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Geographical distribution of Trypanosoma cruzi genotypes in Venezuela.

Carrasco HJ, Segovia M, Llewellyn MS, Morocoima A, Urdaneta-Morales S, Martínez C, Martínez CE, Garcia C, Rodríguez M, Espinosa R, de Noya BA, Díaz-Bello Z, Herrera L, Fitzpatrick S, Yeo M, Miles MA, Feliciangeli MD - PLoS Negl Trop Dis (2012)

Bottom Line: The dataset includes 778 samples collected and genotyped over the last twelve years from multiple hosts and vectors, including nine wild and domestic mammalian host species, and seven species of triatomine bug, as well as from human sources.Most isolates (732) can be assigned to the TcI clade (94.1%); 24 to the TcIV group (3.1%) and 22 to TcIII (2.8%).Importantly, among the 95 isolates genotyped from human disease cases, 79% belonged to TcI - a DTU common in the Americas, however, 21% belonged to TcIV- a little known genotype previously thought to be rare in humans.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de Biología Molecular de Protozoarios, Instituto de Medicina Tropical, Facultad de Medicina, Universidad Central de Venezuela, Caracas, Venezuela. hernan.carrasco@ucv.ve

ABSTRACT
Chagas disease is an endemic zoonosis native to the Americas and is caused by the kinetoplastid protozoan parasite Trypanosoma cruzi. The parasite is also highly genetically diverse, with six discrete typing units (DTUs) reported TcI - TcVI. These DTUs broadly correlate with several epidemiogical, ecological and pathological features of Chagas disease. In this manuscript we report the most comprehensive evaluation to date of the genetic diversity of T. cruzi in Venezuela. The dataset includes 778 samples collected and genotyped over the last twelve years from multiple hosts and vectors, including nine wild and domestic mammalian host species, and seven species of triatomine bug, as well as from human sources. Most isolates (732) can be assigned to the TcI clade (94.1%); 24 to the TcIV group (3.1%) and 22 to TcIII (2.8%). Importantly, among the 95 isolates genotyped from human disease cases, 79% belonged to TcI - a DTU common in the Americas, however, 21% belonged to TcIV- a little known genotype previously thought to be rare in humans. Furthermore, were able to assign multiple oral Chagas diseases cases to TcI in the area around the capital, Caracas. We discuss our findings in the context of T. cruzi DTU distributions elsewhere in the Americas, and evaluate the impact they have on the future of Chagas disease control in Venezuela.

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Isoenzyme profiles of GPI and PGM from representative circulating Venezuelan Trypanosoma cruzi strains.Loading order, left to right: TcI, WA250 cl 10B; TcII, Esmeraldo cl2; TcIV, CanIII cl 1; 1, 8839(TcIV); 2, 8196(TcIV); 3, 10141(TcI); 4, 10610(TcIV); 5, 8089(TcI); 6, 6872(TcI); 7, PGN23(TcI); 8, PGN27(TcIV); 9, PGN31(TcI).
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pntd-0001707-g001: Isoenzyme profiles of GPI and PGM from representative circulating Venezuelan Trypanosoma cruzi strains.Loading order, left to right: TcI, WA250 cl 10B; TcII, Esmeraldo cl2; TcIV, CanIII cl 1; 1, 8839(TcIV); 2, 8196(TcIV); 3, 10141(TcI); 4, 10610(TcIV); 5, 8089(TcI); 6, 6872(TcI); 7, PGN23(TcI); 8, PGN27(TcIV); 9, PGN31(TcI).

Mentions: A phenotypic analysis was initially done using the isoenzyme technique as described by Miles et al,. (1977) [7]. For this analysis we used phosphoglucomutase (E.C.2.7.5.1, PGM) and glucose phosphate isomerase (E.C.S.3.l.9, GPI) enzymes (Figure 1). They were examined by thin-layer starch gel electrophoresis as described by Carrasco et. al. (1996) [26]. Random Amplified Polymorphic DNA (RAPD) genotyping (Figure 2) was performed as in Carrasco et al., (1996) [26]. PCR reactions for RAPD typing were achieved using primers A1, A2, L4 and L5 (Table 3). Each reaction took place in a 20 µL final volume containing 10 mM Tris HCl (pH 8.8) buffer, 0.2 Mm of each dNTP, 20 pg of primer, 1.0 unit of Taq DNA polymerase (Invitrogen, Brazil) and included 5 ng of whole genomic DNA. Reaction conditions were as follows: two cycles at 95°C for 5 min, 30°C for 2 min and 72°C for 1 min, 32 cycles at 95°C for 1 min, 40°C for 2 min, and 72°C for 1 min, and a final extension cycle at 72°C for 5 min. Primer sequences are listed in Table 3. PCR restriction fragment length polymorphism (PCR-RFLP) genotyping (Figure 3) targeted two loci: Glucose phosphate isomerase (GPI) and Heat Shock Protein 60 (HSP60) genes were amplified and cut using restriction enzymes HhaI and EcoRV respectively, following protocols set out in Westenberger et al. 2005 [27].


