Evaluation of a PCR test for the diagnosis of Tritrichomonas foetus infection in bulls: effects of sample collection method, storage and transport medium on the test

Mukhufhi, Ndwakhulu; Irons, Pete Charles; Michel, Anita Luise; Peta, F.

doi:10.1016/s0093-691x(03)00138-9

Cited by 44 publications

(39 citation statements)

References 37 publications

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“…1,10,11 The duration of sample incubation prior to DNA extraction ranged from 24 to 48 hr between the laboratories. One report 14 has noted that T. foetus has been shown to secrete a range of hydrolytic enzymes that cause rapid DNA breakdown following cell lysis, if incubated for more than 48 hr, which may lead to a decreased sensitivity in the ability of molecular methods (e.g., PCR) to detect T. foetus DNA over longer incubation times. None of the participating laboratories exceeded 48 hr of incubation.…”

Section: Discussionmentioning

confidence: 99%

Pooling of cultured samples and comparison of multistate laboratory workflows with the MagMAX sample preparation system and VetMAX quantitative polymerase chain reaction reagents for detection ofTritrichomonas foetus–colonized bulls

et al. 2013

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The objectives of the current study were 1) to compare sample preparation workflows and quantitative real-time polymerase chain reaction assays (qPCR) as currently used in veterinary diagnostic laboratories with a study protocol utilizing commercially available reagents for individual Tritrichomonas foetus testing, 2) to assess the accuracy of pooling cultured smegma samples followed by extraction and qPCR testing as used in the study laboratory, and 3) to assess the specificity of the currently used primers and probes by sequencing all positive and presumptive positive samples identified in the study laboratory in an attempt to capture any nucleotide variability between T. foetus isolates and to rule out false-positive results possibly due to Simplicimonas moskowitzi. Eight hundred three cultured smegma samples were collected from different regions of the United States with the collaboration of 5 veterinary testing laboratories. The samples were processed individually by the respective laboratories, and then sent to the study laboratory and retested using the study protocol. Comparison testing showed an overall agreement of 95.89% between the veterinary testing laboratories and the study laboratory. One hundred seventy-six positive or presumptive positive samples plus 625 negative qPCR samples were combined and retested using a pooling protocol. Pools consisted of 1 positive sample and 4 negative samples (1/5). These pools were processed using the same study laboratory protocols, and 96% of the positive samples were detected in these pools. Nested PCR followed by sequencing confirmed 175 of the 178 samples classified as positive or presumptive positive in the study laboratory as containing T. foetus–specific DNA.

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Section: Discussionmentioning

confidence: 99%

Pooling of cultured samples and comparison of multistate laboratory workflows with the MagMAX sample preparation system and VetMAX quantitative polymerase chain reaction reagents for detection ofTritrichomonas foetus–colonized bulls

et al. 2013

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“…With the reported high analytical sensitivity of PCR, a move to using PCR as a stand-alone test has been seen with many state regulations relying on PCR in lieu of culture [10][11][12][13][14]. However, PCR is costly and may result in false positives.…”

Section: Pcr and Qpcrmentioning

confidence: 99%

Tritrichomonas foetus in Beef Bulls

2014

J Veterinar Sci Technol

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Tritrichomonas foetus is a production and regulatory concern for beef producers in the Western United States and more recently in states of the Mississippi Valley. Traditionally preputial scrapings have been collected, cultured in enriched media, and examined microscopically. PCR techniques are now being used extensively to confirm culture results or as a stand-alone test for the organism. This technology offers the advantage of distinguishing the pathogenic Tritrichomonas foetus organism from other nonpathogenic fecal organisms.

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“…Gerber et al (2010) utilizaram a PCR para a detecção de vírus, Mukhufhi et al (2003) para detecção de protozoários e Saunders et al (2007) utilizaram a técnica de reação em cadeia da polimerase múltipla (mPCR) para obter a padronização da detecção simultânea de Brucella ovis, Actinobacillus seminis e H. somni em sêmen fresco de ovinos.…”

Section: Introductionunclassified

Detecção de Histophilus somni (Haemophilus somnus) no sêmen bovino mediante reação em cadeia pela polimerase (PCR)

Dias

Nunes

Cavalcante

et al. 2014

Rev. Acad. Ciênc. Anim.

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Detecção de Histophilus somni (Haemophilus somnus) no sêmen bovino mediante reação em cadeia pela polimerase (PCR) [I] Detection of Histophilus somni (Haemophilus somnus) in bovine semen using polymerase chain reaction (PCR) [a] ResumoO presente estudo avaliou o uso da PCR para a detecção do Histophilus somni no sêmen bovino. Amostras de sêmen foram contaminadas experimentalmente com H. somni diluída em escalas de 10 7 a 10 1 bactérias/ mL, submetidas à extração de DNA pelo método de fenol/clorofórmio e amplificadas pela PCR. Os produtos da amplificação do DNA foram analisados por eletroforese em gel de acrilamida 8%. Por meio de oligonucleotídeos iniciadores obteve-se a amplificação de um fragmento de 400 pares de bases a partir do DNA da bactéria. Conseguiu-se amplificação positiva até na diluição de 10 1 bactérias/mL. A PCR mostrou-se eficiente na detecção de H. somni. O resultado disponibiliza conhecimento relevante para o diagnóstico de H. somni,

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Evaluation of a PCR test for the diagnosis of Tritrichomonas foetus infection in bulls: effects of sample collection method, storage and transport medium on the test

Cited by 44 publications

References 37 publications

Pooling of cultured samples and comparison of multistate laboratory workflows with the MagMAX sample preparation system and VetMAX quantitative polymerase chain reaction reagents for detection ofTritrichomonas foetus–colonized bulls

Pooling of cultured samples and comparison of multistate laboratory workflows with the MagMAX sample preparation system and VetMAX quantitative polymerase chain reaction reagents for detection ofTritrichomonas foetus–colonized bulls

Tritrichomonas foetus in Beef Bulls

Detecção de Histophilus somni (Haemophilus somnus) no sêmen bovino mediante reação em cadeia pela polimerase (PCR)

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