Molecular characterization of tetracycline- and quinolone-resistant Aeromonas salmonicida isolated in Korea

The antibiotic resistance of 16 Aeromonas (A.) salmonicida strains isolated from diseased fish and environmental samples in Korea from 2006 to 2009 were investigated in this study. Tetracycline or quinolone resistance was observed in eight and 16 of the isolates, respectively, based on the measured minimal inhibitory concentrations. Among the tetracycline-resistant strains, seven of the isolates harbored tetA gene and one isolate harbored tetE gene. Additionally, quinolone-resistance determining regions (QRDRs) consisting of the gyrA and parC genes were amplified and sequenced. Among the quinolone-resistant A. salmonicida strains, 15 harbored point mutations in the gyrA codon 83 which were responsible for the corresponding amino acid substitutions of Ser⁸³→Arg⁸³ or Ser⁸³→Asn⁸³. We detected no point mutations in other QRDRs, such as gyrA codons 87 and 92, and parC codons 80 and 84. Genetic similarity was assessed via pulsed-field gel electrophoresis, and the results indicated high clonality among the Korean antibiotic-resistant strains of A. salmonicida.     

Immunohistochemical and histomorphometric evaluation of vascular distribution in intact canine cranial cruciate ligament

Objectives: To (1) describe vascular distribution in the grossly intact canine cranial cruciate ligament (CCL) using immunohistochemical techniques specific to 2 components of blood vessels (factor VIII for endothelial cells, laminin for basement membrane); and (2) compare the vascularity in different areas of interest (craniomedial versus caudolateral bands; core versus epiligamentous regions; and proximal versus middle versus distal portions) in the intact normal canine CCL. Study Design: In vitro study. Animals: Large, mature dogs (n=7) of breeds prone to CCL disease that were euthanatized for nonorthopedic conditions. Methods: Intact CCL were collected from fresh canine cadavers free from stifle pathology. CCL tissue was processed for immunohistochemistry and stained for factor VIII and laminin. Vascular density was determined by histomorphometric analysis. Results: Specific vascular staining was sparsely identified throughout the CCL; however, the proximal portion of the CCL appears to have a greater number of vessels than the middle or distal portion of the ligament. Conclusions: The CCL is a hypovascular tissue and its vascular distribution is not homogeneous.     

Immunohistochemistry and polymerase chain reaction for the detection of Lawsonia intracellularis in porcine intestinal tissues with proliferative enteropathy

Detection method of Lawsonia intracellularis was studied in formalin-fixed paraffin-embedded intestinal tissues from 5 naturally infected pigs by immunohistochemistry with a monoclonal antibody against outer membrane protein of L. intracellularis. Warthin-Starry silver stain revealed clusters of argyrophilic, slightly curved rod-shaped organisms in the apical cytoplasm of enterocytes. Immunohistochemical staining with a L. intracellularis-specific monoclonal antibody confirmed the presence of the organism in the apical cytoplasm of hyperplastic enterocytes. The presence of L. intracellularis in the ileum of pig with proliferative enteropathy was confirmed by polymerase chain reaction (PCR) further on the basis of amplification of 319 base pair products specific for porcine L. intracellularis chromosomal DNA. Immunohistochemistry and PCR may be a complementary method to confirm the diagnosis of L. intracellularis infection in pigs.     

Determination of Feeder Cell‐Based Cellular Niches Supporting the Colonization and Maintenance of Spermatogonial Stem Cells from Prepubertal Domestic Cat Testes

