Ellen M. Dotson

Risk factors for kala-azar in Bangladesh

Since 1990, South Asia has experienced a resurgence of kala-azar (visceral leishmaniasis). To determine risk factors for kala-azar, we performed cross-sectional surveys over a 3-year period in a Bangladeshi community. By history, active case detection, and serologic screening, 155 of 2,356 residents had kala-azar with onset from 2000 to 2003. Risk was highest for persons 3–45 years of age, and no significant difference by sex was seen. In age-adjusted multivariable models, 3 factors were identified: proximity to a previous kala-azar patient (odds ratio [OR] 25.4, 95% confidence interval [CI] 15–44 within household; OR 3.2 95% CI 1.7–6.1 within 50 m), bed net use in summer (OR 0.7, 95% CI 0.53–0.93), and cattle per 1,000 m2 (OR 0.8, 95% CI 0.70–0.94]). No difference was seen by income, education, or occupation; land ownership or other assets; housing materials and condition; or keeping goats or chickens inside bedrooms. Our data confirm strong clustering and suggest that insecticide-treated nets could be effective in preventing kala-azar.

The United States Trypanosoma cruzi Infection Study: evidence for vector-borne transmission of the parasite that causes Chagas disease among United States blood donors.

BACKGROUND: Screening US blood donors for Trypanosoma cruzi infection is identifying autochthonous, chronic infections. Two donors in Mississippi were identified through screening and investigated as probable domestically acquired vector‐borne infections, and the US T. cruzi Infection Study was conducted to evaluate the burden of and describe putative risk factors for vector‐borne infection in the United States.

Evaluation of long-lasting insecticidal nets after 2 years of household use.

Development of long‐lasting insecticidal nets (LLINs) may eliminate the need for insecticide retreatment of ITNs. While two LLINs (Olyset®, Sumitomo Chemical Co., Japan; and PermaNet® 1.0, Vestergaard‐Frandsen, Denmark) have received recommendations from the World Health Organization Pesticide Evaluation Scheme, field‐testing under normal use has been limited. We used a survival analysis approach to compare time to net failure of conventional polyester bednets treated only with deltamethrin to two LLINs and two candidate LLINs (Olyset®; PermaNet®; Insector, Athanor, France; and Dawa®, Siamdutch Mosquito Netting Co., Thailand). Additionally, we evaluated nets treated with a process designed to increase the wash‐durability of permethrin‐treated nets through the addition of cyclodextrin (a starch) in the treatment process. Houses in western Kenya were randomly assigned to one of the six net types and nets were distributed to cover all sleeping spaces. Households were visited monthly to assess reported side effects in inhabitants and washing frequency. Nets were evaluated for insecticidal activity by periodic WHO cone bioassays with mortality assessed at 24 h. Nets with bioassay mortality <70% were assayed monthly until failure, defined as the first of two consecutive bioassay mortality rates <50%. Time to failure was analyzed using an extended Cox Proportional Hazards model controlling for the cumulative number of washes. We distributed 314 nets to 177 households in June–July 2002; 22 nets (7.0%) were lost to follow‐up and 196 (62.4%) failed during the first 2 years of the evaluation. Controlling for cumulative number of washes, PermaNet® 1.0 [Hazard Ratio (HR) 0.14, 95% Confidence Interval (CI) 0.06–0.31] had a significantly lower risk of failure than conventional nets while Insector had a significantly higher risk of failure (HR 2.57, 95% CI 1.06–4.15). The risks of failure of the remaining nets (Olyset®: HR 1.29, 95% CI 0.79–2.10; Dawa®: HR 0.58, 95% CI 0.32–1.18; cyclodextrin: HR 0.65, 95% CI 0.40–1.1) were not significantly different from that of a conventional net. PermaNet® 1.0 performed significantly better than conventional nets and should be recommended to malaria control programs.

