Received date: February 24, 2015; Accepted date: February 26, 2015; Published date: March 06, 2015
Citation: Gaur K, Nigam R (2015) Pooled Conventional PCR to Detect Tritrichomonas Foetus Infected Beef Bulls. J Biodivers Endanger Species 3:148. doi:10.4172/2332-2543.1000148
Copyright: © 2015 Gaur K, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
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Tritrichomonas craniate may be a production and restrictive concern for beef producers within the Western USA. Historically preputial scrapings are collected, refined in enriched media, and examined microscopically. PCR techniques are a unit currently getting used extensively to verify culture results or as a complete take a look at the organism. This technology offers the advantage of identifying the infective Tritrichomonas foetus organism from different non infective dirty organisms. The samples were collected, cultured, evaluated and analysed by qPCR.
Tritrichomonas; PCR; Beef; Foetus
Tritrichomonas vertebrate (T. foetus) may be a single cell parasite related to Cupid's itch in cattle. The organism produces abortion, sterility, prolonged birth intervals, and pyometra in infected cows . The organism infects the prepuce of the bull and is transmitted to prone females throughout copulation [1-5]. Most frequently the feminine are going to be solely transiently infected, and in a matter of four to five months the infection is cleared and she or he is once more reproductively sound, yet remains liable to re-infection at a later time. For a long period of time, the female remains infected after giving birth to a calf through normal delivery [1,3,5]. All regulative efforts address the suitable testing to work out the presence of infection in bulls. Identification of infection within the female and also the movement of infected feminines aren’t considerably controlled or monitored.
Diagnosis of T. foetus is predicated on clinical signs, history, and laboratory detection of the organism, so as to substantiate a herd is infected with T. foetus it's necessary to demonstrate the organism or it is desoxyribonucleic acid. This can be done by direct microscopic examination, culture followed by microscopic examination and PCR of Preputial Scrapings (PPS) or cervical mucus [1,5,6]. All states through the eastern border of Kansas have some form of T. foetus regulation, and recently states east of that line have developed regulations addressing T. foetus including Missouri, Arkansas and Louisiana. Culture is proscribed by the inherent lack of environmental stability of the organism, collections made or maintained at low temperatures, exposed to direct daylight, microorganism and plant contamination of culture media, and changes in pH might cause a loss of viability and end in failure to observe the organism [7,8] and as the organism dies in the culture media it degrades and the by-products of the degradation results in a breakdown of DNA of the organisms.
Another disadvantage to culture is that there are very few individuals to differentiate the nuances between T. foetus and other Trichomonad spp. and diagnoses require a confirmation through the use of PCR. Potential sample degradation supports to doing PCR is very important that waiting until the highest of the traditional culture quantity may finish in inadequate amount of deoxyribonucleic acid for diagnostic functions. PCR is costly and may result in false positives. Two types of DNA studies are being employed, gel based PCR and qPCR, both methods require the extraction of the T. foetus DNA, the only difference between the two methods is the how the DNA is detected after extraction and the PCR has occurred. Gel based PCR relies on detecting a band at the appropriate base pair level versus qPCR that uses a fluorescent probe, optical density, and cycle threshold (Ct) to detect the presence of the organisms DNA with the Ct being the determining factor on a sample being called positive or negative.
Conventional PCR and/or pooling might minimize the number of false positives. Factors contributing to false positive or false negative PCR results are contamination, false priming sites, and inhibitors. In the case of comparing individual cultures to the pooled result again the false positives associated with the presence of non T. foetus trichomonads was avoided without the cost of individual PCR testing. Conventional PCR and/or pooling might minimize the number of false positives. Factors contributing to false positive or false negative PCR results are contamination, false priming sites, and inhibitors. In the case of comparing individual cultures to the pooled result again the false positives associated with the presence of non T. foetus trichomonads was avoided without the cost of individual PCR testing. The comparison of pooled conventional PCR to individual culture adds a comparison of pooled conventional PCR to individual conventional PCR and also compares pooled conventional PCR to individual qPCR.
Pooled PCR offers the advantage of a lower cost to screen herds for T. foetus. PCR is an expensive alternative to traditional culture, a method to lower the cost of using PCR is to pool samples. Pooling minimizes the potential effects of these factors without any significant impact on diagnostic sensitivity. The inverse relationship between diagnostic sensitivity and diagnostic specificity when viewed in light of the high analytical sensitivity creates a potential for an increase number of false positive tests.