Residual foci of Triatoma infestans infestation: Surveillance and control in Rio Grande do Sul, Brazil, 2001-2018

Abstract INTRODUCTION: This retrospective study conducted from 2001 to 2018 investigated the residual foci of Triatoma infestans infestation in Rio Grande do Sul, Brazil. METHODS: The data were obtained via entomological surveillance and the distribution of vector occurrence. The coverage of active research was mapped. RESULTS: The largest coverage rate for active research was observed in the northwest region of the total of 515,081 domiciles researched. Most T. infestans specimens were captured in the peridomicile. CONCLUSIONS: Infestation has decreased significantly since 2008, and T. infestans has not been captured since 2015.

Triatoma infestans was the main insect vector of the protozoan Trypanosoma cruzi, an etiologic agent of Chagas disease (ChD), one of the most important parasitic diseases in Latin America 1 . In 1991, it was indicated that the control of T. infestans in endemic risk areas should be the focus of a collaborative effort by the Ministries of Health in the Southern Cone Initiative: Argentina, Bolivia, Brazil, Chile, Paraguay, and Uruguay. The Pan American Health Organization/World Health Organization (PAHO/WHO) established and coordinated the "Intergovernmental Commission on Chagas Disease," which prepared the "Triatoma infestans Elimination Plan" for the elimination of T. infestans and the interruption of transfusion transmission of trypanosomiasis. The plan defined vector elimination as follows: no capture of the vector for a minimum period of 3 years in an area with established and functional entomological surveillance 2 . The elimination plan was successful. There was an 83% reduction in the occurrence of T. infestans in Brazil in 1993, with some residual foci remaining in the states of Rio Grande do Sul (RS) and Bahia 3 .
This study aimed to evaluate the elimination of the vector and long-term entomological surveillance in RS. RS is located in south Brazil. RS shares a northern border with the State of Santa Catarina and international boundaries with Uruguay to the south and southwest, and Argentina to the west and northwest. RS has a humid subtropical climate and a variety of temperatures throughout the year, recording sub-zero temperatures in the winter and temperatures near 40 °C in the summer.   The insects were collected from Passive Surveillance (PS) through the Triatomine Information Post (TIP), also known as community surveillance (notified by the population); Active Research (AR); and integral research (IR), the active search of vectors in all of a positive locality for T. infestans in the previous year, and the active search in nearby localities.
The AR followed the field operations planning protocols in the rural areas. Public health agents were trained for AR and received supervision during fieldwork. Triatomine research was carried out using tweezers, an insect dislodging product, and a flashlight to better see the triatomines' hiding places in the intradomicile and peridomicile environments. However, it is important to report that capture failures are likely to occur, both on the part of the population and health workers. Therefore, continuous refreshers and training courses and educational programs must be provided.
The dataset included the following: year; number of municipalities with T. infestans infestation; positive domicile units (DUs) with T. infestans infestation and ecotope of capture in the DU occurrence of the capture in intradomicile (ID) and/or peridomicile (PD) areas; capture of 1 or > 1 insect; the presence of nymphs (N) in the ID and/or PD areas (nymphs characterize colonization of the DU); the total number of DUs researched; the total number of sprayed DUs; number and productivity of TIPs for triatomines; and other insects captured in the year ( Table 1).
The area of vector occurrence was mapped, overlapping the area of AR, with the software QGIS. The intensity of vector occurrence was established by the number of years during which T. infestans were identified in the municipalities between 2001 and 2018. These data were categorized by capture frequency: 1 year, 2-4 years, 5-6 years, and > 6 years. AR was evaluated using the coverage rate of AR (AR%). The AR% by year was defined as the number of DUs found by ChDCP divided by the number of rural DUs, from 497 municipalities in RS, by year, over 18 years. The AR% (total average) is the result of the division of the total accumulated coverage rate of active research by year and municipality, over 18 years (2001)(2002)(2003)(2004)(2005)(2006)(2007)(2008)(2009)(2010)(2011)(2012)(2013)(2014)(2015)(2016)(2017)(2018). The AR% was classified as follows: < 1%, > 1% to 15%, > 15% to 30%, > 30% to 45%, and > 45% to 65%. Municipalities (eight) with little or no rural population were categorized as < 1. The AR% was mapped and overlapped with the vector occurrence in the RS territory between 2001 and 2018. The T. infestans frequency from 1 to 6 years occurred in 19 municipalities (Figure 1).
The distribution of the last foci of T. infestans infestation from 2012 to 2014 was restricted to 3 municipalities (Figure 2). The PS (notified by the population) was motivated by the installation and maintenance of the TIP and educational activities for health promotion. An awareness campaign was launched in 2012 with promotional materials such as radio spots, posters, and brochures, reprinted and used in subsequent years 4  Greater AR coverage occurred in the region considered high risk for T. infestans, since all municipalities reported the occurrence of the vector in the past. These municipalities had foci of T. infestans between 1 and 6 years. The municipalities' areas were scheduled for AR and IR, and an AR% of 45-65% was found in 7 municipalities. The number of AR decreased in 2012 because of the consistent drop in the number of triatomines captured by health agents in rural DU. Active surveillance and spraying carried out by the health agents proved to be effective in the presented scenario within the areas with the occurrence of T. infestans.
Most T. infestans specimens were captured in PD areas, and the presence of more than one specimen was also more likely in PD areas than in ID areas. The number of colonies (the presence of nymphs in the ecotope) was also higher in the PD environment. Infestations with one T. infestans vector were more common in ID areas. With respect to the invasion site, the infestation of DUs was 4/6 consistently more common in PD areas than in ID areas, similar to results verified in Argentina 6 .
The foci were treated with residual action insecticide and integrated into the FUNASA's Housing Improvement Program for Chagas Disease Control (HIPCDC). Additionally, educational activities were implemented, as recommended by the Brazilian Consensus on Chagas disease. For housing improvement, the SES-RS allocated financial resources, based on presence of T. infestans, to the HIPCDC for the construction and renovation of DUs with outhouses in rural localities in northwest RS, making these areas inhospitable to triatomine colonization 7 . It is important to note that most improvements occurred in the peridomicile areas, including painting and renovations. Moreover, the teams intensified their work in hen houses and storerooms to prevent the entry of animals that could serve as T. cruzi reservoirs or triatomine food sources. This process triggered a series of trainings, domicile research, educational workshops, and the administrative and technical reorganization of ChDCP. Indeed, this contributed to the disappearance of the residual foci of T. infestans infestation, found after the implementation of the HIPCDC 8 . With respect to the consistent decrease in the number of triatomines, the entomologic scenario indicated other directions for the program: a major investment in the passive or community surveillance, with the installation and maintenance of TIPs in all municipalities of the state with the collected insects. Community participation in ChD vector surveillance is fundamental to the success of entomological control and, with community involvement, surveillance becomes continuous and is not performed by only ChDCP agents, resulting in long-term, sustainable control 9,10 . The number of reports of triatomines decreased during this period, but the productivity of TIPs has increased, as observed by the increase in the total number of arthropods in recent years, demonstrating that the educational actions carried out with the disclosure of TIPs have achieved the intended pedagogical effect.
The causes that may have influenced the occurrence and maintenance of populations of T. infestans in the northwestern region of RS, especially in the first years of the investigation, are as follows: climate of the region, since northwest RS has warmer annual average temperatures of 20-22°C (higher than that in other areas of the state: variations between 15 and 18°C, and a minimum

