Triatomine fauna in the state of Bahia, Brazil: What changed after 40 years of the vector-control program?

ABSTRACT Background: Neglected tropical diseases are a growing threat to global health, and endemic Chagas disease has emerged as one of the most important health problems in America. The main strategy to prevent Trypanosoma cruzi transmission is chemical control of vectors. This study presents a descriptive analysis of synanthropic triatomines before and after the implementation of a vector-control program in Bahia, Brazil. Methods: Descriptive analysis and geospatial statistics were performed on triatomine data, (1) the relative abundance and (2) proportional spatial distribution, from Bahia during two periods: (A) 1957 to 1971 and (B) 2006 to 2019. Results: We observed a decrease in the relative abundance of Panstrongylus megistus (A: n=22.032, 61.9%; B: n=1.842, 1.0%) and Triatoma infestans (A: n=1.310, 3.7%; B: n=763, 0.43%), as well as an increase in the relative abundance of T. sordida (A: n=8.314, 23.4%, B: n=146.901, 81.6%) and T. pseudomaculata (A: n=894, 2.5%, B: n=16.717, 9.3%). Conclusions: Our results indicate a clear reduction in the occurrence of P. megistus and T. infestans (last record in 2015) and an increase in the relative abundance and geographical distribution of T. sordida and T. pseudomaculata after 40 years of the vector-control program. The high frequency of other triatomine species in the municipalities of the state of Bahia and their abundance in recent years highlight the need to reinforce permanent entomological surveillance actions to prevent Chagas disease.


INTRODUCTION
Triatomines are blood-sucking insects (Hemiptera: Reduviidae) that are vectors of the parasite Trypanosoma cruzi (Chagas, 1909), the etiological agent of Chagas disease, also known as American trypanosomiasis. Some synanthropic triatomine species adapt to anthropic changes within their natural landscape, colonizing the household environment (animal breeding sites) and occasionally inside houses, increasing the risk of T. cruzi transmission 1,2 . Many triatomines invade houses, but few can initiate the processes of colonization and domiciliation, which depend on the characteristics of the invasive species 3,4 , the invaded dwelling 5,6, and the environment around the household 7 .
In Brazil, some triatomine species have succeeded in occupying the domestic environment and expanding their spatial distribution beyond their original biomes 8 through active and passive dispersal 9,10 . One of these species is Triatoma infestans (Klug, 1835), which occupied household units among 12 states of Brazil and was considered the main vector involved in T. cruzi transmission in Brazil 11,12 . After the first standardized chemical-based control actions were implemented by the National Chagas Disease Control Program in 1975 and by the integrated initiatives of the Southern Cone of Latin America to eliminate T. infestans in 1991, a sustained decrease in domestic populations of vectors was observed [13][14][15] . In 2006, Brazil received certification of interruption of Chagas disease transmission by T. infestans from the Pan American Health Organization (PAHO) and the World Health Organization (WHO) 16 . However, many regions of Brazil, such as the state of Bahia, still have several species of native vectors that can transmit T. cruzi to humans and domestic animals. Pirajá da Silva initially described the existence of triatomines in the state of Bahia in 1911, shortly after T. cruzi was described by Carlos Chagas 17 . Pirajá da Silva identified Conorhinus megistus triatomines -syn. Panstrongylus megistus (Burmeister, 1835) -in the city of Mata de São João, near Salvador, the capital city of Bahia. Pirajá da Silva also identified triatomines among the cities of Feira de Santana, Candeias, São Francisco do Conde, and Salvador 18 . Almost 40 years after Pirajá's initial research description, Chagas disease was recognized as a serious health problem in the state of Bahia, where triatomines were captured simultaneously to the record of autochthonous cases of Chagas heart disease in Salvador 19 . P. megistus and Triatoma rubrofasciata (De Geer, 1773) naturally infected by T. cruzi were found at the historic center of Salvador city, associated with human cases, which motivated a chemical control campaign 20 . Nevertheless, some foci of P. megistus maintained T. cruzi transmission to families in the neighborhoods of Salvador 20 . In the early 1970s, more than 600 specimens of P. megistus and T. rubrofasciata were examined in Salvador, of which 16% were infected with T. cruzi 21  Currently, 26 triatomine species have been recognized in Bahia [25][26] . Several factors can modify the spatial distribution and abundance of synanthropic triatomines. To understand the changes in T. cruzi vector occurrence, the use of retrospective epidemiological and entomological data has become a relevant strategy. In this study, we describe the relative abundance and proportional spatial distribution of synanthropic triatomines at the municipal level before (1957 to 1971) and after (2006 to 2019) the vector-control program implementation in the state of Bahia, which is one of the states with the greatest diversity of triatomines and highest rates of epidemiological information related to Chagas disease in northeastern Brazil.

