Single-Tube Nested PCR for Detection of<i>Tritrichomonas</i><i>foetus</i>in Feline Feces

Gookin, Jody L.; Birkenheuer, Adam J.; Breitschwerdt, Edward B.; Levy, Michael G.

doi:10.1128/jcm.40.11.4126-4130.2002

Cited by 105 publications

(112 citation statements)

References 13 publications

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“…Currently, there is a general tendency to opt for direct molecular analysis of feces for the diagnosis of trichomoniasis (Gookin et al 2002, Paris et al 2014). According to these authors it is not possible to differentiate T. foetus from other protozoa with similar appearance and size as Giardia spp.…”

Section: Discussionmentioning

confidence: 99%

Co-infection by Tritrichomonas foetus and Pentatrichomonas hominis in asymptomatic cats

et al. 2015

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RESUMO.-[Co-infecção por Tritrichomonas foetus e Pentatrichomonas hominis em gatos assintomáticos.] Tritrichomonas foetus, um parasito bem conhecido por seu significado como um agente patogênico transmitido venereamente em bovinos, também foi identificado como causa de diarreia crónica do intestino grosso em gatos domésticos em muitos países. No Brasil, vários estudos sobre o diagnósti-co de tricomonose bovina foram realizados, mas até agora, não há informação disponível em relação à trichomonose felina. Assim, este é o primeiro estudo a relatar a ocorrência Tritrichomonas foetus, a parasite well known for its significance as a venereally transmitted pathogen in cattle, has been identified as a cause of chronic large bowel diarrhea in domestic cats in many countries of the world. In Brazil, several studies on the diagnosis of bovine trichomoniasis have been performed, but until now, no study was made regarding feline trichomoniasis. Thus, this is the first study to report the occurrence of T. foetus and Pentatrichomonas hominis in cats using morphological and molecular analysis. Feces from 77 cats were examined, four of which (5.2%) were positive for the presence of parabasalids. Morphological analysis of stained smears revealed piriform trophozoites showing the three anterior flagella, elongated nucleus and axostyle ending abruptly in fillet, characteristic of T. foetus. In scanning and transmission electron microscopy, identification characters similar to those previously reported for T. foetus were observed. The cultures containing trophozoites were submitted for molecular analysis, which resulted positive for T. foetus DNA using specific primers (TFR3 and TFR4), and all samples were positive and subjected to sequencing in which they showed 99.7-100% similarity with another isolate sequencing of T. foetus (JX960422). Although no trophozoite with consistent morphology of P. hominis has been visualized in the samples, differential diagnosis was performed using specific primers for P. hominis (TH3 and TH5) amplicon. In three of the four samples (3.89%) sequencing revealed 100% similarity when compared with another sequence of P. hominis deposited in Genbank (KC623939). Therefore, the present study revealed through the diagnostic techniques employed the simultaneous infection by T. foetus and P. hominis in the feces of cats. However, it was necessary to use more than one technique for the diagnosis of the co-infection. These results demonstrate the importance of a correct diagnosis to allow an appropriate treatment by the veterinarian.

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Section: Discussionmentioning

confidence: 99%

Co-infection by Tritrichomonas foetus and Pentatrichomonas hominis in asymptomatic cats

et al. 2015

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“…10 Feline T. foetus infection is diagnosed by direct fecal smear examination for trichomonads, cultivation of feces using a commercially available system (In Pouch TM TF), 11 or by extraction of DNA from feces and amplification of T. foetus rDNA by use of PCR analysis with species-specific primers. 9 One of the distinctive features of feline trichomonad-induced colitis is that affected cats are typically young. 10 At the time colon specimens were collected, all cats of this study were 1 year old or younger and ranged in age from 10 weeks to 12 months (mean, 6.7 6 1.7 months).…”

Section: Discussionmentioning

confidence: 99%

“…The organism can be diagnosed by microscopic observation of motile trichomonads in saline-diluted fecal smears, cultivation of trichomonads from feces using In Pouch TM TF media, 11 demonstration of T. foetus rDNA in feces by use of PCR analysis, 9 or by histologic evaluation of multiple sections of colon. If biopsies are to be evaluated, it is important to collect several specimens, as current data indicate that the organism is not necessarily present in all sections.…”

Section: Discussionmentioning

confidence: 99%

Histologic Features Associated withTritrichomonas foetus-induced Colitis in Domestic Cats

