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1.
The traditional diagnostic test for Tritrichomonas foetus involves collection of preputial or vaginal samples followed by culture in a growth media and microscopic examination. Recently, polymerase chain reaction (PCR) techniques have been described for use as a diagnostic assay. The objective of this study was to evaluate a previously described PCR assay for detecting T. foetus in cultured preputial material. The detection limits of the assay for T. foetus organisms in a growth medium, in samples prepared from washing microscope slides, and in preputial material cultured in a growth medium were determined. Preputial samples were collected from 13 bulls uninfected with T. foetus. The PCR assay was able to detect 5 T. foetus organisms in the growth medium and the cultured preputial material. Amplification products were obtained from samples prepared from washes of microscope slides containing as few as 3 visualized organisms. The PCR assay was able to detect organisms in culture at a lower concentration than was possible by direct microscopic examination. This low detection limit may allow the PCR assay to be used to enhance the sensitivity of the current diagnostic test. In addition, the assay could be used to confirm the identification of T. foetus organisms observed by direct microscopic examination when other confirmation techniques, such as staining and phase microscopy, are not practical.  相似文献   

2.
Preputial scraping samples from 305 mixed breed beef bulls were examined for the detection of Tritrichomonas foetus infection. All samples were collected by veterinarians and transported in commercial media to an accredited lab. Upon arrival samples underwent microscopic examination for the presence of Tritrichomonas foetus and were then incubated until 5 days postcollection before final microscopic examination. Culture detected 14 samples with Trichomonad spp.; all were confirmed to be Tritrichomonas foetus by polymerase chain reaction (PCR). After final examination samples were randomly placed in groups of 5 samples; technicians were blinded as to culture results of the individual samples constituting each pool. From each sample within a group, a portion of the fluid sediment was removed and pooled with the other samples of the group to form 61 pools. From each of the formed pools an aliquot was removed for PCR. PCR detected 16 positive pools; an additional 2 positive samples were then identified on individual PCR on samples previously diagnosed as culture negative. Relative to culture, the 95% confidence intervals for sensitivity and specificity of PCR pools to detect Tritrichomonas foetus were 76.8% to 100% (mean value: 100%) and 85.5 to 99.5% (mean value: 93.4%), respectively.  相似文献   

3.
The diagnostic test for Tritrichomonas foetus in bulls is microscopic examination of cultured preputial samples. Trichomonads other than T. foetus can be present in a preputial sample. Both a staining technique and a polymerase chain reaction assay were useful in differentiating between T. foetus and another trichomonad observed in samples from virgin bulls.  相似文献   

4.
Efficacy of ipronidazole against trichomoniasis in beef bulls   总被引：3，自引：0，他引：3  
Preputial smegma samples from 195 beef bulls were collected repeatedly and cultured for Tritrichomonas foetus. Seventy-five (38.5%) of these bulls were positive for trichomonads on at least 1 culture. Sensitivity of the culture procedure (number of positive cultures/number of total cultures from known-positive bulls) was 81.6%. Storage of preputial smegma in lactated Ringer's solution at 5 C for 24 hours resulted in a 14% loss of sensitivity. Seventy-three of the 75 infected bulls were available for treatment and were alloted randomly to 2 groups. Bulls in both groups were treated with procaine penicillin (7,000 IU/kg, IM) for 2 days before ipronidazole treatment. Thirty grams of ipronidazole powder was dissolved in 60 ml of sterile water, and was given IM to group 1 bulls. Group 2 bulls were given a similar 30-g ipronidazole solution IM on day 1, and were given 15 g of ipronidazole dissolved in 30 ml of sterile water on days 2 and 3. Efficacy of treatment (ie, negative cultures of preputial smegma for trichomonads for 6 consecutive weeks after treatment) was 92.8% for the 42 bulls treated once and 100% for the 31 bulls treated 3 times.  相似文献   