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average of 10ºC in the winter); socio-cultural profile of individuals from the northwest region (mainly Italian and German immigrants); and several outbuildings (storerooms, sheds, chicken houses, and brick ovens) and the accumulation of deposits and building materials (woods and bricks) in rural DUs in the northwest, which can lead to failures in spraying activities. These factors are known to favor infestation and might explain the recolonization 11,12 . It is presumed that there was operational failure in the actions of the Mobilization Project for the Elimination 13 of T. infestans implemented in 2010 in the municipality of Santa Rosa (Figure 2), since five foci of the vectors were found in 2014. It is suspected that the reported focus of the Church in Santo Cristo (2012-2013), a town near Santa Rosa, may have caused the spread of T. infestans in the region.
With respect to spraying, it has been reported that insect resistance to the chemical treatment employed (pyrethroid insecticides such as alpha-cypermethrin and deltamethrin) was not observed. In addition, Pessoa et al. (2015) 14 and Belisário et al. (2017) 15 demonstrated that the specimens of T. infestans strong genetic structure and little to no gene flow among populations, from the states of Bahia and RS, collected precisely in Santa Rosa and Doutor Maurício Cardoso. These municipalities, together with Santo Cristo, exhibited the last foci of T. infestans in this investigation and were susceptible to pyrethroid insecticide deltamethrin in laboratory tests, corroborating our field observations. The infestation has decreased significantly since 2008, and T. infestans was not captured in 2015, 2016, 2017, or 2018. The last foci were recorded in 2014. Thus, it is essential to note that the persistent occurrence of T. infestans in RS did not occur due to the entry of triatomines from other sources, including neighboring countries, and that the residual infestations cannot be attributed to resistance to pyrethroids.
After achieving the vector elimination plan, the main issue was to maintain the structure and provide surveillance coverage avoiding the reemergence of ChD. Thus, it was emphasized that entomological surveillance should be permanent, with the participation of health services and the community. These are points of vital importance to achieve sustainable control. The operational routine maintenance of AR in in areas without the domiciled vector is justifiable in the public administration only if combined with a vision of environmental health surveillance, including educational activities and the addition of a housing improvement component to ChD control, mandating environmental sanitation and care in rural housing. Although it is too early to determine whether long-term control will be achieved, the results confirm the efforts in the search to eliminate T. infestans in RS, coordinated by SES, municipal departments, public health agents, and the community.

ACKNOWLEDGMENTS
We offer our deepest thanks to the field agents or public health agents of FUNASA-MS in Rio Grande do Sul that provided technical support for development and implementation of these actions.

FINANCIAL SUPPORT
Secretaria Estadual da Saúde-Rio Grande do Sul, Brasil.

AUTHORS' CONTRIBUTION
CB: Conception and design of the study, acquisition of data, drafting the article, analysis and interpretation of data, final approval of the version to be submitted; TW: Conception and design of the study, acquisition of data; MMV: Design of the study, analysis and interpretation of data, final approval of the version to be submitted; APKR: Acquisition of data, analysis and interpretation of data, final approval of the version to be submitted; PS: Analysis and interpretation of data, final approval of the version to be submitted; CGCS: Acquisition of data, mapping of data; FM: Acquisition of data, analysis and interpretation of data, final approval of the version to be submitted.