Bacterial symbiosis and paratransgenic control of vector-borne Chagas disease

The triatomine vectors of Chagas disease are obligate haematophagous insects, feeding on vertebrate blood throughout their entire developmental cycle. As a result of obtaining their nutrition from a single food source, their diet is devoid of certain vitamins and nutrients. Consequently, these insects harbour populations of bacterial symbionts within their intestinal tract, which provide the required nutrients that are lacking from their diet. We have isolated and characterised symbiont cultures from various triatomine species and developed a method for genetically transforming them. We can then reintroduce them into their original host species, thereby producing stable paratransgenic insects in which we are able to express heterologous gene products. Using this methodology, we have generated paratransgenic Rhodnius prolixus that are refractory for infection with Trypanosoma cruzi. Two examples of potentially refractory genes are currently being expressed in paratransgenic insects. These include the insect immune peptide cecropin A and active single chain antibody fragments. We have also developed an approach that would allow introduction of genetically modified bacterial symbionts into natural populations of Chagas disease vectors. This approach utilises the coprophagic behaviour of these insects, which is the way in which the symbionts are transmitted among bug populations in nature. The production and ultimate release of transgenic or paratransgenic insects for public health applications is potentially very promising but also worthy of much careful consideration with respect to environmental, political, and human safety concerns.

Laboratory wash resistance of long-lasting insecticidal nets

Long‐lasting insecticidal nets (LLINs) may eliminate the need for retreatment of mosquito nets used for the control of malaria and other vector‐borne diseases. The efficacy of LLINs after repeated washing under laboratory conditions has been used to predict long‐lasting efficacy under field conditions. We evaluated under laboratory conditions the wash resistance of two LLINs (PermaNet® 1.0, Vestergaard‐Frandsen, Denmark; Olyset®, Sumitomo Chemical Co., Japan), two candidate LLINs (Dawa®, Siamdutch Mosquito Netting Co., Thailand; Insector, Athanor, France) and a net treated with a process designed to increase its wash resistance and compared them with conventionally treated nets (deltamethrin, 25 mg/m2). Nets of all six types were washed using a standard protocol and tested weekly using WHO cone bioassays with Anopheles gambiae (Kisumu strain). The PermaNet® 1.0 was the most wash resistant with >50% mosquito mortality in WHO cone bioassays after as many as 20 washes. The Dawa® net also retained some activity after repeated washing but exhibited wide variation in insecticide retention and biological activity. The remaining nets lost >90% of their biological activity after six washes as measured by 24‐h mortality of A. gambiae in WHO cone tests. After 20 washes, all nets lost >50% of their initial insecticide concentrations except for the Olyset® net. After 20 washes, nets were heated for 4 h at 60 °C to determine whether biological activity could be restored by heat‐assisted regeneration. Only the Olyset® net was regenerated by heating, with average mosquito mortality and knockdown in WHO cone tests rising to >90% after heating for 4 h at 60 °C. However, regeneration of the biological activity of Olyset® nets that had been washed three times did not occur at 30 °C or 35 °C after 12 weeks. The wash resistance of these LLINs corresponded well to their retention of biological activity observed in a field trial, suggesting that wash resistance may be a good predictor of the longevity of insecticidal activity of LLINs under field conditions.

Hidden Sylvatic Foci of the Main Vector of Chagas Disease Triatoma infestans: Threats to the Vector Elimination Campaign?

Background Establishing the sources of reinfestation after residual insecticide spraying is crucial for vector elimination programs. Triatoma infestans, traditionally considered to be limited to domestic or peridomestic (abbreviated as D/PD) habitats throughout most of its range, is the target of an elimination program that has achieved limited success in the Gran Chaco region in South America. Methodology/Principal Findings During a two-year period we conducted semi-annual searches for triatomine bugs in every D/PD site and surrounding sylvatic habitats after full-coverage spraying of pyrethroid insecticides of all houses in a well-defined rural area in northwestern Argentina. We found six low-density sylvatic foci with 24 T. infestans in fallen or standing trees located 110–2,300 m from the nearest house or infested D/PD site detected after insecticide spraying, when house infestations were rare. Analysis of two mitochondrial gene fragments of 20 sylvatic specimens confirmed their species identity as T. infestans and showed that their composite haplotypes were the same as or closely related to D/PD haplotypes. Population studies with 10 polymorphic microsatellite loci and wing geometric morphometry consistently indicated the occurrence of unrestricted gene flow between local D/PD and sylvatic populations. Mitochondrial DNA and microsatellite sibship analyses in the most abundant sylvatic colony revealed descendents from five different females. Spatial analysis showed a significant association between two sylvatic foci and the nearest D/PD bug population found before insecticide spraying. Conclusions Our study shows that, despite of its high degree of domesticity, T. infestans has sylvatic colonies with normal chromatic characters (not melanic morphs) highly connected to D/PD conspecifics in the Argentinean Chaco. Sylvatic habitats may provide a transient or permanent refuge after control interventions, and function as sources for D/PD reinfestation. The occurrence of sylvatic foci of T. infestans in the Gran Chaco may pose additional threats to ongoing vector elimination efforts.