5.
OBJECTIVE: To compare sensitivity for diagnosing Tritrichomonas foetus infection in bulls using 2 sampling tools and to calculate negative predictive values for infection. DESIGN: Randomized clinical trial. ANIMALS: 30 Bos taurus bulls naturally or experimentally infected with T foetus. PROCEDURE: Preputial scrapings were obtained once/wk for 6 weeks using an artificial insemination pipette and a metal brush; which tool was used first for each bull was randomly determined. Samples were collected first from the left side of the prepuce and then from the right side and placed in commercially available transport media chi 2 Values and confidence limits were adjusted for effect of clustering of results by bull. RESULTS: Significant differences in sensitivity of results were not found between samples collected using the brush or pipette. Using the pipette, sensitivity was estimated to be 91.6% (95% confidence interval, 84.3 to 95.7%); negative predictive values ranged from 41 to 99% for prevalence of infection of 90 to 5%, respectively. Sensitivity was 88.8% for first sample obtained and 96.1% for second sample obtained. CONCLUSIONS AND CLINICAL RELEVANCE: Collection of preputial scrapings with an artificial insemination pipette or a metal brush and use of a commercially available culture system can provide a sensitive diagnostic test for T foetus infection in bulls. Calculated negative predictive values indicated that 1 or 2 tests would suffice in most clinical situations. For bulls from herds in which T foetus is endemic, 2 to 4 tests/bull may be required to ensure that each bull is not infected.  相似文献   

6.
Three different methods of collecting preputial material for bacteriological examination were compared using 3 bulls infected with Campylobacter fetus subsp. fetus. The first method utilised a specially designed instrument to scrape the preputial and penile mucosa, int he second method plastic pipettes were used to aspirate material and the third method involved washing the preputial cavity with sterile peptone water. Bacteriological examination of the samples showed conclusively that scraping was the method of choice because more C. fetus positive samples were identified and there was less interference from contaminating organisms.  相似文献   

7.
The current diagnostic test for Tritrichomonas foetus involves the culture of collected preputial or vaginal samples. In an earlier study, which evaluated sampling tools for use with bulls, it was observed that the sensitivity of the diagnostic test was higher for 2nd samples collected from the right side of the prepuce than it was for samples collected 1st from the left side. The study described in this paper was conducted to evaluate which of these factors was responsible for the effect on diagnostic sensitivity. Twenty-nine bulls infected with T. foetus were repeatedly sampled in a 2-factor cross-over design. Samples taken from the right side of the prepuce were 4 times as likely to be positive as samples taken from the left side (P = 0.03). Other factors did not have a significant effect on the outcome of the diagnostic test. Unexpected factors may affect the sensitivity of the diagnostic test for T. foetus.  相似文献   

8.
Preputial exudates were collected from 3 bulls infected with Tritrichomonas foetus by scraping the mucosa with a specially designed instrument and by aspiration. For diagnostic purposes the scraping method was superior direct microscopic examination but both methods were equally good when the samples were cultured within 2 hours of collection. The organism remained viable in a transport medium for 24, 48 and 72 hours showing a lineal decrease in viability with time which was more than 3 times greater in samples aspirated than in samples scraped.  相似文献   

9.
Experimental infection of the reproductive tracts of heifers and bulls with Tetratrichomonas sp. isolated from preputial smegma of virgin bulls was attempted. Nine heifers and four bulls were challenged by inoculation of 7 x 10(6) Tetratrichomonas sp. into the vaginal lumen and preputial cavity, respectively. Vaginal mucus and preputial smegma samples were collected and cultured for Tetratrichomonas sp. Heifers were slaughtered in groups of three at 2, 9 and 21 days after inoculation. Two heifers and two bulls infected with Tritrichomonas foetus and two uninfected heifers were used as controls for the model infection. Tetratrichomonas sp. were only isolated in vaginal mucus of 7/9 inoculated heifers at 6h post-inoculation, and genital secretions taken at slaughter time from vagina, uterus and oviduct were cultural negative. Bulls challenged with Tetratrichomonas sp. remained cultural negative. Since Tetratrichomonas sp. survived only a few hours in the female genitalia and did not survive in the male genitalia after experimental challenge, Tetratrichomonas sp. did not colonize the genital tract. These were likely trichomonads from the digestive tract. Collection of clean samples without fecal contamination from the reproductive tract is proposed as a measure to avoid Tetratrichomonas sp. transitory genital infection.  相似文献   

