Validation of RT‐PCR Assays for Molecular Characterization of Porcine Teschoviruses and Enteroviruses

Porcine enteroviruses (PEVs) and teschoviruses (PTVs) are described as causative agents of neurological disorders, fertility disorders and dermal lesions of swine. Difficulties in the serological detection of these viruses may lead to a significant underestimation of infections with clinical symptoms. With the recent availability of genome sequence data for all the serotypes, molecular diagnosis is a possibility. The present study describes a new approach to molecular ‘serotyping’ of PTVs and PEV‐B viruses, involving the amplification and sequencing of a genomic fragment of the VP1 coding region. A molecular characterization of Italian entero‐teschovirus isolates was performed using a set of previously published and newly designed polymerase chain reaction primers. A total of 33 porcine isolates and 10 reference strains were analysed. Porcine enterovirus‐B samples were first diagnosed as positive for enterovirus by amplification of the 5′‐non‐translated region. Samples were then typed by amplification and sequencing of a portion of the VP1 coding region. Porcine enterovirus‐A and PTVs were detected by a published assay in the 5′‐NC region that allows them to be differentiated according to the size of amplification product, using the same set of primers. For serotype characterization of PTV, we evaluated four different regions: the N terminus of the capsid protein VP2, the region encoding for RNA‐dependent RNA polymerase, and the capsid VP1 and VP4 regions. The newly designed primers in the VP1 region was proved to be broad in range and suitable for serotype assessment and therefore constitute a useful diagnostic tool for molecular diagnosis of porcine teschovirus/enterovirus strains and for the study of molecular epidemiology and evolution of these viruses.     

Molecular systematics in Chrysanthemum × grandiflorum (Ramat.) Kitamura

An attempt was made to understand the molecular systematics and genetic differences between 10 original chrysanthemum cultivars and 11 mutants. The similarity among the cultivars and mutants varied from 0.17 to 0.90 using RAPD analyses, a simple but efficient method to distinguish cultivars and to assess parentage. Two distinct groups were found. Two cultivars were present as a separate group showing differences from all other cultivars. Mutants with different flower colour could be identified at the molecular level using RAPD technique holding promise to identify unique genes as SCAR markers. A high genetic distance among the different chrysanthemums showed that there exists a possibility of introgressing new and novel genes from the chrysanthemum gene pool.     

Pericarp total protein profiles as molecular markers of tomato fruit quality traits in two segregating populations

The tomato fruit quality results of biochemical and physiological changes that occur during the ripening process. Although, the pericarp total protein profiles are less polymorphic than DNA-based markers the polymorphism in those could be directly associated with fruit quality traits. The aim of this work was to identify associations between polymorphic polypeptides from fruit pericarp at two ripening stages and fruit quality traits evaluated in two segregating populations of tomato. A cross between a normal ripening cultivar of Solanum lycopersicum (C, Caimanta) and a genotype carrying the nor (non ripening) gene (N) as well as a cross between Caimanta and a cherry type tomato of S. lycopersicum var. cerasiforme (Ce) showed genetic variance for several fruit quality traits such as fruit weight, shape, solids soluble content, acidity, color and fruit shelf life. The quantitative variations observed at phenotypic level had correspondence with the polymorphism detected in the protein profiles. Indeed, the polymophic polypeptides associated with quality fruit traits and fruit shelf life would be useful to assist tomato breeding programs as protein molecular markers.     

Characterization of semen collected by pharmacologically induced ejaculation from a stallion with seminal vesiculitis

The present study compared the quality of sperm collected by artificial vagina or pharmacologically induced ejaculation from a 10-year-old thoroughbred stallion with seminal vesiculitis. The pharmacological protocol involved intravenous administration of detomidine (0.01 mg/kg) and oxytocin (20 IU) and successfully induced ejaculation in all attempts of semen collection. Sperm motility, plasma membrane and acrosome integrity (PMAI), reactive oxygen species (ROS) levels, polymorphonuclear neutrophil (PMN) percentage, and bacterial profiles of fresh and cooled semen (5°C for 24 hr) were evaluated. Semen obtained by the pharmacological method presented reduced seminal volume, decreased PMN percentage and superior sperm motility in cooled samples. Moreover, higher PMAI and lower ROS levels were observed in semen collected by the pharmacological method. Therefore, pharmacologically induced ejaculation is an alternative to obtain semen with minimal contamination and with sperm of superior quality and longevity from stallions with seminal vesiculitis.     

