Endometrial Tissue Concentrations of Enrofloxacin after Intrauterine Administration to Mares

Endometritis in mares is a common cause of infertility. Conventional treatments of the disease have mostly been unsuccessful, so new therapeutic alternatives need to be investigated. This study evaluated the uterine disposition and plasma pharmacokinetic behaviour of a commercial formulation of enrofloxacin (EFX) given by the intrauterine (i.u.) route (2.5 mg/kg) in healthy mares. In order to evaluate the uterine inflammatory response, an initial histopathological study assessing polymorphonuclear cell infiltration was carried out in 20 mares over a 14-day period after treatment. In a second study, 6 healthy adult mares were used for the pharmacokinetic study. Samples of uterine tissue and plasma were collected from 0 to 24 h after the i.u. treatment with 5% EFX solution. Samples were analysed by conventional microbiological assay using an EFX-sensitive strain of Escherichia coli (ATCC 25922). There was a moderate but statistically nonsignificant inflammatory response following i.u. administration of either the formulation or the vehicle alone. Pharmacokinetic analysis of the uterine concentrations of EFX showed a slow and sustained depletion, with EFX remaining at concentrations above the MIC for 24 h after treatment. The area under the concentration–time curve obtained for the uterus suggested that EFX and its metabolites are specifically retained in the uterus, which is the target tissue for bacterial colonization. Neither study provided any evidence of EFX toxicity. In conclusion, these results are encouraging and suggest that EFX may be a useful local treatment in mares with bacterial endometritis.     

Development of a PCR test for the identification of Staphylococcus intermedius based on the 16S rDNA sequence

The development of a PCR assay based on the 16S ribosomal RNA gene (rDNA) sequence was carried out for the identification of Staphylococcus intermedius. Sixty-six strains of S. intermedius, 70 of Staphylococcus aureus and 2 of Staphylococcus hyicus were examined for the assay. The 16S rDNA, of which the PCR target fragment makes up 901 bp corresponding to the sequence data of the gene, was detected in all strains of S. intermedius, but it was not detected in any strains of either S. aureus or S. hyicus. These results suggest that the PCR allows a simple and precise identification of S. intermedius.     

An outbreak of enzootic bovine leukosis in upper Egypt: clinical,laboratory and molecular-epidemiological studies

In 1989, 220 Holstein Friesian cattle (212 heifers and eight bulls) were imported from Minnesota, USA, to form a closed dairy herd in Arab El-Aoumar, Assiut, Upper Egypt. In November 1996, some abnormal signs such as loss of weight, decreased milk yield, external lymphadenopathy and decreased appetite were observed on this farm. Serological screening by enzyme-linked immunosorbent assay revealed a seroprevalence of antibodies directed against bovine leukaemia virus (BLV) of 37.7% in cattle under 2 years old and of 72.8% in animals more than 2 years old. Diagnosis was confirmed by the detection of BLV proviral DNA using polymerase chain reaction with primers amplifying a fragment of the env gene. Out of 21 tested leucocyte fractions from individual animals, 15 were positive showing a BLV-specific amplicon of 444 base pairs. Analysis of the amplicons for restriction fragment length polymorphisms and DNA sequencing results allowed the isolates to be typed. Since this was the first recorded case of enzootic bovine leukosis in Upper Egypt, strict quarantine measures were adopted and all serologically positive animals in the herd were culled.     

Immunization against gnRH in mature mares: antibody titres,ovarian function,hormonal levels and oestrous behaviour

The aim of the present study was to investigate the effect of active immunization against GnRH in mature Standardbred mares (three experimental and one control mare) on antibody titres, ovarian function, hormonal levels and oestrous behaviour. The mares were individually teased with a stallion once each day. During the first part of the experiment (period I: late April until November), blood was sampled every third day during the first 3 months, thereafter once per week. In the second part of the experiment (period II: December until August), sampling was carried out every second week. Progesterone, oestradiol-17beta and LH were analysed. Rectal gynaecological examination was made with the same intervals as the blood samplings and included palpation of the genital organs and ultrasonography. The experimental mares were immunized against GnRH with a GnRH-BSA conjugate. Equimune (Vetrepharm, Bracetown, Business Park, Clonee, Co. Meath, Republic of Ireland) was used as adjuvant. The mares were immunized on four occasions (20-30 day intervals) and GnRH antibody titre was determined. All immunized mares produced antibodies against GnRH but the maximum titres as well as the duration of a greater than 10% binding capacity varied between the mares (1 : 1600 to 1 : 50 000; 5 to 12 months, respectively). After the first injection, all mares showed one oestrus and ovulated at the regular time. In two of the mares, the immunization resulted in ovarian atrophy. Their hormone levels of progesterone, oestradiol- 17beta and progesterone decreased to basal levels and the cyclical hormone pattern was interrupted from approximately 30 days onwards. They continued to show oestrous signs but with irregular durations and intervals. The third mare showed ovarian suppression only for short periods and not in both ovaries at the same time; the hormone levels were basal for only about 20 days (days 50-70) and the mare ovulated on day 75 after start of immunization. The other mares ovulated after 13.5 and 15 months, respectively. It is concluded that the effect of immunization against GnRH in mature mares was individual concerning antibody titre response and the suppression of ovarian activity and hormone levels. Mares with totally inactive ovaries continued to show oestrous signs but with irregular intervals and durations.     