Recently, isolation and in vitro culture of putative spermatogonial stem cells (SSCs) in the domestic cat have been conducted. However, the cellular niche conditions that facilitate the establishment and long‐term maintenance of feline SSCs (FSSCs) have not been described. Therefore, we investigated the type of feeder cells used to stimulate colony formation and growth of FSSCs among the various factors in the FSSC niche. Spermatogonial stem cells isolated from feline testes were cultured on mitotically inactivated testicular stromal cells (TSCs) derived from cats, dogs and mice, and mouse embryonic fibroblasts (MEFs). The formation and growth of colonies derived from SSCs cultured on each type of feeder cell were identified at passage 0, and the morphology, alkaline phosphatase (AP) activity and expression of SSC‐specific genes in surviving colonies were investigated at passage 4. Among these diverse feeder cells, TSCs from cat showed the greatest colony formation, growth and maintenance of FSSCs, and SSC colonies cultured by passage 4 showed a typical dome‐shaped morphology, AP activity and expression of SSC‐specific genes (NANOG, OCT4, SOX2 and CD9). Accordingly, these results demonstrate that feline TSCs could be used as feeder cells to support the establishment and maintenance of SSCs from domestic cats.     

Anesthetic management of an off-pump open-heart surgery in a dog

ObservationsA 9 year-old, 40 kg, female spayed Bouvier des Flandres was anesthetized for surgical removal of an intra-cardiac mass. Pre-anesthetic work-up included thoracic radiographs, which revealed moderate pleural effusion, and cardiac ultrasound, which identified a mass attached to the wall of the right ventricular outflow tract (RVOT). The mass caused dynamic obstruction of the RVOT during systole. The dog was pre-medicated with intravenous (IV) hydromorphone (0.05 mg kg^?1). Following pre-oxygenation, anesthesia was induced with ketamine (3.75 mg kg^?1, IV) and diazepam (0.18 mg kg^?1, IV). Anesthesia was maintained with isoflurane in oxygen, an intravenous constant rate infusion (CRI) of fentanyl (10–30 μg kg^?1 hour^?1) and a CRI of lidocaine (50–200 μg kg^?1 minute^?1). A right lateral thoracotomy was performed. The heart was stopped transiently with a cold cardioplegic solution for 7.83 minutes to allow the removal of the mass through an open-heart procedure. No cardiopulmonary bypass was used. The heart was successfully restarted after cardiopulmonary resuscitation with internal cardiac massage and internal defibrillation. The dog recovered uneventfully from anesthesia without any apparent neurological sequelae. Post-operative analgesia consisted of intercostal nerve blocks with bupivacaine, CRIs of fentanyl (2–5 μg kg^?1 hour^?1) and lidocaine (40 μg kg^?1 minute^?1) and with oral meloxicam (0.1 mg kg^?1). Five days following surgery, the dog was discharged from the hospital. Histopathology and immunohistochemistry of the mass identified an ectopic thyroid carcinoma.ConclusionsThis case showed the feasibility of whole body hypothermia and using a cold cardioplegic solution to induce cardiac arrest for a short open-heart procedure.     

Effects of mid-season avermectin treatments on pyrethroid resistance in horn fly (Diptera: Muscidae) populations at three locations in Louisiana

Between 1999 and 2002, the effect of mid-season doramectin treatments on the level of resistance in pyrethroid-resistant horn fly populations was examined at three separate Louisiana State University Agricultural Center research stations. The cattle were treated with pyrethroid ear tags in all years at all farms, and each farm received a mid-season doramectin treatment in 1 year. The number of weeks of control at Red River was 11 weeks higher in the year following the mid-season treatment of doramectin. At Macon Ridge, the number of weeks of control was 2 weeks higher in the year following the doramectin treatment. No change was observed at St. Joseph. The LC50s for fly populations tested at Macon Ridge and St. Joseph were found to increase for pyrethroids from the spring populations to the fall populations between 2000 and 2002. The LC50s for fly populations at Red River followed the same trends except in 2000, the year when the doramectin treatment was administered. Flies collected pre and post-treatment each year from St. Joseph and Red River were assayed for two alleles (kdr and skdr) associated with target site resistance to pyrethroids. Flies collected pretreatment at Macon Ridge in 1999 also were assayed for the kdr and skdr, and this population of flies had a frequency of 85.6% R-kdr alleles. At St. Joseph and Red River there was a general decline in the frequency of homozygous susceptible skdr (SS-skdr) and homozygous susceptible kdr (SS-kdr) individuals, as well as a general increase in homozygous resistant skdr (RR-skdr) and homozygous resistant kdr (RR-kdr) individuals, during the 4-year study. At both sites, the frequency of R-kdr alleles increased significantly in flies collected in the fall compared to flies collected in the spring with the exception of Red River in 2000, when dormacetin was applied. The frequency of the R-kdr alleles was significantly higher in flies collected in the fall compared to flies collected in the spring in the following year at both sites in two out of three comparisons. The frequency of R-skdr alleles was significantly lower in fly populations tested in the spring compared to fly populations tested in the fall at both farms in years when doramectin was not applied but there were no differences in the years when doramectin was applied. The frequency of R-skdr alleles was significantly higher in fly populations tested in the fall compared to in the spring the following year during all three comparisons at Red River and in one of three comparisons at St. Joseph.     