Electro-acupuncture and Chinese herbs for treatment of cervical intervertebral disk disease in a dog

A non-ambulatory dog with tetraparesis following a pain episode that had evolved over 2 months was submitted for medical treatment and diagnosed with intervertebral disk disease at C3-C4 and dorsal extradural compression at C1-C2 and C3-C4 using myelography and computed tomography. The dog experienced ambulation recovery after 15 days of treatment with only electroacupuncture and Chinese herbal medicine, with marked improvement occurring after only 10 treatments. Six months of follow-up demonstrated that the dog was stable and had no recurrence of symptoms. Therefore, it was concluded that the combination of electroacupuncture and Chinese herbal medicine was responsible for motor rehabilitation.     

Effects of α-tocopherol and Ascorbic Acid on Equine Semen Quality after Cryopreservation

This study investigated the effects of ascorbic acid and α-tocopherol supplementation on semen quality parameters of equine thawed-frozen semen. Semen was divided in seven different treatments in a final concentration of 100 × 10⁶ sperm/mL by using Gent extender containing no supplements (control) and the following supplements with three different concentrations: α-tocopherol (0.5, 1, and 2 mM) and ascorbic acid (0.45, 0.9, and 1.8 g/L). After thawing, all samples were maintained at 37°C, while analyses were performed at 0, 60, and 120 minutes. Evaluation of viability and acrosome status (using Pisum sativum agglutinin conjugated to fluorescein isothiocyanate and propidium iodide), mitochondrial membrane potential (5,5′,6,6′-tetrachloro-1,1′,3,3′tetraethylbenzimidazolyl carbocyanine iodine [JC-1]), membrane lipid peroxidation (LPO; C₁₁-BODIPY^581/591), and stability of the plasmatic membrane (merocyanine 540 and Yo-Pro-1) of each sample was determined by flow cytometry. Relative to the control group, supplementation with α-tocopherol improved (P ≤ .05) postthaw membrane LPO, yet the higher concentrations of ascorbic acid (0.9 and 1.8 g/L, respectively) showed a negative effect on membrane LPO. Neither antioxidant significantly increased (P > .05) the acrosome integrity and mitochondrial membrane potential of frozen-thawed spermatozoa, although supplementation with α-tocopherol and ascorbic acid (0.9 and 1.8 g/L, respectively) had a positive effect on membrane integrity and stability (P ≤ .05). For all semen parameters, the lower concentration of ascorbic acid (0.45 g/L) did not show significant differences (P > .05) compared with the control. In conclusion, α-tocopherol seems to be an efficient antioxidant for reducing the oxidative stress provoked by cryopreservation, decreasing lipid peroxidation on equine spermatozoa.     

MALDI‐MS Lipid Profiles of Oocytes Recovered by Ovum Pickup from Bos indicus and 1/2 indicus × taurus with High vs Low Oocyte Yields

The aim of the present study was to compare the lipid profile in oocytes of indicus and 1/2 indicus × taurus cows with high and low antral follicle count (AFC)/oocyte yields. After an OPU procedure (D0), antral follicles ≥3 mm were counted by ultrasonography (D4, 19, 34, 49, 64), and cows were assigned to groups with either high AFC (≥30 follicles; indicus, NH group; 1/2 indicus × taurus, AH group) or low AFC (≤15 antral follicles; indicus, NL group; 1/2 indicus × taurus, AL group). The lipid profiles of the oocytes were determined by MALDI‐MS. For GI, GII and GIII oocytes, the indicus samples tend to cluster separately from the 1/2 indicus × taurus samples. The lipid species [PC (P‐38:5) + H]⁺ and/or [PC (P‐36:2) + Na]⁺, [PC (38:2) + H]⁺, [PC (38:5) + Na]⁺ and [TAG (60:8) + NH⁴]⁺ were more abundant in indicus (NH and NL groups) than 1/2 indicus × taurus. The higher lipid content in the indicus oocytes likely reflects differences in the rate of lipid metabolism and may contribute to oocyte competence and embryo development.     

