Gene discovery in Eimeria tenella by immunoscreening cDNA expression libraries of sporozoites and schizonts with chicken intestinal antibodies

Specific antibodies were produced ex vivo from intestinal culture of Eimeria tenella infected chickens. The specificity of these intestinal antibodies was tested against different parasite stages. These antibodies were used to immunoscreen first generation schizont and sporozoite cDNA libraries permitting the identification of new E. tenella antigens. We obtained a total of 119 cDNA clones which were subjected to sequence analysis. The sequences coding for the proteins inducing local immune responses were compared with nucleotide or protein databases and with expressed sequence tags (ESTs) databases. We identified new Eimeria genes coding for heat shock proteins, a ribosomal protein, a pyruvate kinase and a pyridoxine kinase. Specific features of other sequences are discussed.     

Detection of Cryptosporidium oocysts in goat kid faeces: comparison of a latex agglutination test with three other conventional techniques

Detection of Cryptosporidium oocysts from goat kid faeces: comparison of a latex agglutination test with three other conventional techniques. A quantitative latex agglutination test (QLAT) with monoclonal antibodies for the detection of Cryptosporidium oocysts in faeces was compared with 3 other conventional techniques: Heine staining on faecal smears (HS) giving semi-quantitative results (scores from 1 to 5), sucrose flotation on diluted faeces (SF) with results expressed in oocysts/g of faeces (opg), direct ELISA (DE) giving qualitative results. Goat kid unconcentrated faecal samples (234) from 8 farms were processed according to the 4 techniques. Data were analyzed with Win Episcope 1.0 and Testview 1.1 softwares. The oocyst outputs ranged from 100 000 (detection limit for SF) to 200 millions opg (mean: 15.2 millions opg). A very good agreement was recorded between QLAT and HS, SF, DE: Kappa values ranged between 0.82 and 0.90. When considering the samples exhibiting oocysts (or not) as positive (or negative) using both HS and SF (n = 219), the sensitivity and specificity of QLAT were respectively 95.1 and 96.0%. The lack of sensitivity was observed in faeces harboring a few oocysts (< or = 200 000 opg, scores < or = 2) whereas the lack of specificity was only observed in 3 samples originating from the same farm. A significant correlation was calculated between the percentage of agglutination in QLAT and the number of oocysts in SF or scores in HS (Spearman correlation ranging from 0.45 to 0.48, p < 0.001). QLAT is a rapid, simple and reliable tool for routine detection of Cryptosporidium oocysts in faeces.     

Serologic and genetic characterization of North American H3N2 swine influenza A viruses

The H3N2 subtype of influenza A viruses isolated from pigs in the United States and Canada has shown both genetic and antigenic diversity. The objective of this study was to determine the serologic and genetic characteristics of contemporary strains of these viruses. Genetic analysis of 18 reference strains and 8 selected strains demonstrated differences in 1% to 9% of the nucleotides of the hemagglutinin (HA) gene. Phylogenetic analysis of the HA gene revealed 3 genetic clusters, as well as divergence of cluster III viruses from a cluster III prototype virus (A/Swine/Illinois/21587/99). By means of 1-way cross-hemagglutination inhibition with antiserum against 5 field isolates and 3 vaccine viruses, most of 97 isolates tested could be placed in 1 of 3 serogroups. The several isolates that did not react with any antiserum were in genetic cluster III, which suggests that continuous antigenic drift in cluster III may have resulted in virus variants. The efficacy of commercial vaccines against these virus variants should be evaluated with vaccination and challenge studies.     

Protocol: a highly sensitive RT-PCR method for detection and quantification of microRNAs

MicroRNAs (miRNAs) are a class of small non-coding RNAs with a critical role in development and environmental responses. Efficient and reliable detection of miRNAs is an essential step towards understanding their roles in specific cells and tissues. However, gel-based assays currently used to detect miRNAs are very limited in terms of throughput, sensitivity and specificity. Here we provide protocols for detection and quantification of miRNAs by RT-PCR. We describe an end-point and real-time looped RT-PCR procedure and demonstrate detection of miRNAs from as little as 20 pg of plant tissue total RNA and from total RNA isolated from as little as 0.1 μl of phloem sap. In addition, we have developed an alternative real-time PCR assay that can further improve specificity when detecting low abundant miRNAs. Using this assay, we have demonstrated that miRNAs are differentially expressed in the phloem sap and the surrounding vascular tissue. This method enables fast, sensitive and specific miRNA expression profiling and is suitable for facilitation of high-throughput detection and quantification of miRNA expression.     

Mild to moderate overweight in dogs: is there an impact on routine hematological and biochemical profiles,echocardiographic parameters and cardiac autonomic modulation?

Veterinary Research Communications - Obesity is considered the most common nutritional disease of dogs. Even though overt obesity is more likely to impair health, even moderately overweight dogs...     

