Kinetoplast DNA signatures of Trypanosoma cruzi strains obtained directly from infected tissues.
We report here a polymerase chain reaction (PCR)-based DNA profiling technique that permits Trypanosoma cruzi strain characterization by direct study of infected tissues. This is based on application of a recently developed method of DNA fragment identification, called low-stringency single specific primer PCR (LSSP-PCR), to the study of the variable region of kinetoplast DNA (kDNA) minicircles from T. cruzi Thus, we can translate the intraspecific polymorphism in the nucleotide sequence of kDNA minicircles into a specific and highly reproducible kDNA signature. Comparison with the phenogram obtained by DNA fingerprinting analysis of a set of T. cruzi strains showed good qualitative correlation between the degree of divergence of the LSSP-PCR profiles and the genetic distance between the strains. kDNA signatures of heart tissue from acutely or chronically infected animals revealed perfect concordance with the patterns obtained from cultured parasites for the CL and Colombiana strains but not for the Y strain, which is known to be multiclonal. However, the match was perfect for studies with two clones of the Y strain. We take this as evidence that in some multiclonal strains there is heterogeneity among the clones in the degree of tropism for the heart tissue. Finally, we showed that it is possible to obtain a T. cruzi kDNA signature from the heart of a human patient with chronic Chagasic myocardiopathy. kDNA signatures obtained by LSSP-PCR of sequences amplified from infected tissues constitute a new tool to study the molecular epidemiology of Chagas' disease.
Vago
A R
AR
Departamento de Bioquímica e Imunologia, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil.
Macedo
A M
AM
Oliveira
R P
RP
Andrade
L O
LO
Chiari
E
E
Galvão
L M
LM
Reis
D
D
Pereira
M E
ME
Simpson
A J
AJ
Tostes
S
S
Pena
S D
SD
D000818:Animals; D014355:Chagas Disease / Q000235:genetics* / Q000469:parasitology*; D002908:Chronic Disease; D016172:DNA Fingerprinting; D018105:DNA, Kinetoplast / Q000302:isolation & purification*; D006321:Heart / Q000469:parasitology; D006801:Humans; D008297:Male; D008875:Middle Aged; D010274:Parasitology / Q000379:methods*; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D015203:Reproducibility of Results; D014349:Trypanosoma cruzi / Q000235:genetics* / Q000254:growth & development
D018105:DNA, Kinetoplast