Geographical distribution of Trypanosoma cruzi genotypes in Venezuela.

Carrasco HJ, Segovia M, Llewellyn MS, Morocoima A, Urdaneta-Morales S, Martínez C, Martínez CE, Garcia C, Rodríguez M, Espinosa R, de Noya BA, Díaz-Bello Z, Herrera L, Fitzpatrick S, Yeo M, Miles MA, Feliciangeli MD - PLoS Negl Trop Dis (2012)

Isoenzyme profiles of GPI and PGM from representative circulating Venezuelan Trypanosoma cruzi strains.Loading order, left to right: TcI, WA250 cl 10B; TcII, Esmeraldo cl2; TcIV, CanIII cl 1; 1, 8839(TcIV); 2, 8196(TcIV); 3, 10141(TcI); 4, 10610(TcIV); 5, 8089(TcI); 6, 6872(TcI); 7, PGN23(TcI); 8, PGN27(TcIV); 9, PGN31(TcI).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3383755&req=5

pntd-0001707-g001: Isoenzyme profiles of GPI and PGM from representative circulating Venezuelan Trypanosoma cruzi strains.Loading order, left to right: TcI, WA250 cl 10B; TcII, Esmeraldo cl2; TcIV, CanIII cl 1; 1, 8839(TcIV); 2, 8196(TcIV); 3, 10141(TcI); 4, 10610(TcIV); 5, 8089(TcI); 6, 6872(TcI); 7, PGN23(TcI); 8, PGN27(TcIV); 9, PGN31(TcI).
Mentions: A phenotypic analysis was initially done using the isoenzyme technique as described by Miles et al,. (1977) [7]. For this analysis we used phosphoglucomutase (E.C.2.7.5.1, PGM) and glucose phosphate isomerase (E.C.S.3.l.9, GPI) enzymes (Figure 1). They were examined by thin-layer starch gel electrophoresis as described by Carrasco et. al. (1996) [26]. Random Amplified Polymorphic DNA (RAPD) genotyping (Figure 2) was performed as in Carrasco et al., (1996) [26]. PCR reactions for RAPD typing were achieved using primers A1, A2, L4 and L5 (Table 3). Each reaction took place in a 20 µL final volume containing 10 mM Tris HCl (pH 8.8) buffer, 0.2 Mm of each dNTP, 20 pg of primer, 1.0 unit of Taq DNA polymerase (Invitrogen, Brazil) and included 5 ng of whole genomic DNA. Reaction conditions were as follows: two cycles at 95°C for 5 min, 30°C for 2 min and 72°C for 1 min, 32 cycles at 95°C for 1 min, 40°C for 2 min, and 72°C for 1 min, and a final extension cycle at 72°C for 5 min. Primer sequences are listed in Table 3. PCR restriction fragment length polymorphism (PCR-RFLP) genotyping (Figure 3) targeted two loci: Glucose phosphate isomerase (GPI) and Heat Shock Protein 60 (HSP60) genes were amplified and cut using restriction enzymes HhaI and EcoRV respectively, following protocols set out in Westenberger et al. 2005 [27].

Bottom Line: The dataset includes 778 samples collected and genotyped over the last twelve years from multiple hosts and vectors, including nine wild and domestic mammalian host species, and seven species of triatomine bug, as well as from human sources.Most isolates (732) can be assigned to the TcI clade (94.1%); 24 to the TcIV group (3.1%) and 22 to TcIII (2.8%).Importantly, among the 95 isolates genotyped from human disease cases, 79% belonged to TcI - a DTU common in the Americas, however, 21% belonged to TcIV- a little known genotype previously thought to be rare in humans.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de Biología Molecular de Protozoarios, Instituto de Medicina Tropical, Facultad de Medicina, Universidad Central de Venezuela, Caracas, Venezuela. hernan.carrasco@ucv.ve

ABSTRACT
Chagas disease is an endemic zoonosis native to the Americas and is caused by the kinetoplastid protozoan parasite Trypanosoma cruzi. The parasite is also highly genetically diverse, with six discrete typing units (DTUs) reported TcI - TcVI. These DTUs broadly correlate with several epidemiogical, ecological and pathological features of Chagas disease. In this manuscript we report the most comprehensive evaluation to date of the genetic diversity of T. cruzi in Venezuela. The dataset includes 778 samples collected and genotyped over the last twelve years from multiple hosts and vectors, including nine wild and domestic mammalian host species, and seven species of triatomine bug, as well as from human sources. Most isolates (732) can be assigned to the TcI clade (94.1%); 24 to the TcIV group (3.1%) and 22 to TcIII (2.8%). Importantly, among the 95 isolates genotyped from human disease cases, 79% belonged to TcI - a DTU common in the Americas, however, 21% belonged to TcIV- a little known genotype previously thought to be rare in humans. Furthermore, were able to assign multiple oral Chagas diseases cases to TcI in the area around the capital, Caracas. We discuss our findings in the context of T. cruzi DTU distributions elsewhere in the Americas, and evaluate the impact they have on the future of Chagas disease control in Venezuela.

Show MeSH
Related in: MedlinePlus