10.
Portions of penis and prepuce were collected from 24 bulls with current or recent Tritrichomonas foetus infection. Epididymides were collected from seven of the bulls, and seminal vesicles and prostate were collected from four. Following immunohistochemical staining with two monoclonal antibodies (34.7C4.4 and TF1.15) prepared against T. foetus surface antigens, trichomonads were identified in sections from 15 of the bulls. Organisms were most often located in penile crypts in the midshaft and caudal regions and less often in preputial crypts. Trichomonads were not observed in sections from other genitalia or in subepithelial tissue. T. foetus antigen, however, was present in the cytoplasm of some epithelial cells and the cytoplasm of some mononuclear cells in subepithelial lymphoid aggregates and follicles. Preputial smegma was collected from 16 T. foetus-infected bulls and from 16 control bulls with negative T. foetus cultures. Preputial antibody levels to TF1.17, a surface antigen of T. foetus, were determined by an enzyme-linked immunosorbent assay. Preputial secretions from infected bulls contained specific antibody of each isotype and subisotype tested. IgG1 responses were the greatest, IgM and IgA responses were approximately equal, and IgG2 responses were low. Each isotype and subisotype response in infected bulls was significantly greater than that in the controls. These results confirm previous speculation concerning anatomical sites of infection and suggest that parasite antigen can be taken up and processed locally, resulting in deposition of specific IgG1, IgG2, IgA, and IgM antibodies in the preputial cavity.  相似文献   

11.
Accurate identification of the bovine pathogen Tritrichomonas foetus is sometimes complicated by the presence of other trichomonadid protozoa in clinical samples. A highly specific and reproducible approach for differentiating 3 common types of bovine trichomonadid protozoa found in the bovine preputial cavity, T. foetus, Pentatrichomonas hominis, and a Tetratrichomonas species, was developed using polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis. Universal trichomonadid protozoa primers, TFR1 and TFR2, were used to amplify the 5.85 rRNA gene and internal transcribed spacer regions (ITSRs), and the products were digested with the restriction enzyme HpyCH4IV. Restriction fragment length polymorphism analysis was performed on 55 trichomonad isolates from bovine preputial washing and scraping samples. The RFLP results correlated 100% with 5.85 rRNA gene and ITSR sequence resultsand PCR results with primers specific for T. foetus. The results of this study demonstrate that PCR and RFLP analysis can be used in lieu of DNA sequencing to identify the specific trichomonadid protozoa isolated from the bovine preputial cavity.  相似文献   

12.
Tritrichomonas foetus is a serious veterinary pathogen that causes bovine trichomonosis, a sexually transmitted disease that eventually leads to abortion and infertility. T. foetus has a simple life cycle that consists of only a trophozoitic form. During unfavorable environmental conditions, the trophozoites, which are polar and flagellated, can adopt a spherical shape and internalize their flagella. These rounded organisms are known as pseudocysts. Although it is currently assumed that T. foetus pseudocyst formation is reversible and that it represents a response to stressful conditions, there are no reports showing the presence of this form in vivo. For this reason, the aim of this study was to verify whether T. foetus pseudocysts are encountered in naturally infected bulls. Towards this goal, fresh preputial samples obtained from seven mature bulls that were naturally infected with T. foetus were analyzed using complementary techniques, such as video microscopy, fluorescence microscopy, scanning and transmission electron microscopy. The analyses revealed that approximately 55% of the parasites were in pseudocyst form in each preputial sample, whereas approximately 25% of T. foetus displayed pear-shaped bodies. Previous research demonstrated that in vitro T. foetus pseudocysts are able to divide by a budding process. Here, this division mode was observed in approximately 20% of fresh T. foetus obtained from preputial bovine samples. Thus, this study shows that in infected bulls, pseudocysts are present and occur more frequently than the pear-shaped parasites.  相似文献   

13.
A number of different culture media have been described for use in the diagnosis of Tritrichomonas foetus infection in bulls, and recently, a commercial culture kit has become available. The objective of this study was to compare the sensitivity of 2 culture-based diagnostic tests for T. foetus in bulls. One test used a commercial kit for transport and culture of the samples. The other test used a thioglycollate transport medium (TFTM) for transport and a modified Diamond's medium (MDM) for culture of the samples. Twenty-one bulls infected with T. foetus were sampled repeatedly. On each sampling day, samples collected from the left and right sides of the bull were tested with one of the 2 diagnostic tests being compared. The effect of the type of diagnostic test on the outcome of the test was evaluated with a chi-square test for the calculated odds ratio. Because repeated tests from the same bull cannot be considered independent measures, unadjusted chi-square tests were adjusted for the effect of clustering by bull. Samples tested using the commercial kit were 6.95 times as likely to be positive as samples tested with a diagnostic test using MDM (P < 0.001).  相似文献   

