Pharmacokinetics of articaine hydrochloride and its metabolite articainic acid after subcutaneous administration in red deer (Cervus elaphus)

AIM: To develop and validate a simple and sensitive method using liquid chromatography-mass spectrometry (LC-MS) for quantification of articaine, and its major metabolite articainic acid, in plasma of red deer (Cervus elaphus), and to investigate the pharmacokinetics of articaine hydrochloride and articainic acid in red deer following S/C administration of articaine hydrochloride as a complete ring block around the antler pedicle.

METHODS: The LC-MS method was validated by determining linearity, sensitivity, recovery, carry-over and repeatability. Articaine hydrochloride (40?mg/mL) was administered S/C to six healthy male red deer, at a dose of 1?mL/cm of pedicle circumference, as a complete ring block around the base of each antler. Blood samples were collected at various times over the following 12 hours. Concentrations in plasma of articaine and articainic acid were quantified using the validated LC-MS method. Pharmacokinetic parameters of articaine and articainic acid were estimated using non-compartmental analysis.

RESULTS: Calibration curves were linear for both articaine and articainic acid. The limits of quantifications for articaine and articainic acid were 5 and 10?ng/mL, respectively. Extraction recoveries were >72% for articaine and >68% for articainic acid. After S/C administration as a ring block around the base of each antler, mean maximum concentrations in plasma (C_max) of articaine were 1,013.9 (SD 510.1) ng/mL, detected at 0.17 (SD 0.00) hours, and the C_max for articainic acid was 762.6 (SD 95.4) ng/mL at 0.50 (SD 0.00) hours. The elimination half-lives of articaine hydrochloride and articainic acid were 1.12 (SD 0.17) and 0.90 (SD 0.07) hours, respectively.

CONCLUSIONS AND CLINICAL RELEVANCE: The LC-MS method used for the quantification of articaine and its metabolite articainic acid in the plasma of red deer was simple, accurate and sensitive. Articaine hydrochloride was rapidly absorbed, hydrolysed to its inactive metabolite articainic acid, and eliminated following S/C administration as a ring block in red deer. These favourable pharmacokinetic properties suggest that articaine hydrochloride should be tested for efficacy as a local anaesthetic in red deer for removal of velvet antlers. Further studies to evaluate the safety and residues of articaine hydrochloride and articainic acid are required before articaine can be recommended for use as a local anaesthetic for this purpose.     

Policies for the vaccination of cats and dogs in New Zealand veterinary practices

AIMS: To determine current practices and attitudes towards vaccination of dogs and cats of veterinarians in New Zealand; the methods used for informing clients on which vaccines to use, and the preferred site for vaccination of cats.

METHODS: A postal questionnaire was sent to all 483 listed veterinary practices in New Zealand during February 2012. Some questions were specific to pet dogs, cats, or working farm dogs. Responses were categorised according to practice type and geographical region of the respondent. Factors associated with respondent recommendation of annual vaccination with modified live viral (MLV) vaccines were examined using logistic regression analysis. Vaccines that were considered to be essential for every animal were defined as core; those that may be recommended for animals whose location or lifestyle placed them at risk, were defined as non-core.

RESULTS: There were 204 useable returns, equivalent to a response rate of 42.2%, distributed across the country. Annual vaccination with MLV vaccines of dogs was recommended by 54/198 (27.3%) respondents, and of cats by 107/181 (59.1%) respondents. Factors associated with the recommendation of annual administration of MLV vaccines to dogs included being a companion animal practice, a desire for policies on vaccination to be left to individual clinics, and having one veterinarian in the practice. Administration of the final vaccination for puppies was recommended at ≥14 weeks old by 55/185 (29.7%) respondents, and for kittens at ≥13 weeks old by 42/183 (23%) respondents. Of respondents that administered MLV vaccines annually, 62/103 (60.2%) believed reducing the frequency of vaccination would reduce income, and 52/103 (50.5%) considered it would have a negative effect on animal health. Advice to enable clients to decide which non-core vaccines were administered was given by 181/199 (91%) respondents. Factors considered when recommending a vaccine included consideration of risk to individual patients (190/203; 93.6%), requirements of boarding kennels/catteries (165/203; 81.3%) and clinic vaccination policy (142/203; 70%). The preferred site for administering MLV vaccines to cats was the dorsal neck or inter-scapular region (137/198; 69.2%). Amongst respondents, 18 wanted disease surveillance information to allow for truly informed decisions to be made about vaccination.

