Prevalence of cpb2, encoding beta2 toxin,in Clostridium perfringens field isolates: correlation of genotype with phenotype

Beta2 toxin, encoded by the cpb2 gene, has been implicated in the pathogenesis of porcine, equine and bovine enteritis by type A Clostridium perfringens. By incorporating primers to cpb2 into a multiplex genotyping PCR, we screened 3270 field isolates of C. perfringens. Of these, 37.2% were PCR positive for the cpb2 gene. The majority of isolates from cases of porcine enteritis were positive for cpb2 (>85%), and this was even more true for C. perfringens isolated from cases of porcine neonatal enteritis (91.8%). In contrast, isolates from normal pigs only contained cpb2 in 11.1% of cases. The correlation between enteritis in other animal species and the presence of cpb2 was not so strong. cpb2 was found in 21.4% of C. perfringens isolates from cattle with enteritis, and in 47.3% of isolates from calves with enteritis or abomastitis. The prevalence of cpb2 varied with genotype, with type A isolates being positive for this gene in 35.1% of cases. Furthermore, enterotoxigenic type D or type E strains almost always carried cpb2. We cloned a 6xHIS-tagged beta2 (HIS-beta2) and used this protein to raise antiserum against beta2. Culture supernatants from 68 cpb2-positive and 13 cpb2-negative strains were tested for the presence of beta2 by Western blotting. In cpb2-positive isolates of porcine origin, beta2 was almost always detected (96.9%). However, in cpb2-positive isolates from other animal species, only 50.0% expressed beta2 protein. The high rate of cpb2-positivity among strains from neonatal pigs with enteritis and the high correlation of genotype with phenotype, supports the contention that beta2 toxin plays a role in the pathogenesis of these infections. However, it may be important to consider the use of an additional method for the detection of beta2 toxin in non-porcine cpb2-positive isolates when making claims about the role of beta2 in enteritis in non-porcine species.     

A prospective, case control study evaluating the association between Clostridium difficile toxins in the colon of neonatal swine and gross and microscopic lesions.

Clostridium difficile infection in swine has most often been described in suckling pigs, where it has been associated with mesocolonic edema and typhlocolitis. This prospective study was designed to assess the correlation between the presence of C. difficile toxins (TCd) in the colon contents of neonatal pigs and a number of parameters, including gross evidence of diarrhea, mesocoloninc edema, typhlitis, and colitis. C. difficile was isolated from 51% (66/129) of large intestines and TCd was detected in the colon contents of 50% (65/129) of the piglets. Fifty-eight percent (38/65) of TCd-positive piglets had normal to pelleted colon and rectal contents, whereas 75% (48/64) of TCd-negative pigs had gross evidence of diarrhea. Clostridium difficile toxin-positive animals were significantly more likely to have normal to pelleted feces. Edema of the mesocolon was observed in 38/65 (59%) of TCd-positive piglets. Because a high number of TCd-positive piglets (41%) lacked edema of the mesocolon and a high number of TCd-negative pigs had mesocolonic edema (51%), a statistically significant association between TCd and mesocolonic edema was not identified. Seventy-five percent (49/65) of TCd-positive piglets had colitis and 47/65 (72%) had typhlitis. The association between TCd and both colitis and typhlitis was statistically significant. Apparently healthy piglets were obtained from 5 separate sites. Because TCd was detected in the colon contents of 23/29 (79%) apparently healthy piglets obtained from 5 separate sites, and 70% of TCd-positive control pigs had colitis, C. difficile may represent an important subclinical issue in neonatal swine.     

Analysis of FMO genes and off flavour in pork

Off flavours in pork sometimes produce tastes such as sourness, fishy, metallic or other non‐typical flavours and are often caused by low pH. Loss of function mutations in flavin containing mono‐oxygenase 3 (FMO3) are known to be associated with a fishy off flavour in both chicken eggs and cow's milk and a similar autosomal recessive disorder is present in humans resulting in a fishy odour. FMO3 is a member of a gene family that is clustered on human chromosome 1. Comparative mapping suggested that FMO3 and the remaining FMO genes (ex. FMO1 and FMO5) might map to the orthologous region on pig chromosome 9 (SSC9) where a quantitative trait locus (QTL) for off flavour was previously identified. Primers were designed to amplify FMO1, FMO3 and FMO5 gene fragments and several SNPs were discovered and genotyping tests developed. The genotypes from the Iowa State University Berkshire × Yorkshire resource population were used to linkage map FMO1 and FMO3 to SSC9 and FMO5 to pig chromosome 4 (SSC4). QTL and associations analyses were performed using the map containing FMO1 and FMO3. Results demonstrated that FMO3 and FMO1 mapped less than 1 cM away from the peak for the off flavour QTL previously detected on SSC9 and provide indications of an association between the FMO3 polymorphism and off flavour in pork.     

The arrhythmogenic dose of epinephrine in halothane and isoflurane anesthetized dogs: an assessment of repeatability.

