimony and neighbour-joining methods, was performed using these sequences and other nucleotide sequences available in the databases or in the literature. Results obtained indicate that all these new nodaviruses should be classified into the striped jack nervous necrosis virus (SJNNV) genotype. This finding suggests that SJNNV genotype is emerging in the Iberian Peninsula and could easily spread throughout the Mediterranean, representing a serious threat to the fish farming industry.     

Endometrial Explant Culture to Study the Response of Equine Endometrium to Insemination

Mating‐induced endometritis (MIE) is ubiquitous in the horse after natural mating and artificial insemination with frozen/thawed semen causing the most aggressive response. The majority of mares eliminate MIE 24–48 h after insemination. An endometrial explant culture was tested as a potential in vitro exemplar for sperm‐induced MIE. Endometrial prostaglandin F_2α (PGF_2α) secretion and expression of interleukin‐8 (IL‐8) were used as markers of inflammation. Endometrial explants were cultured from uteri collected from follicular phase mares. Explants were challenged with 1 or 10 × 10⁶ sperm/ml frozen/thawed semen, chilled semen, washed sperm or seminal plasma. Medium was collected 24 and 72 h after challenge and assayed for PGF_2α by radioimmunoassay. Treatment of endometrial explants with frozen/thawed, chilled semen or washed sperm did not change the secretion of PGF_2α compared with untreated controls. However, 24 h after challenge cultured explants expressed IL‐8. The in vitro endometrial explant system did not represent the in vivo response to semen when PGF_2α was used as a marker of inflammation, yet the use of gene expression as an inflammatory marker warrants further investigation.     

New method for the determination of benzoic and sorbic acids in commercial orange juices based on second-order spectrophotometric data generated by a pH gradient flow injection technique

Two widely employed antimicrobials, benzoic and sorbic acids, were simultaneously determined in commercial orange juices employing a combination of a flow injection system with pH gradient generation, diode array spectrophotometric detection, and chemometric processing of the recorded second-order data. Parallel factor analysis and multivariate curve resolution-alternating least-squares were used for obtaining the spectral profiles of sample components and concentration profiles as a function of pH, including provisions for managing rank-deficient data sets. An appropriately designed calibration with a nine-sample set of binary mixtures of standards, coupled to the use of the second-order advantage offered by the applied chemometric techniques, allowed quantitation of the analytes in synthetic test samples and also in commercial orange juices, even in the presence of unmodeled interferents (with relative prediction errors of 8.7% for benzoic acid and 2.5% for sorbic acid). No prior separation or sample pretreatment steps were required. The comparison of results concerning commercial samples with a laborious reference technique yielded satisfactory statistical indicators (recoveries were 99.0% for benzoic acid and 101.4% for sorbic acid).     

Presence and diversity of mixed avian Plasmodium spp. infections in introduced birds whose distribution overlapped with threatened New Zealand endemic birds

ABSTRACT

Aims: To determine the presence of infection and co-infection of Plasmodium lineages in introduced birds at translocation sites for the North Island saddleback (Philesturnus rufusater), to investigate their role as Plasmodium spp. reservoirs.

Methods: Blood samples were collected from introduced bird species, with a special focus on blackbirds (Turdus merula) and song thrushes (Turdus philomelos), at six locations in the North Island of New Zealand that were the origin, or translocation sites, for North Island saddleback. Where available, blood smears were examined, and blood samples were tested using nested PCR with subsequent sequence analysis, for the presence of Plasmodium spp.

Results: Of the 55 samples tested using PCR analysis, 39 (71%) were positive for Plasmodium spp., and 28/40 (62%) blood smears were positive for Plasmodium spp. Overall, 31 blood samples were from blackbirds with 28/31 (90%) samples positive for Plasmodium spp. Six distinct avian Plasmodium lineages were identified, including three cosmopolitan lineages; Plasmodium vaughani SYAT05 was detected in 16 samples, Plasmodium matutinum Linn1 in 10 samples and Plasmodium elongatum GRW6 in eight samples. Mixed infections with more than one lineage were detected in 12 samples. Samples from two Australian magpies (Gymnorhina tibicen) were positive for Plasmodium. sp. lineage MYNA02, previously not identified in New Zealand.

Conclusions and clinical relevance: This is the first report from New Zealand in which specific Plasmodium spp. mixed infections have been found in introduced birds. Co-infections with several cosmopolitan Plasmodium lineages were identified, as well as the first report in New Zealand of an exotic avian Plasmodium sp. lineage, in Australian magpies. Whilst the role of introduced birds in maintaining and spreading pathogenic avian malaria in New Zealand is unclear, there is a potential infection risk to native birds, especially where distributions overlap.     

Experimental infection of turbot, Psetta maxima (L.), with strains of viral haemorrhagic septicaemia virus isolated from wild and farmed marine fish

The susceptibility of turbot, Psetta maxima, to infection with two strains of viral haemorrhagic septicaemia virus (VHSV) obtained from wild Greenland halibut, Reinhardtius hippoglossoides, and from farmed turbot was examined. A marine VHSV strain known to be highly pathogenic for turbot was also utilized for comparative purposes. Fish were infected by intra-peritoneal (i.p.), immersion or cohabitation, and maintained at two different temperatures (8 and 15 degrees C). Infection trials showed that the three VHSV isolates were pathogenic for turbot fingerlings by i.p. injection at both temperatures, with high levels of mortality. Virus was recovered from most pools of dead fish i.p. challenged, but not from surviving fish. Although clinical signs were not induced following waterborne exposure, viral growth was obtained from some pools of surviving fish challenged by immersion with strain GH40 from Greenland halibut, which indicates that the virus can survive in sea water and infect other fish via horizontal transmission. Furthermore, although low, the clinical signs and mortality observed in fish cohabitating with turbot challenged with strain GH40 confirms horizontal transmission and indicates that the passage through fish increases the virulence of this strain for turbot. These findings indicate that Greenland halibut, as other wild fish, may play an important role in the epizootiology of VHSV and suggest a potential risk for the turbot farming industry.