METHODS
The state of Bahia has 417 municipalities and is located in the southern region of northeastern Brazil, bordering eight other Brazilian states and the Atlantic Ocean in the east (Figure 1). Descriptive analyses of both the spatial and temporal distribution of the synanthropic triatomine species data from Bahia was performed during two periods: Information regarding period B was obtained from the following state government databases after the certification of interruption of Chagas disease transmission by T. infestans: Epidemiologic Surveillance team of the State of Bahia's Health Service (SESAB), Epidemiological Surveillance Office (DIVEP), and Central Public Health Laboratory of Bahia (LACEN/BA) 29 .
Entomological data were obtained in three different ways: (a) notification of triatomines (surveillance and community participation) performed by the population itself by taking suspicious insects to health agents; (b) notification attendance (active surveillance) performed by the health agents of each municipality, dependent on (a); and (c) active search (active surveillance), in which vectors were searched among household units around the area, regardless of any notification performed by the population.
To compare T. cruzi vector species information from both periods of the study, we evaluated: (1) the relative abundance, calculated as the proportion of each triatomine species divided by the total number of triatomines for each period; and (2) proportional spatial distribution, calculated as the proportion of municipalities that recorded a triatomine species divided by the number of sampled municipalities.
Spreadsheets were used to collect the following data: species, municipality, date, geographic information system (GIS) coordinates, sampling environment (intradomestic, peridomestic), and data reference. Graphs and descriptive statistics were computed using SPSS 24. In the absence of specific GIS information, the coordinates of the city's headquarters from the Brazilian Institute of Geography and Statistics (IBGE) were used 30 . Data processing was performed using the ArcGis® Software 10.5 31 .
By analyzing the spatial distribution of the main triatomine species before and after T. cruzi vector-control actions, we observed a reduction in the municipal occurrence of synanthropic populations of T. infestans and P. megistus. T. infestans was no longer detected in some municipalities in the west (e.g., Santa Maria Vitória, Barreiras) and north (e.g., Juazeiro, Curaçá) of Bahia (Figure 3) and had been first detected among other municipalities in the Caatinga (e.g., Itaguaçu da Bahia, Novo Horizonte) and Atlantic Forest biomes (e.g., Tremedal and Presidente Tancredo Neves). During period B, T. infestans was found only at residual foci and was last recorded in 2015 in the municipality of Novo Horizonte (Figure 3).
During period B, we observed a higher occurrence of T. sordida in western Bahia, where the Cerrado biome predominates, in south-central Bahia, and in some municipalities of Recôncavo Baiano, eastern Bahia. However, we observed a remarkable change in the spatial distribution of T. pseudomaculata, which expanded its area of occurrence among western municipalities (Cerrado biome) and the central region (Caatinga biome) of Bahia (Figure 3).

DISCUSSION
This study indicated changes in the spatial distribution and relative abundance of synanthropic triatomines in the state of Bahia, Northeast Brazil, before and after 44 years of structured Legend: Color gradients represent the species richness at each period of the study. "X" axis shows the name of the triatomine species evaluated, while "Y" axis shows the proportional (%) spatial distribution of them.