Yaeger

Gookin

2005

Vet Pathol

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Abstract. Tritrichomonas foetus is a venereal pathogen of naturally bred cattle. In domestic cats, T. foetus colonizes the colon, resulting in chronic, large-bowel diarrhea. The infection is prevalent among young, densely housed cats, and there is no effective treatment. To the authors' knowledge, the characteristic microscopic lesions of T. foetus infection in naturally infected cats have not been described. The aim of the study reported here was to characterize the histologic changes in the colon of seven cats with T. foetus infection and chronic diarrhea. All cats were 1 year old or younger (mean, 6.7 6 1.7 months), and a diagnosis of T. foetus infection was made on the basis of direct fecal smear examination (five cats), fecal culture in InPouch TM TF medium (four cats), single-tube nested polymerase chain reaction (PCR) analysis of DNA extracted from feces (two cats), or observation of trichomonads in sections of colon followed by PCR confirmation on DNA extracted from paraffinembedded tissue (two cats). The presence of colonic trichomonads was the most diagnostic histologic feature. Organisms were identified in all cats, but in only 24 of 43 (56%) sections of colon. Trichomonads were generally present in close proximity to the mucosal surface and less frequently in the lumen of colonic crypts. The presence of colonic trichomonads was consistently associated with mild-to-moderate lymphoplasmacytic and neutrophilic colitis, crypt epithelial cell hypertrophy, hyperplasia and increased mitotic activity, loss of goblet cells, crypt microabscesses, and attenuation of the superficial colonic mucosa. In two of the cats, histologic lesions were more severe and were associated with invasion of trichomonads into the lamina propria and/or deeper layers of the colon.

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“…On the basis of previous studies, the stool kit e has been documented to adequately extract purified DNA for use in PCR amplification. 11,24 However using DNA that was purified following strict adherence to the manufacturer's directions only resulted in amplification of samples with fairly high numbers of cells present (5 3 10 6 colony-forming units [CFU]/g). Also the amount of inhibitors present in DNA extractions required the use of 50-ml PCR reaction volumes for amplification.…”

Section: Optimization Of the Dna Extraction Methodsmentioning

confidence: 99%

“…16,24,26,43 The sensitivity of PCR amplification can be affected by the DNA extraction method and types of enhancers used in each reaction. 11,24 The sensitivity of the community DNA approach can also be affected by the amount of sample used and the number of replicate extractions made from each sample. 28,30 The use of community DNA from a sample as a means for detecting antibiotic resistance genes and for identifying potential gene reservoirs is becoming more common.…”

Section: Introductionmentioning

confidence: 99%

Development of a Polymerase Chain Reaction Assay for the Detection of Antibiotic Resistance Genes in Community DNA

Patterson

Singer

2006

J VET Diagn Invest

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Abstract. Many methods are used to detect antibiotic resistance genes in samples. The objective of the study reported here was to compare polymerase chain reaction (PCR) analysis of community DNA with fecal culturing for detecting antibiotic resistance genes in cattle samples. In the laboratory-based portion of this study, known concentrations of an Escherichia coli strain with 3 antibiotic resistance genes (cmy-2, flo, and cat) were added to feces from dairy cattle. These genes were used to assess the effect of various primer pairs, chromosomally versus plasmid-encoded genes, and gene copy number on the sensitivity of PCR amplification. Gene-specific PCR amplification was performed on the community DNA extracted from the feces. Feces were cultured for the inoculated strain. In the field-based portion of the study, 80 cattle fecal samples of unknown gene status were compared by use of similar methods. Culture and PCR amplification from community DNA extractions produced variable results, and this variability was most noticeable at dilutions that approached the detection limit of the assay. Typically, PCR amplification had a higher sensitivity than did culture for detecting the gene of interest. However, the sensitivity of culture was improved by plating on selective media containing antibiotics. The community DNA approach enables assessment of bacterial communities in complex samples such as feces, a task that can be prohibitive by budget or time constraints associated with culture methods. Through a strategic combination of culture and community DNA approaches, the relationship between specific selection pressures and the persistence and dissemination of specific resistance genes can be elucidated.

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Single-Tube Nested PCR for Detection ofTritrichomonasfoetusin Feline Feces

Cited by 105 publications

References 13 publications

Co-infection by Tritrichomonas foetus and Pentatrichomonas hominis in asymptomatic cats

Co-infection by Tritrichomonas foetus and Pentatrichomonas hominis in asymptomatic cats

Histologic Features Associated withTritrichomonas foetus-induced Colitis in Domestic Cats

Development of a Polymerase Chain Reaction Assay for the Detection of Antibiotic Resistance Genes in Community DNA

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