Comparison of antibody titers in rabbits following immunization with inactivated influenza virus via subarachnoidal or subcutaneous route

Rabbits were immunized with inactivated influenza virus via the subarachnoidal (SA) or subcutaneous (SC) route, and the antibody titers in cerebrospinal fluid (CSF) and serum were assayed. There were no nervous signs or morphological lesions related to SA immunization. In the SC group, the antibody titer was elevated in serum, but not elevated in CSF. In the SA group, the antibody titer was significantly elevated in serum and even in CSF, and their antibody titers were much greater than in the SC group. The present results suggest that intrathecal immunization is more effective than SC immunization at inducing a protective immune response against the transneural spread of viruses.     

Tibial plateau symmetry and the effect of osteophytosis on tibial plateau angle measurements

A novel technique was developed to estimate the caudal medial tibial plateau landmark in the face of osteophytosis to improve accuracy in tibial plateau angle measurements. Using this technique, tibial plateau angles were evaluated in 31 normal dogs before and 8 months after right cranial cruciate ligament transection. There was no significant difference in mean tibial plateau angle before or after induction of osteophytosis. Additionally, it was determined that 90% of dogs had a difference of =2 degrees between right and left tibial plateau angles, which was considered symmetrical.     

Ameliorative Effects of Melatonin against Hydrogen Peroxide‐Induced Oxidative Stress on Boar Sperm Characteristics and Subsequent In Vitro Embryo Development

Melatonin, the major secretory product of the pineal gland, scavenges a variety of reactive oxygen and nitrogen species in vivo and in vitro, indicating that melatonin is a potent function as an antioxidant. The objective of this study was to investigate the effect of melatonin in the presence or absence of hydrogen peroxide (H₂O₂) on sperm characteristics (motility, viability, survival rate, membrane integrity, lipid peroxidation (LPO) and mitochondria activity) and also to examine the developmental rates to the blastocysts stage of porcine oocytes fertilized in vitro with semen treated with or without melatonin (100 nm ) in the presence or absence of H₂O₂ (250 μm ). The sperm were treated with melatonin in the presence or absence of H₂O₂ for 3, 6, 9 and 12 h at 37°C and then analysed for the sperm characteristics. The porcine embryos were produced by in vitro maturation and in vitro fertilization (IVM/IVF) using semen treated with or without melatonin (100 nm ) in the presence or absence of H₂O₂ (250 μm ) for 6 h. The semen characteristics, including motility, viability, survival rate, membrane integrity and mitochondria activity, were higher in the groups that were treated with melatonin in comparison to other groups, irrespective of incubation periods. Malondialdehyde levels in control, melatonin and melatonin + H₂O₂ groups were lower than H₂O₂ only group. A positive correlation was shown among motility, viability, survival rate and membrane integrity, but a negative correlation was observed between LPO and the other evaluation methods. The developmental rates to blastocysts of IVM/IVF porcine oocytes fertilized by semen treated with melatonin were significantly increased compared with any other groups, with the cell number of blastocysts shown to have a similar trend to the developmental rates. These results demonstrate that melatonin can improve the semen characteristics during in vitro storage and support the developmental ability of IVM/IVF embryos in pigs.