Development of an in vitro culture system for producing eel larvae from immature ovarian follicles in Japanese eel <Emphasis Type="Italic">Anguilla japonica</Emphasis>

To standardize conditions during the final maturation and ovulation of ovarian follicles from Japanese eel, we have developed a culture system for the production of fertilizable eggs from post-vitellogenic ovarian follicles in vitro. Post-vitellogenic ovarian follicles were incubated in culture medium supplemented with 17α,20β-dihydroxy-4-pregnen-3-one (DHP) with or without bovine serum albumin (BSA) to assess the effects of protein concentration. Eggs that ovulated during incubation were fertilized, and the remaining follicles were incubated in prostaglandin F_2α (PGF_2α) for a further 3 or 6 h before fertilization. Male eels were injected repeatedly with human chorionic gonadotropin. The quality of eggs obtained under the different culture conditions was evaluated after artificial fertilization in terms of hatching success. Hatching rates tended to decrease with increasing concentrations of BSA in the incubation medium in a dose-dependent manner. The addition of PGF_2α drastically increased the number of eggs that ovulated, but the rate of hatching was greatly decreased compared with eggs obtained earlier by DHP incubation alone. The larvae obtained from artificially fertilized eggs produced in vitro survived for 14 days without feeding. We conclude that in vitro culture systems thus have a great potential for the acquisition of good quality eggs under tightly controlled artificial conditions, culminating in the production of eel larvae.     

Protection conferred against viral nervous necrosis by simultaneous inoculation of aquabirnavirus and inactivated betanodavirus in the sevenband grouper, Epinephelus septemfasciatus (Thunberg)

An aquabirnavirus (ABV) and a formalin-inactivated betanodavirus [redspotted grouper nervous necrosis virus (RGNNV)] were investigated for their potential to prevent RGNNV-induced viral nervous necrosis (VNN) in the sevenband grouper, Epinephelus septemfasciatus (Thunberg). Three groups of fish were injected intramuscularly with ABV, intraperitoneally with inactivated RGNNV (iRGNNV) or with both ABV and iRGNNV. At 3, 7, 14, 21 and 28 days post-injection (p.i.), fish were challenged by intramuscular injection of RGNNV. Control fish, which received neither ABV nor iRGNNV, showed high mortalities in all RGNNV challenges. Fish that received only ABV exhibited relative percent survival (RPS) of >60 against RGNNV challenges at 3, 7, 14 and 21 days p.i., but not at 28 days p.i., while fish that received only iRGNNV showed significantly higher protection against RGNNV challenges only at 21 and 28 days p.i. In contrast, fish that received both ABV and iRGNNV showed 60 or higher RPS against all RGNNV challenges. Fish inoculated with iRGNNV with or without ABV exhibited similar high titres of neutralizing antibodies to RGNNV at 14, 21 and 28 days p.i. These results indicate that combined inoculation with iRGNNV and ABV conferred both rapid non-specific and delayed specific protection against VNN.     