Vaccination of one-day-old broiler chicks against infectious coryza

In order to evaluate the protection conferred by an experimental inactivated vaccine against infectious coryza, three challenge trials were undertaken using 112 1-day-old broilers. The vaccine "Hepa Inmuno NC" included bacterial antigens of Avibacterium paragallinarum (serogroups A, B, variant B, and C) as well as antigens of Newcastle virus and hepatitis virus. Fifty-six broiler chicks were vaccinated at the first day of life at the hatchery while another 56 chicks were left unvaccinated. Three infection trials were conducted simultaneously using each of the three serogroups A, B, or C of Av. paragallinarum. In each trial, 17 vaccinated and 17 unvaccinated broilers were used. Challenge was performed at day 31 of life by injection, into the left infraorbital sinus, of approximately 1 x 10(5) colony forming units of the corresponding Av. paragallinarum strain. Clinical signs were recorded on day 2 postchallenge. All broilers were euthanatized and both infraorbital sinuses were bacteriologically examined for the presence of Av. paragallinarum on day 5 postchallenge. In comparison with the unvaccinated broilers, the vaccine significantly reduced the number of broilers with clinical signs after challenge with serogroup B, and significantly fewer vaccinated broilers were positive for the presence of Av. paragallinarum after challenge with serogroup C. On the other hand, no significant protection was observed when broilers were challenged with Av. paragallinarum from serogroup A. Despite the high infection rates in vaccinated chicks after an experimental infection with Av. paragallinarum, it was possible to reduce colonization of Av. paragallinarum (serogroup B) and clinical signs (serogroup C) in broiler chicks by vaccination at the first day of life. Further cross-protection trials should be done, including other Av. paragallinarum strains in the vaccine, especially those from serogroup A.     

Chitosan associated with whole raw soybean in diets for Murrah buffaloes on ruminal fermentation,apparent digestibility and nutrients metabolism

This study aimed to investigate the effects of chitosan and whole raw soybean on nutrient intake, apparent digestibility, nitrogen utilization, microbial protein synthesis, blood metabolites, feeding behavior, ruminal fermentation, digesta kinetics, and reticular flow of nutrients of buffaloes. Four ruminally‐cannulated Murrah buffaloes (351 ± 15 kg of initial BW) were randomly assigned according to a 4 × 4 Latin square design. Treatments were arranged as 2 × 2 factorial arrangement: the first factor was whole raw soybean (WRS), and the second factor was chitosan (CHI) with or without their inclusion in diets. Intake and apparent digestibility of ether extract (p < .01; p = .04, respectively), non‐fiber carbohydrates intake (p = .03) and apparent ruminal digestibility of dry matter (p = .01) were affected by diets. An interaction effect or tendency was observed for microbial nitrogen (p = .09), concentrations, ruminal ammonia nitrogen (p = .05), total volatile fatty acid (p = .03). Association of chitosan with whole raw soybean has potential effects as a modulator of rumen fermentation; therefore, chitosan can be applied as an alternative non‐ionophore for Murrah buffaloes.     

Laparoscopic nephron-sparing surgery for the treatment of canine dioctophymosis

This paper reports on two cases of laparoscopic nephrotomy employed in the treatment of canine dioctophymosis, which is considered a unusual procedure and a new treatment proposal heretofore not performed in veterinary medicine. Two patients were treated, one with a history of hematuria and the other with incidental finding of the parasite in the abdominal cavity during elective ovariohysterectomy. Both dogs were subjected to abdominal ultrasound, which produced images indicating the presence of the parasite in the right side kidney, but with partial parenchymal preservation. The patients were therefore subjected to laparoscopic nephrotomy. The surgical procedure was effective in treating dioctophymosis and enabled minimum tissue invasion during surgery, in addition to preservation of the kidney.