Populations of the common bed bug, Cimex lectularius, have recently undergone explosive growth. Bed bugs share many important traits with triatomine insects, but it remains unclear whether these similarities include the ability to transmit Trypanosoma cruzi, the etiologic agent of Chagas disease. Here, we show efficient and bidirectional transmission of T. cruzi between hosts and bed bugs in a laboratory environment. Most bed bugs that fed on experimentally infected mice acquired the parasite. A majority of previously uninfected mice became infected after a period of cohabitation with exposed bed bugs. T. cruzi was also transmitted to mice after the feces of infected bed bugs were applied directly to broken host skin. Quantitative bed bug defecation measures were similar to those of important triatomine vectors. Our findings suggest that the common bed bug may be a competent vector of T. cruzi and could pose a risk for vector-borne transmission of Chagas disease.

Triatomine vectors transmit Trypanosoma cruzi, the etiological agent of Chagas disease in humans. Transmission to humans typically occurs when contaminated triatomine feces come in contact with the bite site or mucosal membranes. In the Southern Cone of South America, where the highest burden of disease exists, Triatoma infestans is the principal vector for T. cruzi. Recent studies of other vector-borne illnesses have shown that arthropod microbiota influences the ability of infectious agents to colonize the insect vector and transmit to the human host. This has garnered attention as a potential control strategy against T. cruzi, as vector control is the main tool of Chagas disease prevention. Here we characterized the microbiota in T. infestans feces of both wild-caught and laboratory-reared insects and examined the relationship between microbial composition and T. cruzi infection using highly sensitive high-throughput sequencing technology to sequence the V3-V4 region of the 16S ribosomal RNA gene on the MiSeq Illumina platform. We collected 59 wild (9 with T. cruzi infection) and 10 lab-reared T. infestans (4 with T. cruzi infection) from the endemic area of Arequipa, Perú. Wild T. infestans had greater hindgut bacterial diversity than laboratory-reared bugs. Microbiota of lab insects comprised a subset of those identified in their wild counterparts, with 96 of the total 124 genera also observed in laboratory-reared insects. Among wild insects, variation in bacterial composition was observed, but time and location of collection and development stage did not explain this variation. T. cruzi infection in lab insects did not affect α- or β-diversity; however, we did find that the β-diversity of wild insects differed if they were infected with T. cruzi and identified 10 specific taxa that had significantly different relative abundances in infected vs. uninfected wild T. infestans (Bosea, Mesorhizobium, Dietzia, and Cupriavidus were underrepresented in infected bugs; Sporosarcina, an unclassified genus of Porphyromonadaceae, Nestenrenkonia, Alkalibacterium, Peptoniphilus, Marinilactibacillus were overrepresented in infected bugs). Our findings suggest that T. cruzi infection is associated with the microbiota of T. infestans and that inferring the microbiota of wild T. infestans may not be possible through sampling of T. infestans reared in the insectary. Chagas disease in humans is caused by the parasite Trypanosoma cruzi and it is endemic to the Americas. Poor populations are most at risk. The parasite infects an estimated six million people of 21 endemic countries in the Americas, with 30,000 new infections yearly. The main mode of transmission is vector-borne by triatomine bugs, which tend to live in close association with humans. The main Chagas disease vector in the Southern Cone of South America, where the highest burden of disease exists, is Triatoma infestans. As blood-sucking insects, triatomines become infected when they bite a T. cruzi-positive human and once infected they transmit the parasites in their feces. Controlling the vector populations is the main strategy of Chagas disease transmission reduction efforts. Microbiota-mediated methods to control this vector-borne disease are now being explored to determine whether microbes typically found in the vectors’ gut have a detrimental effect on T. cruzi and how they may be used to modify the vector and curb the ability for T. cruzi to be transmitted to humans. To advance this new field, we first must gain better knowledge of the gut microbiota of triatomines. Our study is the first to use sensitive high-throughput methods to study the gut microbes of T. infestans, using both laboratory-reared and wild insects. We have found that the microbial composition of T. infestans in the laboratory does not reflect the complete collection of gut microbes of wild T. infestans and inferring the gut microbiota profile of wild insects through studying lab insects alone may not be possible. We also found evidence that in wild insects T. cruzi affects the composition of the gut microbiota and identified some bacterial taxa which may be important in modulating the T.infestans-T.cruzi relationship.

