A specific DNA probe which identifies Babesia bovis in whole blood.

A genomic library of Babesia bovis DNA from the Mexican strain M was constructed in plasmid pUN121 and cloned in Escherichia coli. Several recombinants which hybridized strongly to radioactively labeled B. bovis genomic DNA in an in situ screening were selected and further analyzed for those which specifically hybridized to B. bovis DNA. It was found that pMU-B1 had the highest sensitivity, detecting 25 pg of purified B. bovis DNA, and 300 parasites in 10 microliters of whole infected blood, or 0.00025% parasitemia. pMU-B1 contained a 6.0 kb B. bovis DNA insert which did not cross-hybridize to Babesia bigemina, Trypanosoma evansi, Plasmodium falciparum, Anaplasma marginale, Boophilus microplus and cow DNA. In the Southern blot analysis of genomic DNA, pMU-B1 could differentiate between two B. bovis geographic isolates, Mexican strain M and Thai isolate TS4. Thus, the pMU-B1 probe will be useful in the diagnosis of Babesia infection in cattle and ticks, and in the differentiation of B. bovis strains.     

Comparison of dot blot hybridization, polymerase chain reaction, and virus isolation for detection of bovine herpesvirus-1 (BHV-1) in artificially infected bovine semen.

Bovine semen samples spiked with bovine herpesvirus 1 (BHV-1) were used to compare dot blot hybridization, polymerase chain reaction (PCR), and virus isolation for detection of BHV-1 in bovine semen. The PCR amplification used primers targeting the BHV-1 thymidine kinase gene and a nucleic acid releasing cocktail (GeneReleaser); the PCR product was used as the DNA probe in dot blot hybridization; virus isolation was done in primary bovine fetal testis (BFT) cell cultures. Semen diluted 1:20 in tissue culture medium had the least cytotoxicity and inhibition of viral cytopathic effects in BFT cells, allowing detection of 1 TCID50/100 microL of BHV-1 suspension by virus isolation. The presence of foreign DNA such as bovine sperm DNA or salmon sperm DNA increased the sensitivity of dot blot hybridization in detecting BHV-1, allowing detection of 20,000 TCID50/100 microL of neat semen. The inhibition of PCR amplification of BHV-1 DNA in bovine semen was eliminated by diluting the samples 1:20 in tissue culture medium. The best PCR amplification was obtained when semen was diluted 1:20 and when a reaction buffer of pH 9.0, with 1.0 mM MgCl2 was used. Under these conditions, the PCR followed by ethidium bromide staining of agarose gels could detect 1 TCID20/100 microL of sample, whereas PCR followed by Southern blot hybridization could detect 0.01 TCID50/100 microL of sample.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dynamics of changes in the cytological picture of vaginal smears and circulating ovarian hormones during the puerperal period in ewes]

A postparturient period is characterized by low basal secretion of adenohypophysis gonadotropins with the following appropriate changes in ovarian hormones and their response to the morphology of vaginal epithelium. In this study the dynamics of the cytological picture of vaginal swabs and ovarian hormones 17 beta-oestradiol (E2) and progesterone was investigated in the puerpery of ewes. The objective was to obtain and extend the knowledge of cytological changes in vaginal epithelium and levels of ovarian hormones of ewes after parturition and of their relationships from the first several days after lambing until the 51st day of the period of observation. Vaginal swabs for vaginal cytology were taken from nine ewes on days 1, 4, 7, 14, 17, 21, 25, 34, 42 and 51 after parturition. These swabs were fixed in ether-alcohol 1:1, stained according to the Faltínová-Zidovsky method, embedded in Canada balsam and evaluated by differentiation of cells according to Luksh (1953). Blood samples for E2 and P4 determinations were taken from the jugular vein in the same intervals as vaginal swabs. The serum was centrifuged and stored at -18 degrees C until use. E2 and P4 concentrations were determined radioimmunologically, using kits RIA-test ESTRA and RIA-test PROG from URVJT Kosice. A statistically significant decline (P < 0.05) of percentual representation of basal and parabasal cells (Fig. 1, Tab. I) on day 7 after lambing was replaced by their multiplication from day 14 reaching the values of 66.07 +/- 3.95 on day 42. A statistically significant decrease in intermediary flat cells (Fig. 2, Tab. II) was observed on days 14 (P < 0.001), 34 and 42 (P < 0.01; P < 0.001), in comparison with the first day after lambing. An evaluation of intermediary convoluted cells revealed their highest percentage on days 1 and 17 after parturition (34.65 +/- 4.77-20.62 +/- 12.57) and their decline to values in the range of 6.77 +/- 1.46-7.66 +/- 2.25 on the remaining days of the period of observation. Percent occurrence of superficial flat cells (Fig. 3, Tab. I) ranged from 3.9 +/- 1.10 to 10.63 +/- 7.23 from day 1 to day 51 after lambing. The lowest percentual representation (1.32 +/- 0.79-4.10 +/- 1.89) was recorded for superficial convoluted cells. Multiplication of the evaluated cells was observed, reaching the highest but insignificant representation (P > 0.05) on day 25 of postparturient investigation: 4.10 +/- 1.89 (Fig. 3, Tab. I). 17 beta-oestradiol (E2) concentrations were compared to the -1st day before parturition, when its values varied at the level of 2.45 +/- 0.64 nmol/l serum.(ABSTRACT TRUNCATED AT 400 WORDS)     

