Primers design and optimization of PCR and nested-PCR assays for the specific detection of Tritrichomonas foetus

被引:6
作者
Fernandes, Paula Rogerio [1 ]
Da Silva, Andrea Caetano [2 ]
Gambarini, Maria Lucia [3 ]
Linhares, Guido Fontgalland C. [3 ]
机构
[1] Univ Fed Goias, Programa Posgrad Ciencia Anim, BR-74001970 Goiania, Go, Brazil
[2] Univ Fed Goias, Inst Patol Trop, BR-74001970 Goiania, Go, Brazil
[3] Univ Fed Goias, DMV, EV, BR-74001970 Goiania, Go, Brazil
来源
REVISTA BRASILEIRA DE PARASITOLOGIA VETERINARIA | 2008年 / 17卷 / 03期
关键词
Polymerase chain reaction; rDNA; molecular diagnosis; trichomonosis; bovine;
D O I
10.1590/S1984-29612008000300003
中图分类号
R38 [医学寄生虫学]; Q [生物科学];
学科分类号
07 ; 0710 ; 09 ; 100103 ;
摘要
Tritrichomonas foetus is a pathogenic protozoan that causes a venereal disease in cattle known as bovine genital tricomonosis. In spite of the efficacy to recognize the target genomic DNA, the protocols so far developed for the diagnosis of this organism by PCR promote some inespecific amplifications or they are unable to discriminate T foetus against other species within the genus. The objective of this study was to assess and optimize PCR and nested-PCR assays for the specific diagnosis of T foetus, using novel primers selected from the alignment of sequences of the genes 18S rRNA, 5.8S rRNA, 28S rRNA and of the internal transcribed spacers of the rDNA (ITS I and ITS2). A pair of primers was constructed for the genus-specific amplification of a 648 bp fragment and two others to amplify T. foetus species-specific fragments of 343 and 429 bp. No cross amplification was observed against Bos taurus genomic DNA neither against the DNA of usual bovine genital pathogens. Both, single and nested-PCR assays, presented analytical sensitivity to detect at least two T. foetus organisms.
引用
收藏
页码:133 / 138
页数:6
相关论文
共 26 条
[1]  
ALTSCHUL S, 1990, J MOL BIOL, V3, P403
[2]  
[Anonymous], 1843, Comptes Rendus Acad Sci
[3]  
BRENT R, 1994, CURRENT PROTOCOLS MO
[4]   Comparative sequence analysis of 5 center dot 8S rRNA genes and internal transcribed spacer (ITS) regions of trichomonadid protozoa [J].
Felleisen, RSJ .
PARASITOLOGY, 1997, 115 :111-119
[5]   Detection of Tritrichomonas foetus by PCR and DNA enzyme immunoassay based on rRNA gene unit sequences [J].
Felleisen, RSJ ;
Lambelet, N ;
Bachmann, P ;
Nicolet, J ;
Müller, N ;
Gottstein, B .
JOURNAL OF CLINICAL MICROBIOLOGY, 1998, 36 (02) :513-519
[6]  
FERREIRA JM, 1977, PATOLOGIA CLIN VET
[7]  
*FL STAT U, PRIM DES
[8]   Single-tube nested PCR for detection of Tritrichomonas foetus in feline feces [J].
Gookin, JL ;
Birkenheuer, AJ ;
Breitschwerdt, EB ;
Levy, MG .
JOURNAL OF CLINICAL MICROBIOLOGY, 2002, 40 (11) :4126-4130
[9]   An improved molecular assay for Tritrichomonas foetus [J].
Grahn, RA ;
BonDurant, RH ;
van Hoosear, KA ;
Walker, RL ;
Lyons, LA .
VETERINARY PARASITOLOGY, 2005, 127 (01) :33-41
[10]   DETECTION OF BOVINE TRICHOMONIASIS WITH A SPECIFIC DNA-PROBE AND PCR AMPLIFICATION SYSTEM [J].
HO, MSY ;
CONRAD, PA ;
CONRAD, PJ ;
LEFEBVRE, RB ;
PEREZ, E ;
BONDURANT, RH .
JOURNAL OF CLINICAL MICROBIOLOGY, 1994, 32 (01) :98-104