Sexual reproduction provides an evolutionary advantageous mechanism that combines favorable mutations that have arisen in separate lineages into the same individual. This advantage is especially pronounced in microparasites as allelic reassortment among individuals caused by sexual reproduction promotes allelic diversity at immune evasion genes within individuals which is often essential to evade host immune systems. Despite these advantages, many eukaryotic microparasites exhibit highly-clonal population structures suggesting that genetic exchange through sexual reproduction is rare. Evidence supporting clonality is particularly convincing in the causative agent of Chagas disease, Trypanosoma cruzi, despite equally convincing evidence of the capacity to engage in sexual reproduction. In the present study, we investigated two hypotheses that can reconcile the apparent contradiction between the observed clonal population structure and the capacity to engage in sexual reproduction by analyzing the genome sequences of 123 T. cruzi isolates from a natural population in Arequipa, Peru. The distribution of polymorphic markers within and among isolates provides clear evidence of the occurrence of sexual reproduction. Large genetic segments are rearranged among chromosomes due to crossing over during meiosis leading to a decay in the genetic linkage among polymorphic markers compared to the expectations from a purely asexually-reproducing population. Nevertheless, the population structure appears clonal due to a high level of inbreeding during sexual reproduction which increases homozygosity, and thus reduces diversity, within each inbreeding lineage. These results effectively reconcile the apparent contradiction by demonstrating that the clonal population structure is derived not from infrequent sex in natural populations but from high levels of inbreeding. We discuss epidemiological consequences of this reproductive strategy on genome evolution, population structure, and phenotypic diversity of this medically important parasite. The rearrangement of alleles among individuals in a population during sexual reproduction maintains high allelic diversity within individuals in a population at polymorphic genes. Allelic diversity within individuals can be particularly important for parasites as it enhances their ability to evade host immune systems. Despite the potential benefits of sexual reproduction for parasites, natural populations of the protozoan parasite—and causative agent of human Chagas disease—Trypanosoma cruzi, exhibit clonal population structures indicative of asexual reproduction. This is particularly surprising as T. cruzi has the capacity for sexual reproduction. Here, we resolve this apparent contradiction by sequencing whole genomes of 123 T. cruzi isolates from a natural population in Arequipa, Peru. Evidence of past sexual reproduction and allelic rearrangements are common in this T. cruzi population. However, the majority of sexual reproduction events occur between close relatives resulting in an apparent clonal population structure. Sexual reproduction with distant relatives in areas with greater strain diversity has the potential to affect public health by increasing diversity in immune evasion genes within individuals and enhancing within-host survival, rapidly diversifying antigens that could affect the sensitivity of serological diagnostics, and by generating diversity in pathogenicity or drug resistance.

Chagas disease vector control campaigns are being conducted in Latin America, but little is known about medium-term or long-term effectiveness of these efforts, especially in urban areas. After analyzing entomologic data for 56,491 households during the treatment phase of a Triatoma infestans bug control campaign in Arequipa, Peru, during 2003–2011, we estimated that 97.1% of residual infestations are attributable to untreated households. Multivariate models for the surveillance phase of the campaign obtained during 2009–2012 confirm that nonparticipation in the initial treatment phase is a major risk factor (odds ratio [OR] 21.5, 95% CI 3.35–138). Infestation during surveillance also increased over time (OR 1.55, 95% CI 1.15–2.09 per year). However, we observed a negative interaction between nonparticipation and time (OR 0.73, 95% CI 0.53–0.99), suggesting that recolonization by vectors progressively dilutes risk associated with nonparticipation. Although the treatment phase was effective, recolonization in untreated households threatens the long-term success of vector control.

Chagas disease affects millions of people in Latin America. The control of this vector-borne disease focuses on halting transmission by reducing or eliminating insect vector populations. Most transmission of Trypanosoma cruzi, the causative agent of Chagas disease, involves insects living within or very close to households and feeding mostly on domestic animals. As animal hosts can be intermittently present it is important to understand how host availability can modify transmission risk to humans and to characterize the host-seeking dispersal of triatomine vectors on a very fine scale. We used a semi-field system with motion-detection cameras to characterize the dispersal of Triatoma infestans, and compare the behavior of vector populations in the constant presence of hosts (guinea pigs), and after the removal of the hosts. The emigration rate – net insect population decline in original refuge – following host removal was on average 19.7% of insects per 10 days compared to 10.2% in constant host populations (p = 0.029). However, dispersal of T. infestans occurred in both directions, towards and away from the initial location of the hosts. The majority of insects that moved towards the original location of guinea pigs remained there for 4 weeks. Oviposition and mortality were observed and analyzed in the context of insect dispersal, but only mortality was higher in the group where animal hosts were removed (p-value <0.01). We discuss different survival strategies associated with the observed behavior and its implications for vector control. Removing domestic animals in infested areas increases vector dispersal from the first day of host removal. The implications of these patterns of vector dispersal in a field setting are not yet known but could result in movement towards human rooms. Chagas disease is transmitted by triatomine bugs that actively disperse by walking and flying. The control of this vector-borne disease focuses on reducing or eliminating the insect vector populations. Most transmission of Trypanosoma cruzi, the causative agent of Chagas disease, involves insects living within or very close to households and feeding mostly on domestic animals. As animal hosts can be removed due to migration, slaughter, or death, it is important to understand how host availability can modify transmission risk to humans and to characterize the dispersal of triatomine vectors on a very fine scale. We used a semi-field system to characterize the dispersal of Triatoma infestans, and compare the behavior of vector populations in the constant presence of hosts and after the removal of the hosts. The emigration rate – net insect population decline in original refuges – following host removal was on average 19.7% of insects per 10 days compared to 10.2% in constant host populations. Activity of insects was significantly increased when hosts were removed. The removal of domestic animals in infested areas increases vector dispersal, possibly towards nearby human sleeping spaces.

The vector of Chagas disease, Triatoma infestans, is largely controlled by the household application of pyrethroid insecticides. Because effective, large-scale insecticide application is costly and necessitates numerous trained personnel, alternative control techniques are badly needed. We compared the residual effect of organophosphate-based insecticidal paint (Inesfly 5A IGR™ (I5A)) to standard deltamethrin, and a negative control, against T. infestans in a simulated natural environment. We evaluated mortality, knockdown, and ability to take a blood meal among 5^th instar nymphs. I5A paint caused significantly greater mortality at time points up to nine months compared to deltamethrin (Fisher's Exact Test, p < 0.01 in all instances). A year following application, mortality among nymphs in the I5A was similar to those in the deltamethrin (χ2 = 0.76, df=1, p < 0.76). At months 0 and 1 after application, fewer nymphs exposed to deltamethrin took a blood meal compared to insects exposed to paint (Fisher's Exact Tests, p < 0.01 and p < 0.01, respectively). Insecticidal paint may provide an easily-applied means of protection against vectors of Chagas disease.