Total intravenous anesthesia with ketamine–medetomidine–propofol in horses

Propofol is a potentially useful intravenous anesthetic agent for total intravenous anesthesia (TIVA) in horses. The purpose of this study was to compare the anesthetic and cardiorespiratory effects of TIVA following the administration of propofol alone(P–TIVA) and ketamine–medetomidine–propofol (KM–P–TIVA) in adult horses. The carotid artery was translocated to a subcutaneous position during TIVA with P–TIVA (n = 6) or KM–P–TIVA (n = 6). All horses were premedicated with medetomidine [0.005 mg kg^–1, intravenously (IV)]. Anesthesia was induced with midazolam (0.04 mg kg^–1 IV) and ketamine (2.5 mg kg IV). All horses were orotracheally intubated and breathed 100% oxygen. The KM drug combination (ketamine 40 mg mL^–1 and medetomidine 0.05 mg mL^–1) was infused at a rate of 0.025 mL kg^–1 hour^–1. Subsequently, a loading dose of propofol (0.5 mg kg^–1, bolus IV) was administered to all horses; surgical anesthesia (determined by horse response to incision and surgical manipulation, positive response being purposeful or spontaneous movement of limbs or head) was maintained by varying the propofol infusion rate as needed. Arterial blood pressure and HR were also monitored. Both methods of producing TIVA provided excellent general anesthesia for the surgical procedure. Anesthesia time was 115 ± 17 (mean ± SD) and 112 ± 11 minutes in horses anesthetized with KM–P–TIVA and P–TIVA, respectively. The infusion rate of propofol required to maintain surgical anesthesia with KM–P–TIVA was significantly less than for P–TIVA (mean infusion rate of propofol during anesthesia; KM–P–TIVA 0.15 0.02 P–TIVA 0.23 ± 0.03 mg kg^–1 minute^–1, p = 0.004). Apnea occurred in all horses lasting 1–2 minutes and intermittent positive pressure ventilation was started. Cardiovascular function was maintained during both methods of producing TIVA. There were no differences in the time to standing after the cessation of anesthesia (KM–P–TIVA 62 ± 10 minutes versus P–TIVA 87 ± 36 minutes, p = 0.150). The quality of recovery was good in KM–P–TIVA and satisfactory in P–TIVA. KM–P–TIVA and P–TIVA produced clinically useful general anesthesia with minimum cardiovascular depression. Positive pressure ventilation was required to treat respiratory depression. Respiratory depression and apnea must be considered prior to the use of propofol in the horse.     

Serum C-reactive protein (CRP) in horses: the effect of aging, sex, delivery and inflammations on its concentration.

The serum concentration of C-reactive protein (CRP) in horses was measured by single radial immunodiffusion (SRID) using rabbit anti-equine CRP serum to evaluate the significance of CRP as an acute-phase reactive protein. In serum samples of clinically normal newborn thoroughbred foals before being given colostrum, serum CRP was not detectable (less than or equal to 1 microgram/ml). The serum CRP concentration was found to increase quickly and reach a peak level of approximately 14.1 micrograms/ml in 12-month-old horses. Then, it decreased gradually to reach a low value of 5.4 micrograms/ml in the 4-year-old. In horses over 5-year-old, serum CRP concentration was stable at values of 7 to 8 micrograms/ml. In mares during the peri-natal period, serum CRP concentration decreased at 2 months before delivery, showed moderate changes just before and after delivery, and then increased to relatively high level by 2 months after delivery. The concentration of serum CRP was found to be increased at 24 hours after experimentally inflammatory stimuli in adult horses, and reached peak values of 3 to 6 times as high as the base line values at 3 to 5 days. Serum CRP concentration returned to base line values by 2 to 3 weeks after treatments. It is clear from these data that CRP is one of the acute-phase reactive proteins in horses.     

Isolation and species distribution of staphylococci from animal and human skin

From April 1999 to December 2000, a survey was made on the distribution of Staphylococcus species on the skin of 7 kinds of animals and humans. Staphylococci were isolated from 12 (100%) of 12 pigs, 17 (89.5%) of 19 horses, 30 (100%) of 30 cows, 73 (90.1%) of 81 chickens, 10 (40%) of 25 dogs, 23 (76.7%) of 30 laboratory mice, 20 (52.6%) of 38 pigeons, and 80 (88.9%) of 90 human beings. The predominant staphylococci isolated from a variety of animal species were novobiocin-resistant species, S. xylosus and S. sciuri regardless of the animal host species. The novobiocin-resistant species including S. xylosus and S. sciuri were only occasionally isolated from human skin. The predominant staphylococci found on human skin were novobiocin-sensitive species, S. epidermidis (63.8%), followed by S. warneri (28.8%) and S. hominis (13.8%). The results suggest that the staphylococcal flora inhabiting animal skin are different from those of human skin in regard to the predominant species isolated. In this study, we used pulsed-field gel electrophoresis to examine the chromosomal polymorphisms of S. epidermidis isolated most frequently from human skin. Strains of S. epidermidis showed the greatest genomic diversity in their fragment patterns.     