Enzootic abortion in a goat herd, caused by mixed infection with Coxiella burnetii and Chlamydia psittaci. Case report]

We report on an outbreak of enzootic abortion in a herd of goats with high losses caused by mixed infection with Coxiella burnetii and Chlamydia psittaci. Both agents could be detected by microscopic investigation of smears from afterbirths by Giménez staining and by a capture enzyme-linked immunofluorescence assay (Capture ELIFA). Additionally, Coxiella burnetii was isolated from the same tissues in BGM cell cultures. Serological investigations with CF, indirect ELIFA, and IF tests revealed high antibody titers against both agents in most animals. Coxiella antigen was still detected 91 days later in milk samples of two infected goats by Capture ELIFA.     

Differential effects of heparin on NO and tumor necrosis factor-alpha production in bovine blood mononuclear cells stimulated with Salmonella typhimurium lipopolisaccharide.

We investigated the influence of heparin, one of the extracellular matrix (ECM) components, on nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) production by bovine peripheral blood mononuclear cells (PBMC) and monocytes left to adhere for 2 (freshly adherent monocytes) and 48 h (resting monocytes), activated with Salmonella typhimurium lipopolysaccharide (LPS). After 24-h stimulation with LPS, heparin (100 microg/ml) increased (by about 40%) NO production by peripheral blood mononuclear cells and by freshly adherent monocytes. However, it did not change NO synthesis by the resting monocytes. Unlike its influence on NO level, heparin diminished TNF-alpha production by PBMC and monocytes stimulated with LPS. Microscopical examination of PBMC stained with biotin-labeled heparin, showed that both lymphocytes and monocytes were able to bind this glycosaminoglycan. We suggest that heparin, as a component of ECM, modulates the early response of monocytes to exogenous stimuli.     

The effect of repeated administration of carbetocin (Depotocin, Spofa) in the first days postpartum on levels of thyroxine, triiodothyronine, 17 beta-estradiol, progesterone and on fertilization]

Application of new procedures in the sphere of the control of sexual functions requires an extension of present knowledge of postparturient endocrinium or endogenic factors comprised in postparturient physiology of sexual activity. According to recent data, oxytocin, besides its uterotonic and luteolytic activity, acts as an ovarian factor in the local intrafollicular regulation of stereidogenesis and as a modulator of uterine secretion of prostaglandines. Based on present knowledge of oxytocin effects, this study was aimed at investigation of the influence of repeated carbetocin (Depotocin inj. Spofa) administration on the dynamics of changes in thyroxine (T4), triiodothyronine (T3), 17 beta-estradiol (E2), progesterone (P4) concentrations and their mutual correlations from the 36th hour till the 51st day after parturition. Simultaneous study of a possible delayed influence of applied carbetocin on conception of ewes after oestrus evocation on day 51 after lambing was carried out. Nineteen ewes of the Slovak Merino breed, lambed in the first decade of February, were assigned to the experimental (n = 9) and to the control group (n = 10). Experimental ewes were subjected to repeated postparturient carbetocin treatment at the dose 0.07 mg per animal. The first dose was applied i. m. in 24 hours, and the second in 72 hours after parturition. On day 51 oestrus was induced in nine ewes of each group by combined treatment with chlorsuperlutin (Agelin, vaginal pessaries, Spofa) and PMSG (500 I.U./animal). On the day of PMSG application ewes were housed together with rams for the period of the next six days. Samples of blood were taken 24 hours before parturition (-1st day), up to 36 h after parturition and on days 4, 7, 14, 17, 21, 25, 34, 42 and 51 after parturition. Concentrations of T4, T3, E2 and P4 were determined by commercial kits RIA-test-T4; RIA-test-T3; RIA-test-ESTRA and RIA-test-PROG (URVJT Kosice). Animal of the control group showed variations of T4 concentrations (Tab. I, Fig. 1) at the level of original values (59.4 +/- 9.69 nmol.l-1) up to the 21st day with the exception of temporary drop on day 4 and rise on day 7, insignificant compared to the -1st day. T4 concentrations of the control group displayed an intermittent increasing trend with the statistically insignificant peak after 36 h and on day 17, compared to the -1st day. After the 21st day controls revealed a sustained moderate increase while the experimental ewes displayed a decline of its concentrations until the 51st day.(ABSTRACT TRUNCATED AT 400 WORDS)     