CONCLUSIONS AND CLINICAL RELEVANCE: Veterinarians can now compare their own vaccination practices and attitudes with those of veterinarians nationally, and internationally. There is a need for national surveillance information and for continued education of the public and commercial kennel and cattery owners for optimal vaccination strategies to be developed.     

Attitudes and perceptions of veterinary paraprofessionals in New Zealand to postoperative pain in dogs and cats

AIM: To survey the attitudes and perceptions of veterinary paraprofessionals in New Zealand to postoperative pain in dogs and cats.

METHODS: In December 2011, veterinary paraprofessionals (VP) from throughout New Zealand were invited to participate in an online survey. Eleven questions, which were divided into five sections, were used to determine demographic information, the respondents’ assessment of pain after commonly performed surgeries in dogs and cats, their opinions on provision of analgesia, who had responsibility for pain monitoring and the use of any formal pain scoring system in the practice.

RESULTS: Data from 165 respondents were able to be used, and 162 (98%) respondents to the survey were female. According to the respondents’ estimates, fracture repair in dogs and repair of diaphragmatic hernias in cats had the highest pain score following surgery. Neutering procedures involving dogs were scored higher than for cats (p<0.01). All respondents agreed that animals benefit from perioperative analgesia. The veterinary nurse was reported to be predominantly responsible for monitoring pain in animals postoperatively by 116/165 (70.3%) respondents. Of 165 respondents, 154 (93%) considered that their knowledge of pain and assessment of pain could be enhanced.

CONCLUSIONS: This survey reflects the attitudes and perceptions of a sample of VP in New Zealand to postoperative pain in dogs and cats. The results indicate that all respondents believe that surgery results in sufficient pain to warrant analgesic therapy. Routine neutering surgeries were considered to be more painful in dogs than in cats. The current survey also provides information to educators on potential areas of focus, given that 93% of respondents felt that their knowledge of pain and assessment of pain could be enhanced.     

Utilization of nylon membranes for the specific isolation of Escherichia coli O157 with DNA probes and checking for relationship to the shiga toxin Escherichia coli (STEC) group

A method for specific isolation of Escherichia coli strains of serotype O157 is given. DNA-hybridization technique by using DIG-labeled specific O157 PCR-amplificates as probes is the basis. These investigations can be used for the detection and isolation of shigatoxin-producing Escherichia coli (STEC) of this serotype in combination with other tests for detection of shigatoxin genes (stx). No background is seen by using 'DIG Easy Hyb' solution and Nylon membranes for colony-and plaque-hybridization (Roche Diagnostics Boehringer Mannheim). Dark brown spots (E. coli colonies) are visible on the membranes after staining. After comparing the membrane with the masterplate it is possible to isolate the colonies from the masterplate, respectively. It is necessary to make proof that the isolates possess the rfb-genes (verification of serotype O157) and the stx-genes (test for belonging to the STEC-group). Both tests could be done by using PCR. In case of positive results other virulence factors of the isolates can be detected.     

Detection and prevalence of verotoxin-producing Escherichia coli (VTEC) in raw sausage

We investigated 158 samples of shortly ripened raw sausages bought in supermarkets of Dessau within 4 month. In 14 (8.8%) samples Verotoxin-producing E. coli were detected. 13 VT-positive samples were found in the group of easily spread raw sausages. The 14 isolates belonged to 6 different O-serotypes. 4 VT1-, 8 VT2- and 2 VT1/VT2-producers were found. 4 isolates belonged to serogroups which were already described in WHO tables and associated with EHEC infections in human beings. One strain of serogroup O22: H8, isolated from a "Teewurst", possessed the complete virulence gene combination of EHEC (eae, hlyA, stx). The detection procedure, already successfully used for detection and isolation of VTEC from raw milk, soft cheese and raw minced beef showed a sensitivity of approximately 10 CfU/25 g of raw sausages. It has to be considered that VTEC are frequently (8.8%) present in shortly ripened raw sausages. The group of easily spread raw sausages has a higher VTEC-contamination rate than firm raw sausages. Raw sausages, especially easy to spread types, belong to the risk foods for EHEC-infections in human beings.     