14.
Tritrichomonosis is a widespread, economically important venereal disease caused by Tritrichomonas foetus. The traditional diagnosis of this disease, which causes infertility and abortion in cattle, is based on the culture of the parasite. This process is time consuming, has low sensitivity, and is prone to contamination with intestinal or coprophilic trichomonadid protozoa, resulting in false positive diagnostics of T. foetus. In order to avoid the shortcomings of the traditional method, we developed a simple PCR assay based on TFR3 and TFR4 primers, which does not require parasite culturing. The sensitivity of the PCR assay resulted comparable to that of the classical method, being able to detect as few as five T. foetus parasites. In addition the method is highly specific. The analysis of preputial fluid washing samples showed that 58 out of 203 samples were positive by both, the PCR and the culture method (+/+), 9 samples were positive by PCR and negative by the traditional method (+/-) and only one sample resulted negative by PCR and negative by culture (-/+). The samples for the PCR assay can be stored for a week at 4 degrees C or 72h at room temperature. In summary, our study demonstrated that the PCR assay is an effective method for the diagnosis of T. foetus from preputial samples, and that it compares advantageously to the classical method.  相似文献   

15.
Prevalence of trichomoniasis among California beef herds   总被引：5，自引：0，他引：5  
Sixty cow-half herds of more than 50 cows each were randomly selected for a prevalence survey of bovine trichomoniasis in California. Herd size, as judged by the number of bulls, ranged from 1 to 210 bulls (median = 8; mean = 59 +/- 15.8). Preputial smegma was collected from 729 bulls (median = 6 bulls/herd) and cultured for Tritrichomonas foetus. Of 57 herds from which samples were collected, 9 (15.8%) had at least one infected bull. Of the 729 bulls from which samples were cultured, 30 (4.1%) were infected. Correcting for sensitivity of the diagnostic test yielded a prevalence of 5.0%. Infection rates for bulls greater than 3 years old and less than or equal to 3 years old were 6.7% and 2.0%, respectively (P less than 0.025). Median herd sizes were 14 bulls (range, 6 to 114) for infected herds and 7 (range, 1 to 210) for uninfected herds. These findings suggest that trichomoniasis is common in California beef herds. Because several bulls less than 4 years old were infected, we suggest that control measures stressing replacement of older bulls with younger ones should be combined with diagnostic procedures in those younger replacements, to ensure that they are not already infected.  相似文献   

16.
Accuracy of culture for diagnosis of Tritrichomonas foetus was investigated in 2832 naturally exposed range beef bulls from 124 herds. Preputial fluid samples were inoculated into the culture medium, incubated at 37 degrees C, and daily examined. Diagnostic test was evaluated using Bayesian techniques to estimate sensitivity and specificity without a gold standard. Median posterior test sensitivity was 72.04% (95% probability interval: 58.07-86.38%) and specificity was 95.37% (95% probability interval: 94.07-96.65%). Low diagnostic test accuracy may have resulted from host and/or diagnostic test procedure related factors. Under natural range conditions, more accurate methods for T. foetus diagnostic and repeated preputial samplings of bulls may be necessary on trichomonosis control programs.  相似文献   

17.
The immune response in cattle infected with Tritrichomonas foetus   总被引：4，自引：0，他引：4  
Holando-Argentina calves (males and females) were experimentally infected with Tritrichomonas foetus var. Belfast (T. foetus) by introducing 10(7) protozoa into the preputial and vaginal cavities, in order to analyse the course of the immune response to infection. Samples of serum, vaginal mucus and preputial secretion were taken periodically and assayed by means of microagglutination of living protozoa. The serum antibody titre, which averaged 32 before infection and was equivalent to titres in a non-infected group, increased to 512 in the heifers 11 weeks later and to 128 in the bulls 4 months post-infection. Agglutinating antibodies were not detected in the preputial cavity, but heifers showed antibodies in the vaginal mucus and became trichomoniasis free after 4 months. Conversely, genital secretions from the bulls gave rise to positive cultures during the whole period of experimentation. The intradermal sensitivity was checked using a soluble antigen from T. foetus. The diameter of the papula increased up to three times in heifers, while in bulls the results were no different than those from the non-infected group. Serum antibodies were of the IgG2 subclass, while those isolated from vaginal mucus were characterized as IgG1, an opsonizing antibody. Heifers were refractory to challenge infection after 1 year. The poor immune response in bulls is consistent with their role as carriers of T. foetus.  相似文献   