Seasonal atopic dermatitis in dogs sensitive to a major allergen of Japanese cedar (Cryptomeria japonica) pollen

Three dogs were examined because of episodes of recurrent pruritic dermatitis in the spring, the season of Japanese cedar (Cryptomeria japonica, CJ) pollination in Japan. The dogs were shown to be sensitive to CJ pollen allergen using intradermal testing and antigen-specific IgE measurement. Fluorometric enzyme-linked immunosorbant assay (ELISA) showed increased concentrations of IgE specific to Cry j 1 and a negative result for Cry j 2 in the three dogs. The concentrations of IgE specific to Cry j 1 during the season of CJ pollination were higher than the concentrations found during the off-season in all the dogs, and the variation in the concentrations correlated with the variation in clinical signs. Peripheral blood mononuclear cells showed apparent proliferative responses to crude CJ pollen antigen and Cry j 1 during CJ pollination season. These findings indicated that Cry j 1 was the major allergen recognized by IgE and lymphocytes and resulted in the development of type I hypersensitivity to CJ pollen allergen in these atopic dogs.     

Comparison of response to immunotherapy by intradermal skin test and antigen-specific IgE in canine atopy

The intradermal skin test (IDST) and serologic allergy test (SAT) has been developed for confirming a diagnosis of canine atopy and determining allergens for immunotherapy. To determine the prevalence of causative allergens for canine atopic dermatitis in Japan, IDST and SAT were performed with the CMG Immunodot strips on 95 atopic dogs using 9 allergens. In addition, we compared agreement rate, sensitivity and specificity between them (using IDST as the standard). The allergen most commonly positive in both tests was house dust mites (IDST: 69.5%, SAT: 48.4%). Moreover, Japanese cedar, mugwort and grass mix were detected as attendant causative allergens. Agreement rates between the two tests ranged from 67.4% to 96.8%; the overall mean agreement rate were 81%. SAT was shown to have sensitivity to IDST ranging from 16.7 to 68.2%. The specificities were very high for all allergens, on the order of 94.9-100% (median=98.7%). Finally, the efficacy of immunotherapy was evaluated on 27 atopic dogs based on IDST (15 dogs) and SAT (12 dogs) results. Overall, 60% (9/15) of the IDST group and 66.8% (8/12) of the SAT group experienced a 50% to 100% reduction in their symptomatology. No significant differences were found in response to immunotherapy during the follow-up period between allergen selection methods. These results indicate the value of serologic tests as an aid to identifying an allergen solution for immunotherapy.     

Cardiovascular effects of medetomidine, detomidine and xylazine in horses

The cardiovascular effects of medetomidine, detomidine, and xylazine in horses were studied. Fifteen horses, whose right carotid arteries had previously been surgically raised to a subcutaneous position during general anesthesia were used. Five horses each were given the following 8 treatments: an intravenous injection of 4 doses of medetomidine (3, 5, 7.5, and 10 microg/kg), 3 doses of detomidine (10, 20, and 40 microg/kg), and one dose of xylazine (1 mg/kg). Heart rate decreased, but not statistically significant. Atrio-ventricular block was observed following all treatments and prolonged with detomidine. Cardiac index (CI) and stroke volume (SV) were decreased with all treatments. The CI decreased to about 50% of baseline values for 5 min after 7.5 and 10 microg/kg medetomidine and 1 mg/kg xylazine, for 20 min after 20 microg/kg detomidine, and for 50 min after 40 microg/kg detomidine. All treatments produced an initial hypertension within 2 min of drug administration followed by a significant decrease in arterial blood pressure (ABP) in horses administered 3 to 7.5 microg/kg medetomidine and 1 mg/kg xylazine. Hypertension was significantly prolonged in 20 and 40 microg/kg detomidine. The hypotensive phase was not observed in 10 microg/kg medetomidine or detomidine. The changes in ABP were associated with an increase in peripheral vascular resistance. Respiratory rate was decreased for 40 to 120 min in 5, 7.5, and 10 microg/kg medetomidine and detomidine. The partial pressure of arterial oxygen decreased significantly in 10 microg/kg medetomidine and detomidine, while the partial pressure of arterial carbon dioxide did not change significantly. Medetomidine induced dose-dependent cardiovascular depression similar to detomidine. The cardiovascular effects of medetomidine and xylazine were not as prolonged as that of detomidine. KEY WORDS: cardiovascular effect, detomidine, equine, medetomidine, xylazine.