Molecular biological method cascade for detection, specific isolation and characterization of shiga toxin-producing Escherichia coli (STEC) in food

Shigatoxin-producing E. coli (STEC) and their subgroup, the enterohaemorrhagic E. coli (EHEC) are known since about 25 years. Only EHEC can cause diseases in humans. A molecularbiological method cascade is described in this article. It contains the following steps: preenrichment, enrichment, preparation of samples (DNA extraction), screening PCR, specific STEC isolation by using DNA DNA hybridization, verification of isolates as STEC by using PCR, characterization of STEC isolates. Pulsed field gel electrophoresis and DNA sequencing are additional methods for the detection of clonal correlations. They should be performed in special labs, only. Furthermore, some information about an interlaboratory ring trial within Germany and some conclusions are given.     

Methods for checking up screening tests for detection of shiga toxin producing Escherichia coli (STEC) using PCR

Methods for making check ups of STEC-Screening-PCRs by using internal and external systems are given. A control-DNA with identical primer binding sites in relation to the target-DNA (stx-genes) was produced and by HPLC-chromatography quantified. It was used in an internal system. But we had to use too high concentrations of this DNA for making the check up in each sample. A false negative result could be shown in samples with a low stx content as a result of this coamplification process. Therefore we used an external system. A control-DNA with heterologous primer binding sites in relation to the stx-screening-PCR and the special primers were given in each vessel. The optimized system was used for making investigations of some faecal samples from cattle.     

Follicular Dynamics in Heifers during Pre-pubertal and Pubertal Period Kept under Two Levels of Dietary Energy Intake

The objective of this study was to characterize follicular dynamics in pre-pubertal, pubertal and post-pubertal periods, as well as the effect of high-energy intake on follicular development and age at puberty in heifers. Thirty-one Nelore (Bos indicus) heifers, 6 months old, were randomly assigned to receive two different diets: one of low (GI) and other of high dietary energy intake (GII). Animals were evaluated in relation to body weight gain by being weighed every 21 days. Heifers were evaluated every other day by real-time linear ultrasonography to characterize ovarian structures development from weaning to post-pubertal period. Blood samples were collected to determine plasmatic concentrations of progesterone by RIA method. The ovulation was determined when progesterone concentrations were >1 ng/mL in three consecutive samples, and by ultrasound images of corpus luteum; and oestrous behaviour in some animals. Age at puberty differed among heifers of GII (17.00 +/- 0.46 months) compared with heifers of GI (19.87 +/- 0.47 months; p < or = 0.05). Maximum size of the dominant follicles at pre-pubertal period was greater in GII heifers than in GI (10.52 +/- 0.33 and 9.76 +/- 0.15 mm, respectively; p < or = 0.05). As heifers approached first ovulation time, size of dominant follicle increased (11.75 +/- 0.37 mm for GI and 12.52 +/- 0.91 mm for GII; p < or = 0.05). Body weight at puberty was not different in both groups (302.33 +/- 27.31 kg for GI and 326.19 +/- 27.78 kg for GII heifers; p > 0.05). We conclude that animals receiving high dietary energy intake attained the puberty earlier and the development of follicles were different than in low dietary energy intake.     

Diversity among isolates of Actinobacillus equuli and related organisms as revealed by ribotyping

Objective The objective of this work was to examine the diversity within Australian isolates of Actinobacillus equuli and related organisms by the genotypic method of ribotyping.
Design Ribotyping, performed using the enzyme Hae III, was used to examine the diversity in 12 field isolates of A equuli (five being capable of fermenting L-arabinose), one field isolate of Pasteurella caballi and two unclassifiable field isolates. Isolates were obtained from Australian horses, except for three isolates of A equuli (one L-arabinose positive and two L-arabinose negative) which were obtained from horses and a pig in Africa. In addition, the type strains for A equuli and P caballi and a reference strain for Bisgaard Taxon 9 were included in the study.
Results The ribotype patterns were analysed by computerised cluster analysis, yielding five clusters (A to E). All five of the L-arabinose positive A equuli were assigned to cluster A, with all the other seven A equuli isolates (all L-arabinose negative) and the type strain being assigned to cluster B. One of the two unclassified isolates formed cluster C along with the reference strain for Bisgaard Taxon 9. The remaining unclassified isolate formed cluster D. Cluster E consisted of the field isolate and reference strain of P caballi .
Conclusion The results of this study indicate that A equuli is a diverse species, with L-arabinose positive isolates of A equuli being quite distinct from typical L-arabinose negative isolates. Ribotyping appears to be a useful tool in confirming the identity of A equuli -like organisms from horses.