Patricia L. Dorn

Autochthonous Transmission of Trypanosoma cruzi, Louisiana

Autochthonous transmission of the Chagas disease parasite, Trypanosoma cruzi, was detected in a patient in rural New Orleans, Louisiana. The patient had positive test results from 2 serologic tests and hemoculture. Fifty-six percent of 18 Triatoma sanguisuga collected from the house of the patient were positive for T. cruzi by PCR.

Two Distinct Triatoma dimidiata (Latreille, 1811) Taxa Are Found in Sympatry in Guatemala and Mexico

Approximately 10 million people are infected with Trypanosoma cruzi, the causative agent of Chagas disease, which remains the most serious parasitic disease in the Americas. Most people are infected via triatomine vectors. Transmission has been largely halted in South America in areas with predominantly domestic vectors. However, one of the main Chagas vectors in Mesoamerica, Triatoma dimidiata, poses special challenges to control due to its diversity across its large geographic range (from Mexico into northern South America), and peridomestic and sylvatic populations that repopulate houses following pesticide treatment. Recent evidence suggests T. dimidiata may be a complex of species, perhaps including cryptic species; taxonomic ambiguity which confounds control. The nuclear sequence of the internal transcribed spacer 2 (ITS2) of the ribosomal DNA and the mitochondrial cytochrome b (mt cyt b) gene were used to analyze the taxonomy of T. dimidiata from southern Mexico throughout Central America. ITS2 sequence divides T. dimidiata into four taxa. The first three are found mostly localized to specific geographic regions with some overlap: (1) southern Mexico and Guatemala (Group 2); (2) Guatemala, Honduras, El Salvador, Nicaragua, and Costa Rica (Group 1A); (3) and Panama (Group 1B). We extend ITS2 Group 1A south into Costa Rica, Group 2 into southern Guatemala and show the first information on isolates in Belize, identifying Groups 2 and 3 in that country. The fourth group (Group 3), a potential cryptic species, is dispersed across parts of Mexico, Guatemala, and Belize. We show it exists in sympatry with other groups in Peten, Guatemala, and Yucatan, Mexico. Mitochondrial cyt b data supports this putative cryptic species in sympatry with others. However, unlike the clear distinction of the remaining groups by ITS2, the remaining groups are not separated by mt cyt b. This work contributes to an understanding of the taxonomy and population subdivision of T. dimidiata, essential for designing effective control strategies.

Genetics and evolution of triatomines: from phylogeny to vector control

Triatomines are hemipteran bugs acting as vectors of the protozoan parasite Trypanosoma cruzi. This parasite causes Chagas disease, one of the major parasitic diseases in the Americas. Studies of triatomine genetics and evolution have been particularly useful in the design of rational vector control strategies, and are reviewed here. The phylogeography of several triatomine species is now slowly emerging, and the struggle to reconcile the phenotypic, phylogenetic, ecological and epidemiological species concepts makes for a very dynamic field. Population genetic studies using different markers indicate a wide range of population structures, depending on the triatomine species, ranging from highly fragmented to mobile, interbreeding populations. Triatomines transmit T. cruzi in the context of complex interactions between the insect vectors, their bacterial symbionts and the parasites; however, an integrated view of the significance of these interactions in triatomine biology, evolution and in disease transmission is still lacking. The development of novel genetic markers, together with the ongoing sequencing of the Rhodnius prolixus genome and more integrative studies, will provide key tools to expanding our understanding of these important insect vectors and allow the design of improved vector control strategies.

Identification of a large hybrid zone between sympatric sibling species of Triatoma dimidiata in the Yucatan peninsula, Mexico, and its epidemiological importance.

Triatoma dimidiata is one of the major Chagas disease vectors, with an extensive diversity in its morphology, habitat, and level of domiciliation. Molecular studies based on the internal transcribed spacer 2 (ITS-2) have subdivided this species into four potential taxonomic groups. Using both ITS-2 and cytochrome B markers, we confirmed the sibling species status of ITS-2 Group 3 and detected an apparent sympatry of ITS-2 Groups 2 and 3 in the Yucatan peninsula, Mexico. Here we examine the geographic distribution of T. dimidiata ITS-2 genotypes in the region and compare their egg production and Trypanosoma cruzi infection rates, as indicators of biological differences between groups. PCR genotyping of large natural populations showed an extensive sympatry of Groups 2 and 3 in most of the peninsula, often within the same house. We also detected a large proportion of individuals displaying ITS-2 sequences from both Groups 2 and 3, suggesting hybridization. Analysis of ITS-2 genotype frequencies indicated a strong departure from Hardy-Weinberg equilibrium in female hybrids, but not in males, due to a large heterozygote deficit. These results suggest random mating between ITS-2 Groups 2 and 3 combined with reduced viability and/or survival in female hybrids. This and other factors may allow for the maintenance of distinct ITS-2 Groups 2 and 3 populations despite high hybrid frequencies. Importantly, T. cruzi infection was much higher in hybrids compared to ITS-2 Groups 2 and 3 individuals, but all three genotypes appeared to seasonally infest houses in a similar manner in the region. These findings warrant further studies on T. dimidiata taxonomy and its epidemiologic implications.

Inhibition of protein synthesis results in super-induction of procyclin (PARP) RNA levels.