Recent developments in the epidemiology of virus diseases

There is continual variation in viral epidemics regarding clinical symptoms, duration and disappearance, and the emergence of new diseases. This can be observed in both human and animal diseases. This evolution of virus diseases is mainly related to three factors: aetiological agent, host and environment. As far as genetic alterations of the virus are concerned, two major mechanisms are involved: mutations such as recombination and reassortment; and selection for resistance or susceptibility. This review focuses on the epidemiology of newly emerged virus diseases in man and animals, such as acquired immunodeficiency syndrome, haemorraghic fevers, bovine spongiform encephalopathy, canine haemorraghic disease and respiratory syndrome in horses.     

Differential serological testing by simultaneous indirect immunofluorescent antibody test in canine leishmaniosis and ehrlichiosis

A mixed indirect fluorescence antibody test (IFAT), based on cultured promastigotes Leishmania infantum and formol-inactivated suspension of cells infected with the bacteria Ehrlichia canis, was applied to make a differential diagnosis between canine ehrlichiosis and leishmaniosis. A titre greater than 80 was considered positive for antibodies to E. canis and suggestive of antibodies to L. infantum. Positive sera were titrated subsequently by serial dilutions to confirm antibodies positive to Leishmania and establishing the antibody titre of both pathogens. Fluorescence was absent with negative control sera and background staining was minimal. No serological cross-reactions between positive sera for L. infantum or E. canis were detected. Results obtained by mixed IFAT did not differ when the same serum IFAT standard was compared. The test showed equivalent sensitivity (100%). The specifities were 100% for L. infantum and 98.5% for E. canis. The equivalence in sensitivity was confirmed by calculating the correlation coefficient between IFAT standards and mixed IFAT (r>or=0.99 for both pathogens). The results of our investigations demonstrated that mixed IFAT is a specific means of establishing serological differential diagnosis of canine leishmaniosis and ehrlichiosis.     

Molecular phylogenetic analysis of Onchocerca lupi and its Wolbachia endosymbiont

The morphology of Onchocerca lupi, responsible for canine ocular onchocercosis, is unique within the genus. Earlier analyses of the 5S ribosomal RNA gene spacer region sequence of the parasite and the 16S ribosomal RNA gene sequence of its Wolbachia endosymbiotic bacteria (Rickettsiales) supported the morphological and biological arguments that O. lupi is a distinct species. However, the exact phylogenetic position of O. lupi and its endosymbiont could not be unambiguously determined. Herein we report analyses based on the mitochondrial cytochrome oxidase I (COI) gene of the filarial species and the Wolbachia surface protein (wsp) and the bacterial cell-cycle ftsZ genes of their wolbachiae. Our results indicate that O. lupi separated from other Onchocerca spp. early in evolution. This is in line with the previous morphological analysis demonstrating that O. lupi is an atypical Onchocerca species showing both primitive and evolved characters. The phylogenetic trees generated for the COI sequences of filariae and the wsp and ftsZ sequences of their wolbachiae were congruent with each other, which supports the hypothesis that nematodes and their Wolbachia endobacteria share a long co-evolutionary history.     

Avian paramyxovirus serotype 1 isolates from the spinal cord of parrots display a very low virulence