Temperature, Larval Diet, and Density Effects on Development Rate and Survival of Aedes aegypti (Diptera: Culicidae)

Many environmental factors, biotic and abiotic interact to influence organismal development. Given the importance of Aedes aegypti as a vector of human pathogens including dengue and yellow fever, understanding the impact of environmental factors such as temperature, resource availability, and intraspecific competition during development is critical for population control purposes. Despite known associations between developmental traits and factors of diet and density, temperature has been considered the primary driver of development rate and survival. To determine the relative importance of these critical factors, wide gradients of conditions must be considered. We hypothesize that 1) diet and density, as well as temperature influence the variation in development rate and survival, 2) that these factors interact, and this interaction is also necessary to understand variation in developmental traits. Temperature, diet, density, and their two-way interactions are significant factors in explaining development rate variation of the larval stages of Ae. aegypti mosquitoes. These factors as well as two and three-way interactions are significantly associated with the development rate from hatch to emergence. Temperature, but not diet or density, significantly impacted juvenile mortality. Development time was heteroskedastic with the highest variation occurring at the extremes of diet and density conditions. All three factors significantly impacted survival curves of experimental larvae that died during development. Complex interactions may contribute to variation in development rate. To better predict variation in development rate and survival in Ae. aegypti, factors of resource availability and intraspecific density must be considered in addition, but never to the exclusion of temperature.

Infection of Kissing Bugs with Trypanosoma cruzi, Tucson, Arizona, USA

A survey of triatomine insects found that 41.5% were infected with the causative agent of Chagas disease.

Molecular Population Genetics and Phylogeography of the Chagas Disease Vector Triatoma infestans in South America

ABSTRACT Knowledge of the genetic variability, population structure, and evolutionary history of Triatoma infestans may be useful for developing rational vector control strategies. A 661-bp fragment of the mitochondrial gene cytochrome oxidase I (COI) was sequenced and analyzed in bugs from Argentina, Uruguay, Peru, and Bolivia, including peridomestic, domestic, Andean, and Chaco sylvatic bugs. A total of 48 polymorphic sites among 37 haplotypes were described. Nucleotide variation fluctuated among samples, with the highest nucleotide diversity observed in seven Argentinean provinces. Within this group, some populations showed patterns of variability compatible with population expansions and/or fine-scale population structure, whereas others suggested population bottlenecks and/or population admixture processes. A maximum parsimony analysis of the haplotypes showed the presence of a Bolivian/Peruvian and an Argentinean/Uruguayan clade. Bolivian sequences were further divided in Chaco sylvatic and Andean domestic and sylvatic. Two different nested clades were found within the Argentinean/Uruguayan cluster. Analysis of molecular variance (AMOVA) and KST* analysis supported a strong population structure in Argentina, where genetic differentiation was correlated with geographic distance. Departures from neutrality expectations and a nested cladistic analysis suggest a recent population expansion of T. infestans in Argentina, followed by restricted gene flow and patterns of isolation by distance. This expansion could have taken place as a two-wave process, as was shown by the phylogenetic analysis and signatures of population admixture in the southernmost Argentinean populations.

Genetic structure of Triatoma infestans populations in rural communities of Santiago del Estero, northern Argentina.

To gain an understanding of the genetic structure and dispersal dynamics of Triatoma infestans populations, we analyzed the multilocus genotype of 10 microsatellite loci for 352 T. infestans collected in 21 houses of 11 rural communities in October 2002. Genetic structure was analyzed at the community and house compound levels. Analysis revealed that vector control actions affected the genetic structure of T. infestans populations. Bug populations from communities under sustained vector control (core area) were highly structured and genetic differentiation between neighboring house compounds was significant. In contrast, bug populations from communities with sporadic vector control actions were more homogeneous and lacked defined genetic clusters. Genetic differentiation between population pairs did not fit a model of isolation by distance at the microgeographical level. Evidence consistent with flight or walking bug dispersal was detected within and among communities, dispersal was more female-biased in the core area and results suggested that houses received immigrants from more than one source. Putative sources and mechanisms of re-infestation are described. These data may be use to design improved vector control strategies.