Procyclin is an abundant surface antigen found exclusively on the procyclic forms of African trypanosomes. We are interested in the induction of procyclin gene expression during differentiation from bloodstream forms. We find that increased levels of procyclin RNA are evident as early as 15 min after triggering differentiation. The increase in procyclin RNA levels requires the temperature shift from 37 degrees C to 27 degrees C and is aided by addition of the tricarboxylic acid cycle intermediate cis-aconitate. Maximal induction is observed with a combination of three triggers of differentiation: citrate, cis-aconitate and the temperature shift. Protein synthesis does not appear to be required for induction of procyclin RNA during differentiation. In fact, addition of protein synthesis inhibitors results in super-induction of procyclin RNA levels, even under conditions where no induction is normally observed (i.e., at 37 degrees C in the absence of citrate and cis-aconitate). This super-induction was observed with four different protein synthesis inhibitors that affect different stages of translation. Thus, the accumulation of procyclin transcripts may be under the control of a negative regulator whose effective levels are reduced during differentiation from bloodstream to procyclic forms.

The Chagas Vector, Triatoma dimidiata (Hemiptera:Reduviidae), is Panmictic within and Among Adjacent Villages in Guatemala

Abstract Trypanosoma cruzi, the hemoflagellate parasite and cause of Chagas disease in Latin America, is carried by Triatomine vectors, principally Triatoma dimidiata and Rhodnius prolixus in Central America. To assist control efforts and to understand the epidemiology of the disease in Guatemala, the population genetics of T. dimidiata was analyzed among three houses within a village and two adjacent villages in Guatemala. Eleven Randomly Amplified Polymorphic DNA-polymerase chain reaction (RAPD-PCR) primers were screened and three used to amplify bands, 29 of which were scored, from T. dimidiata DNA of ≈50 bugs per house from three houses within a village and from 66 and 33 bugs, respectively, from adjacent villages. Results show very small genetic distances among the three T. dimidiata subpopulations from the houses (D = 0.013–0.022) and the two villages (D = 0.0199). The amount of differentiation among houses (fixation index, FST) was also very small, FST = 0.025 among the houses and the two villages FST = 0.019. These fixation indices give an average number of mating migrants per generation (Nm) of 9.7 (among houses) and 12 (among villages). Average heterozygosity (H) appears to be high, ranging from H = 0.299–0.325 among the houses and H = 0.273 among the villages. The low genetic distance and fixation indices, and high heterozygosity suggest that the subpopulations in the houses and in the adjacent villages are not reproductively isolated but are in fact, one large panmictic population. Therefore the geographic coverage necessary for effective control must include, at least, the area encompassing adjacent villages.

Vector blood meals and Chagas disease transmission potential, United States.

A high proportion of triatomine insects, vectors for Trypanosoma cruzi trypanosomes, collected in Arizona and California and examined using a novel assay had fed on humans. Other triatomine insects were positive for T. cruzi parasite infection, which indicates that the potential exists for vector transmission of Chagas disease in the United States.

A Preliminary Assessment of Genetic Differentiation of Triatoma dimidiata (Hemiptera: Reduviidae) in Guatemala by Random Amplification of Polymorphic DNA-Polymerase Chain Reaction

Abstract The population genetics of Triatoma dimidiata (Latreille, 1811) from five different provinces in Guatemala, including three sylvan and three domestic populations, was investigated by random amplification of polymorphic DNA-polymerase chain reaction. There is a high degree of genetic variation in all of the T. dimidiata populations as evidenced by high levels of average expected heterozygosity and polymorphism. Domestic populations are more closely related to each other (D = 0.05–0.085, Nei’s genetic distance) than are the sylvan (D = 0.121–0.189). Within the limited sample size of three populations, there was a correlation with geographic and genetic distance for the domestic populations, but not for the sylvan. Surprisingly, one of the sylvan populations was genetically very similar to the domestic populations. The FST demonstrated a high degree of differentiation at the country-wide level (FST = 0.175) and a moderate degree of differentiation within the sylvan (FST = 0.135) or domestic (FST = 0.097) populations. Although these results demonstrated that gene flow is limited between different provinces in Guatemala, hierarchical analysis showed that barriers between the Atlantic and Pacific drainage slopes were not biologically significant limiters of gene flow.

Comparison of two immunochromatographic assays and the indirect immunofluorscence antibody test for diagnosis of Trypanosoma cruzi infection in dogs in south central Louisiana

Two rapid tests evaluated in dogs considered to be of high risk of infection with the Chagas parasite Trypanosoma cruzi using two immunochromatographic assays: Trypanosoma Detect for canine, InBios, Seattle, WA and CHAGAS STAT-PAK assay, Chembio Diagnostic Systems, Medford, NY, in south central Louisiana. For this purpose a serological survey was carried out in a total of 122 dogs and a serum bank was created. These 122 animals were first tested by IFAT that was used as the standard test. From the serum bank 50 samples were tested using the two rapid Chagas assays and results compared to the standard test IFAT. The serological survey using IFAT showed a prevalence of T. cruzi infection in 22.1% of the tested dogs. In the immunochromatographic assays, 13 and 11 animals were positive on rapid assay: Trypanosoma Detect for canine, InBios and CHAGAS STAT-PAK, Chembio Diagnostic Systems, respectively compared to 11 positive by IFAT. These two immunochromatographic tests have shown high susceptibility and specificity compared to our standard method IFAT. The rapid, easy and accurate screening assays used in conjunction with confirmatory tests, would be an excellent tool for veterinarians to diagnose T. cruzi infection. Early detection of T. cruzi infection may prevent complications through an effective treatment. Greater awareness by veterinarians of the risk, clinical findings, history along with diagnostic methods will contribute greatly to an understanding of the true prevalence of Chagas disease in dogs in Louisiana.

Triatoma sanguisuga blood meals and potential for Chagas disease, Louisiana, USA.

To evaluate human risk for Chagas disease, we molecularly identified blood meal sources and prevalence of Trypanosoma cruzi infection among 49 Triatoma sanguisuga kissing bugs in Louisiana, USA. Humans accounted for the second most frequent blood source. Of the bugs that fed on humans, ≈40% were infected with T. cruzi, revealing transmission potential.