Allometric model of the Reineke equation for Japanese cypress (<Emphasis Type="Italic">Chamaecyparis obtusa</Emphasis>) and red pine (<Emphasis Type="Italic">Pinus densiflora</Emphasis>) stands

An allometric model that explains the mechanism of the difference in the slope of the Reineke equation (A) among species was proposed based on the allometric relationships of mean tree height (H) to quadratic mean diameter D (H ∝ D ^θ ) and stand density N (H ∝ N ^δ ), i.e., A = θ/δ. The proposed model was fitted to Japanese cypress (Chamaecyparis obtusa Endl.) and red pine (Pinus densiflora) stands. The allometric exponents θ and δ were, respectively, 0.8995 and −0.5000 for cypress and 0.8612 and −0.6619 for pine. The difference between cypress and pine was significant for δ but not for θ. Inserting the exponents into the model resulted in predicted slopes of −1.7991 for cypress and −1.3011 for pine. The difference in the slope of the Reineke equation between the two species was produced by characteristics related to the tree crown, rather than characteristics related to stem slenderness. The proposed model enables us to estimate the slope of the Reineke equation from commonly measured stand attributes, such as mean tree height and quadratic mean diameter. Therefore, the proposed model is expected to be practical and convenient for estimating the slope of the Reineke equation and for explaining the mechanism of its variation among species. The model should be also accepted as a generalized model of the stand density versus quadratic mean diameter relationship, whereas the original Reineke equation should be seen as a specific case of this model.     

Antibacterial and antioxidant activities of Artemisia annua essential oil

The essential oil of Artemisia annua aerial parts, consisting of camphor (44%), germacrene D (16%), trans-pinocarveol (11%), beta-selinene (9%), beta-caryophyllene (9%) and artemisia ketone (3%), was screened for its antimicrobial activity. The essential oil remarkably inhibited the growth of tested Gram-positive bacteria Enterococcus hirae and both tested fungi. This oil has shown an antioxidant activity equivalent to 18% of the reference compound (alpha-tocopherol).     

Effects of two different anesthetic protocols on 64‐MDCT coronary angiography in dogs

Heart rate is a major factor influencing diagnostic image quality in computed tomographic coronary artery angiography (MDCT‐CA), with an ideal heart rate of 60–65 beats/min in humans. The purpose of this prospective study was to compare effects of two different clinically applicable anesthetic protocols on cardiovascular parameters and 64‐MDCT‐CA quality in 10 healthy dogs. Scan protocols and bolus volumes were standardized. Image evaluations were performed in random order by a board‐certified veterinary radiologist who was unaware of anesthetic protocols used. Heart rate during image acquisition did not differ between protocols (P = 1), with 80.6 ± 7.5 bpm for protocol A and 79.2 ± 14.2 bpm for protocol B. Mean blood pressure was significantly higher (P > 0.05) using protocol B (protocol A 62.9 ± 9.1 vs. protocol B 72.4 ± 15.9 mmHg). The R‐R intervals allowing for best depiction of individual coronary artery segments were found in the end diastolic period and varied between the 70% and 95% interval. Diagnostic quality was rated excellent, good, and moderate in the majority of the segments evaluated, with higher scores given for more proximal segments and lower for more distal segments, respectively. Blur was the most commonly observed artifact and mainly affected the distal segments. No significant differences were identified between the two protocols for optimal reconstruction interval, diagnostic quality and measured length individual segments, or proximal diameter of the coronary arteries (P = 1). Findings indicated that, when used with a standardized bolus volume, both of these anesthetic protocols yielded diagnostic quality coronary 64‐MDCT‐CA exams in healthy dogs.     

Lumbo-sacral epidural anaesthesia as a complement to dissociative anaesthesia during scrotal herniorrhaphy of livestock pigs in the field

Background

In Sweden, scrotal or inguinal herniorrhaphy of livestock pigs in the field has traditionally been an important part of the surgical skills training of veterinary students. Few substances meet the legal requirements for field anaesthesia of production animals in the European Union but a protocol based on azaperone-detomidine-butorphanol-ketamine does. Unfortunately the anaesthesia is characterised by unpredictable duration and depth and of abrupt awakenings which is not acceptable from an animal welfare perspective and impedes surgical training. Lumbo-sacral epidural analgesia is proven to provide sufficient analgesia to allow abdominal surgery, but there are few reports on the field use of this loco-regional technique. The study aim was to evaluate whether lumbo-sacral anaesthesia can be safely and successfully used in the field by a veterinary student and whether the combination of dissociative and lumbo-sacral epidural anaesthesia improves analgesia and anaesthesia to guarantee animal welfare during herniorrhaphy in livestock pigs, enabling surgical skills training.

Results

Pigs in the control-group (placebo) responded significantly stronger to surgery, with five out of 11 requiring additional doses of detomidine and ketamine. There were no significant differences between groups in respiratory rate, heart rate, blood pressure, SpO₂ or blood gases. SpO₂ levels <94 % were recorded in several pigs in both groups. No post-injection complications were reported at follow-up.

Conclusions

The results from this study showed that lumbo-sacral epidural anaesthesia with lidocaine could successfully be administered during dissociative anaesthesia of livestock pigs by a veterinary student and without reported post-injection complications. It improved analgesia and anaesthesia during herniorrhaphy of sufficient duration to enable surgical skills training. The risks and consequences of hypoxaemia and hypoventilation should be considered.