The spinal cord of 32 psittacines suffering from proventricular dilatation disease (PDD) was investigated. In six cases, a virus was isolated which upon electron microscopic examination revealed morphological details typical of members of the Paramyxoviridae. All isolates were subsequently characterized as avian paramyxovirus serotype 1 (APMV-1) by type-specific polyclonal antisera. According to their reactivity with APMV-1 specific monoclonal antibodies, the six isolates shared epitopes within the haemagglutinin-neuraminidase spike protein, distinct from pigeon-type paramyxoviruses and the LaSota vaccine strain. This grouping was further corroborated by properties of the haemagglutinin: all isolates showed a very thermosensitive haemagglutination activity and were rapid eluters. Virulence of the APMV-1 isolates in 1-day-old specific pathogen free (spf) chicken was very low, with intracerebral pathogenicity indices between 0 and 0.1. In embryonated spf chicken eggs, psittacine isolates replicated to high titres (10(8.6)-10(10.7) EID50/ml). However, they exhibited a reduced lethality over an observation time of 7 days (10(6.1)-10(8.3) ELD50/ml). In a haemagglutination inhibition test with parrot sera from birds with no history of APMV-1 vaccination, sera reacted preferentially with two isolates compared with APMV-1 vaccine strains LaSota and B1. The other four isolates exhibited a differentiated reaction pattern with the parrot sera, indicating an antigenic inhomogeneity. This is the first report of isolating very low virulent APMV-1 from neuronal tissue of parrots and implications for a possible role in slow progressing disease will be discussed.     

Prevalence of and resistance to anti-microbial drugs in selected microbial species isolated from bulk milk samples

The prevalence of strains of Staphylococcus aureus, coagulase-negative (CN) staphylococci, Listeria monocytogenes, Escherichia coli, Enterococcus faecalis, E. faecium and Bacillus cereus, was investigated in 111 bulk milk samples. Staphylococcus aureus was isolated from 38 samples, CN staphylococci from 63 samples, E. coli from 49 samples, E. faecalis or E. faecium from 107 samples, and L. monocytogenes from two samples. Bacillus cereus was not found in any of the samples and three samples were free of any of the selected species. Sensitivity to the anti-microbial drugs amikacin, ampicillin, ampicillin + sulbactam, cephalothin (CLT), cephotaxime, clindamycin, chloramphenicol (CMP), co-trimoxazole, erythromycin (ERY), gentamicin, neomycin, norfloxacin, oxacillin, penicillin, streptomycin (STR), tetracycline (TTC) and vancomycin was tested using the standard dilution technique. Minimum inhibitory concentration (MIC) characteristics (MIC50, MIC90, MIC range) were determined for each microbial species. Resistance against one or more anti-microbial drugs was found in 93% of S. aureus, 40% of CN staphylococci, 73% of E. coli, 88% of E.faecalis, 55% of E.faecium, and one L. monocytogenes strain. Most of the strains, particularly enterococci, were resistant to STR, TTC, and ERY (MIC50 4 microg/ml). A high percentage of staphylococci were resistant to beta-lactam antibiotics. High resistance to CLT was found in 11 strains of E. coli (MIC 256 microg/ml) and strains resistant to CMP (MIC90 16 microg/ml) were detected. The highest numbers of resistance phenotypes were found in E. coil (16) and CN staphylococci (12). Eighteen identical resistance phenotypes were demonstrated in indicator bacteria (E. coli, E. faecalis, E. faecium) and pathogens (S. aureus, CN staphylococci) isolated from the same bulk milk sample. The obtained resistance data were matched against the herd owners' information on therapeutic use of the drugs. This confrontation could not explain the findings of strains resistant to ERY or CMP. Our findings are evidence of selection of resistant strains among not only pathogenic agents, but also among indicator bacteria which can become significant carriers of transmissible resistance genes.     

Pharmacokinetics of gentamicin C1, C1a and C2 in horses after single intravenous dose

Gentamicin pharmacokinetics has not been studied in horses. Pharmacokinetics of gentamicin C1, C1a and C2 components following i.v. administration of total gentamicin at 6.6 mg/kg bwt to 6 healthy mature horses was determined. Significant differences in clearance, half-life (t 1/2), and mean residence time (MRT) between the gentamicin Cia and the 2 other components were found. The total body clearance (CL) of gentamicin C1a was 1.62 +/- 0.50 ml/min x kg and similar to the glomerular filtration rate (GFR) reported for horses. The CL of gentamicin C1 and C2 were 1.03 +/- 0.08 ml/min x kg and 1.10 +/- 0.15 ml/min x kg, respectively, and significantly slower than that of gentamicin C1a. The values of apparent volume of distribution at steady state were 0.22 +/- 0.05, 0.26 +/- 0.12 and 0.23 +/- 0.05 l/kg for gentamicin C1, C1a and C2, respectively. The MRT values were mean +/- s.d. 3.6 +/- 0.5, 2.7 +/- 0.3 and 3.5 +/- 0.4 h and the t 1/2 values were 3.1 (2.5-4.0), 2.4 (2.0-3.2) and 33 (2.4-4.3) h (harmonic mean and range) for gentamicin C1, C1a and C2, respectively. The MRT and t 1/2 values for gentamicin C1a were significantly shorter than those of gentamicin C1 and C2. It was concluded that the difference in pharmacokinetics between the gentamicin components has potential pharmacological and toxicological implications.