Repeat determinations of the arrhythmogenic dose of epinephrine (ADE) were made over two 6 h periods on 2 separate days during halothane and isoflurane anesthesia. Each of 6 dogs underwent 4 trials (2 halothane and 2 isoflurane). During each trial, the ADE was determined at baseline, 3 and 6 h. Epinephrine was infused for 3.0 min at increasing dose rates (2.5, 5.0, 10.0 and 20.0 mg/kg/min) until the arrhythmia criterion (4 or more intermittent or continuous premature ventricular contractions) was reached. The inter-infusion interval was 20 min. There were no significant differences in the measured cardiovascular parameters (SBP, DBP, MBP, and HR), arterial blood gases, or acid-base status prior to each determination during a single trial. The cardiovascular responses to epinephrine infusion were not significantly different between inhalants or determinations. The range of the ADE determined over both trials during isoflurane anesthesia was 30.12 +/- 12.21 micrograms/kg to 50.83 +/- 9.17 micrograms/kg. The baseline ADE during Day 1 of halothane anesthesia (6.70 +/- 1.36 micrograms/kg) was significantly greater than ADE determinations at 3 (4.65 +/- 0.88 micrograms/kg) and 6 h (4.61 +/- 0.87 micrograms/kg). The reduction in the ADE over time during day 2 of halothane anesthesia was not statistically significant (P = 0.0669). These results suggest that during halothane anesthesia, the ADE is not repeatable over time, and they may influence our interpretation of the results of investigations that measure alterations in the ADE due to pharmacological manipulations without repeated control ADE determinations.     

Environmental impact mitigation and bi-culture: a comparative legal analysis of flexibility within European legal regimes – biofilter deployment

This paper investigates the potential flexibility within the current legal frameworks governing marine aquaculture across Europe to cater for the installation and management of marine biofilters alongside fish farms. The basis for debate is that deployment of these biofilters could be used to facilitate environmental impact mitigation. Furthermore, they have the potential to facilitate the development of bi-culture or polyculture through the harvesting of species from the biofilters themselves, for example, mussels (Mytilus edulis). This study explores the flexibility within and suitability of the current legal systems within Europe to cater for the specific technology’s adoption and management. The challenges identified from the legislative and policy frameworks are discussed along with the applicability of these to the use of biofilters for the development of bi-culture. Penultimately, recommendations are made as to where additional measures are needed and the challenges and difficulties that such measures will need to overcome before biofilters can be applied for environmental impact mitigation. Finally, the paper illustrates an application of ‘Comparative Legal Analysis’, demonstrating links to the question in hand and through so doing to wider coastal issues.     

Population trends in northern spotted owls: Associations with climate in the Pacific Northwest

We used reverse time capture-mark-recapture models to describe associations between rate of population change (λ) and climate for northern spotted owls (Strix occidentalis caurina) at six long-term study areas in Washington and Oregon, USA. Populations in three of six areas showed strong evidence of declining populations, while populations in two additional areas were likely declining as well. At four areas, λ was positively associated with wetter-than-normal conditions during the growing season, which likely affects prey availability. Lambda was also negatively associated with cold, wet winters and nesting seasons, and the number of hot summer days. The amount of annual variation in λ accounted for by climate varied across study areas (3-85%). Rate of population change was more sensitive to adult survival than to recruitment; however, there was considerable variation among years and across study areas for all demographic rates. While annual survival was more closely related to regional climate conditions, recruitment was often associated with local weather. In addition to climate, declines in recruitment at four of six areas were associated with increased presence of barred owls. Climate change models predict warmer, wetter winters and hotter, drier summers for the Pacific Northwest in the first half of the 21st century. Our results indicate that these conditions have the potential to negatively affect annual survival, recruitment, and consequently population growth rates for northern spotted owls.     

Intra-specific variation of wheat grain quality in response to elevated [CO2] at two sowing times under rain-fed and irrigation treatments

In order to investigate the intra-specific variation of wheat grain quality response to elevated atmospheric CO₂ concentration (e[CO₂]), eight wheat (Triticum aestivum L.)cultivars were grown at two CO₂ concentrations ([CO₂]) (current atmospheric, 389 CO₂ μmol mol⁻¹vs. e[CO₂], FACE (Free-Air CO₂ Enrichment), 550  ±  10% CO₂ μmol mol⁻¹), at two water levels (rain-fed vs. irrigated) and at two times of sowing (TOS₁, vs. TOS₂). The TOS treatment was mainly imposed to understand whether e[CO₂] could modify the effects of timing of higher grain filling temperatures on grain quality. When plants were grown at TOS₁, TKW (thousand kernel weight), grain test weight, hardness index, P, Ca, Na and phytate were not significantly changed under e[CO₂]. On the other hand, e[CO₂] increased TKW (16%), hardness index (9%), kernel diameter (6%), test weight (2%) but decreased grain protein (10%) and grain phytate (11%) at TOS₂. In regard to grain Zn, Mn and Cu concentrations and some flour rheological properties, cultivar specific responses to e[CO₂] were observed at both sowing times. Observed genetic variability in response to e[CO₂] in terms of grain minerals and flour rheological properties could be easily incorporated into future wheat breeding programs to enable adaptation to climate change.