18.
We present observations on an unusual tetratrichomonad species isolated from preputial smegma of virgin bulls. Ultrastructural studies were performed using scanning and electron microscopy techniques. This protozoan presents four anterior flagella of unequal length and a recurrent one forming the undulating membrane. It shows one anterior nucleus, a Golgi complex, an axostyle, and a costa. The hydrogenosomes are rather elongated, seen in groups, and presenting different electron densities. Vacuoles of different sizes containing bacteria and material in process of digestion were frequently found. PCR was also used in order to compare the species herein described with other trichomonad species. The amplification products were seen only with primers TFR1 and TFR2 (specific to trichomonads), but not with TFR3 and TFR4 (specific to Tritrichomonas foetus), suggesting that although collected from the genital tract of the bull, this protist was not T. foetus. We propose that the appearance of these tetratrichomonads were probably due to the sodomy practiced among bulls. Concomitant contamination of preputial cavity with feces could explain the presence of the opportunistic organism. The observations presented here show the importance of the correct diagnostic when investigating samples obtained from the urogenital tract of cattle. We also suggest that this flagellate belongs to the species Tetratrichomonas buttreyi.  相似文献   

19.
Bovine trichomonosis (BT) and bovine genital campylobacteriosis (BGC) are sexually transmitted diseases that can be important infectious causes of reproductive failure in extensively managed beef cattle where natural mating is a common practice. However, their prevalence in Europe was thought to be insignificant or very low. The purpose of this study was to investigate the prevalence and risk factors associated with BT and BCG in a representative beef cattle breed, Asturiana de la Monta?a (AM), which is usually managed extensively in the mountain areas of Northern Spain and putative risk factors associated with the two diseases are present on most farms holding AM cattle. Preputial smegma samples were collected from 103 bulls belonging to 65 herds. Pathogen detection was undertaken using culture and PCR. Two scraping methods for sample collection (AI pipette and plastic scraper), as well as different culture media and DNA extraction methods were evaluated on field samples. Campylobacter fetus veneralis infection was not detected in any animal in any herd. However, Tritrichomonas foetus infection was demonstrated in 32% (33/103) and 41.5% (27/65) of bulls and herds tested, respectively. AM bulls older than 3 years (39.7%) were more likely to be infected than young bulls (16%) (OR=3.45, CI=1.07-11.19). An increase in repeat breeder cows was reported in herds from which T. foetus was detected (OR=5.2, CI=1.5-17.18). These findings highlight the re-emergence of this disease in extensively managed beef cattle in Spain. For routine diagnosis, the use of a culture technique and PCR in combination is advisable for testing smegma samples under field conditions.  相似文献   

20.
Preputial fluids from 567 virgin Angus and Hereford bulls, 1-2 years old, were inoculated into Sutherland medium, and approximately 8.4% produced cultures with a protozoan suggestive of Tritrichomonas foetus. Under brightfield microscopy, large numbers of single-celled motile organisms with multiple anterior flagellae, a posterior flagellum, axostyle, and a visible undulating membrane were detectable. Motility was jerky and rolling, as described for T. foetus. Air-dried smears of cultures stained with Giemsa or Diff-Quick + iodine revealed an organism similar to T. foetus, although somewhat more rounded. Several organisms appeared to have four anterior flagellae. Scanning electron microscopy (5000x) of representative samples revealed four anterior flagellae on most organisms, and an axostyle that was consistently longer than that seen in T. foetus. Using pan-trichomonal primers and T. foetus-specific primers in a polymerase chain reaction (PCR) assay, amplification products of 372bp were detected in all virgin bull isolates, but only with the pan-trichomonal primers. Positive control isolates of T. foetus yielded amplification products of the expected size (372 and 347bp) with the two sets of primers, respectively. We conclude that these protozoa are not T. foetus, and note the similarity of these findings with those reported earlier in North American beef cattle. Because in several countries there is no legal treatment for bovine trichomonosis, veterinarians recommend slaughter of bulls with positive preputial cultures. The existence of easily mis-identified non-T. foetus trichomonads in the bovine prepuce suggests that the current "gold standard" diagnostic test (culture of preputial scrapings or washings) should be augmented with a more specific confirming test, such as the PCR employed in this study.  相似文献   

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