Ejaculatory failure associated with aortic-iliac thrombosis in two stallions.

Two mature breeding stallions were evaluated because of specific ejaculatory dysfunction, and each was found to have aortic-iliac thrombosis occluding 60 to 70% of the aortic lumen. In each case, the stallion had strong libido, normal mounting, and vigorous initial thrusting. With continued exertion, thrusting became weak and dismount was awkward. Treatment aimed at maximizing sexual arousal before mounting and reducing hind limb pain and exertion during breeding allowed these stallions to continue breeding. A program of gradually increasing daily exercise also was associated with improved breeding performance during subsequent breeding seasons. In cases of specific ejaculatory dysfunction, aortic-iliac disease should be included among the differential diagnoses.     

Determination of sulfamethazine and trimethoprim in liquid feed premixes by HPLC and diode array detection, with an analysis of the uncertainty of the analytical results.

Sulfamethazine (SMZ) and trimethoprim (TMP) are antibacterials used in veterinary practice. This paper describes a method for their determination in veterinary liquid feed premixes that is based on liquid chromatography with diode array detection. Gradient elution with methanol and ammonium acetate achieved excellent separation of the two analytes within 15 min without any interference from the matrix. Absorbance of the column effluent was monitored at 264 nm for SMZ and at 230 nm for TMP. Detailed analyses of the uncertainties of determinations afford estimated expanded uncertainties of, respectively, 0.2 and 0.1 w/v % for typical SMZ and TMP concentrations of 10.7 and 2.1 w/v %, respectively. At the lower end of the calibrated range of the method, the dominant source of uncertainty is the preparation of standards and the construction of the calibration line.     

Laboratory-based surveillance of salmonellosis of humans in Germany--safety of Salmonella typhimurium and Salmonella enteritidis live vaccines]

The Paul Ehrlich Institute, Langen, in Germany has been licensed different live vaccines of S. Typhimurium and S. Enteritidis for use in the veterinary medicine since I the 90s. The Robert Koch Institute has established a lab-based surveillance system for these live vaccine strains for an evaluation of recent public health safety. Since 2000 all strains of S. Typhimurium and S. Enteritidis from humans were investigated in respect to their phage types and other vaccine markers. 3676 S. Typhimurium strains and 4489 S. Enteritidis strains mainly from Salmonellose patients were investigated after phage typing according to their auxotrophic or antibiotic resistance markers. The live vaccine strains of Zoosaloral, TAD Salmonella vacT or TAD Salmonella vacE and Salmovac SE could not be found from infections in humans.     

The effect of eradication of lice on the occurrence of the grain defect light flecks and spots on cattle hides.

The influence of an eradication programme for lice on the prevalence of light flecks and spots on cattle hides was studied in 33 dairy cattle herds during a period of two and a half years. Lice were eradicated from the main group of herds after 9 to 12 months and the quality of the hides before and after treatment was compared. Hides from slaughtered animals were collected during the study period, tanned and examined with special emphasis on the occurrence of the grain damage light flecks and spots. The prevalence of hides without light flecks and spots increased from 24.2% before treatment to 61.6% after treatment. The prevalence of hides free from the damage increased significantly in all examined anatomical regions. The improvement in hide quality was most marked in the shoulders and neck region which corresponded to the major predilection site of cattle lice. The prevalence of hides with light flecks and spots started to decrease in the first period (2-40 days) after eradication. The changes after treatment suggested that most healing process took place over a period of about 4 months. The eradication programme eliminated the seasonal variation in the prevalence of light flecks and spots which was present before treatment.