A (1957-1971) B (2006-2019) A (n) A (%) B (n) B (%) A (n) A (%) B (n) B (%)
Cavernicola pilosa Barber, 1937   Panstrongylus megistus was the predominant species between 1957 and 1971; it was found in 60% of municipalities, with a relative abundance of 62% of collected triatomines 22 . Its proportional spatial distribution has been greatly reduced in the state of Bahia, especially in the areas of Recôncavo Baiano and the metropolitan region of Salvador. P. megistus spatial distribution has also been reduced in other Brazilian states 11,[36][37][38] . Three hypotheses could explain this reduction in the metropolitan region of Salvador: (a) chemical control was successfully performed over four decades, resulting in the elimination of domestic populations of P. megistus; (b) intense urbanization in these municipalities resulted in deforestation and fragmentation of the Atlantic Forest biome areas, which is the natural habitat of this species 22 ; and (c) housing improvement, with progressive depletion of adobe houses 39,40 , a favorable environment for P. megistus colonization. P. megistus is a native Brazilian species with a wide spatial distribution and high epidemiological and entomological relevance due to the high rates of T. cruzi infection and its proximity to human dwellings, as it can colonize intra-and/or peridomicile areas. In 2021, P. megistus foci were described in the metropolitan region of São Paulo, with T. cruzi-infected triatomines associated with marsupials, revealing the importance of continued surveillance of synanthropic P. megistus 46 .
During period A, T. infestans were found in 12.8% of the sampled municipalities and represented 3.7% of the sampled triatomines. During period B, T. infestans was identified in only seven municipalities with residual colonies [34][35] . Bahia was the last Brazilian state to receive PAHO certification, possibly because of specific T. infestans identification errors and the appearance of new records of this species in the study area 32 . The elimination of T. infestans in several municipalities in western and central Bahia can be explained by the T. infestans elimination plan. This plan was intensified in 2004 and included spraying households with insecticides, followed by research and capture of triatomines, surveying approximately 500000 households 26 . However, residual colonies of this species have been detected in a few municipalities 28,29 , requiring constant monitoring to eliminate residual foci from the state of Bahia. In addition to the chemical control performed in Bahia since 1975, intensified in 1991, which aimed to eliminate T. infestans 11,15 , other social actions were implemented by the federal government. These included the growth acceleration program, intended to improve housing quality, with the replacement of mud houses with brick houses and health education actions on Chagas disease. This may also have influenced the reduction in household colonization by T. infestans and other household species 39,40. Although successful in controlling T. infestans, several native species have been recorded in a large number of municipalities. They were captured in households and frequently colonized peri domestic areas. Among them, some were infected by T. cruzi and many of them fed on domestic animals and human blood 42 . In period A, T. sordida was recorded in 37% of the municipalities, representing 23% of the collected triatomines, while T. pseudomaculata was recorded in 19% of the municipalities, with a relative abundance of 2.5% of the gathered triatomines 23 . In period B, T. sordida was recorded in 60% of the municipalities, representing approximately 81% of the triatomines. T. sordida is the most common species in different regions of Bahia. Similarly, T. pseudomaculata was identified in the largest number of municipalities during period B (67.4%). Systematic vector-control actions had a low impact on the spatial distribution of T. sordida and T. pseudomaculata in Bahia. There was a higher occurrence of T. sordida in western and south-central Bahia, where there are areas of Cerrado, which is the original biome of the species' natural populations 1,2,25 . The highest occurrence of T. pseudomaculata in western and central Bahia followed the occurrence predictions of the species based on environmental variables 42 . The number of specimens of T. sordida and T. pseudomaculata exceeded 90% of all triatomines collected in the state between 2006 and 2019.
We observed a higher occurrence of T. brasiliensis and similar species in Bahia, thus expanding the observations of Ribeiro-Jr et. al. 41 . Before systematized control actions, the T. brasiliensis species complex was registered in 10% of the municipalities of Bahia, and between 2006 and 2019, at least 94 municipalities (36%) registered this species. In the last few years, other species of the T. brasiliensis complex have been described in the state of Bahia, including T. juazeirensis, T. melanica, T. sherlocki, and T. petrocchiae 43 . Among these species, T. juazeirensis is noteworthy because it was predominantly collected inside household units. Other species, such as T. tibiamaculata, P. geniculatus, T. melanocephala, and P. lutzi, were intrusively detected inside the houses, mainly adult specimens.
There was a significant difference in the occurrence of T. tibiamaculata between the two periods. In period A, it was recorded in approximately 1% of the municipalities with a relative abundance of 0.01% 23 . In period B, it was recorded in 7% of the municipalities with a relative abundance of 0.55%. In recent decades, even though T. tibiamaculata has been naturally found inside nests of marsupials, rodents, and epiphytes in forests 1,22 , it has been recorded in peri domestic palms and inside households. Therefore, T. cruzi-infected T. tibiamaculata generates a transmission risk in Salvador, probably in the entire metropolitan region, and in the Atlantic Forest areas 44 .
Triatomine vectors of T. cruzi can be classified as native/non native, wild/non wild, domestic/peridomestic to enable the definition of effective vector-control strategies 45 . While domestic and peridomestic populations of species of triatomines are subject to spraying of chemical pesticides, which is the main strategy of vector control, native-wild species can persistently invade or colonize the household (peri-and intradomicile). Thus, this may represent a challenge for controlling T. cruzi transmission in the domestic environment.
This study has several limitations. Since the use of reference databases does not allow a broad analysis of data, it was impossible to obtain data on triatomines within habitats (intra-and peridomicile) in period A (1957)(1958)(1959)(1960)(1961)(1962)(1963)(1964)(1965)(1966)(1967)(1968)(1969)(1970)(1971). The effects of passive dispersion and seasonal changes on vector behavior were impossible to measure. In addition, the database does not allow descriptive evaluations of nymph occurrence. Moreover, not all municipalities collected data regularly during the evaluated periods, and the health surveillance service classifies municipalities among the high, medium, and low risk of transmission, emphasizing that there is no obligation to conduct regular entomological research among low-risk municipalities based on the classification presented by Brazil's Health Ministry 26 . Future studies reassessing risk classification at the municipal level are urgently needed for the redefinition of areas and risk of transmission. Further studies should analyze the situation of these areas to explain whether the absence of triatomines is due to the functioning of the service or biogeographic issues related to triatomines. In periods A and B, T. brasiliensis complex species were considered as one, and to avoid misinterpretations, data obtained in this study were evaluated at stricto sensu level 44 . In Bahia, nine species that are unlikely to be found within households 24,25,33 were recorded by the surveillance services. These included Belminus laportei, Eratyrus mucronatus, Microtriatoma trinidadensis, Panstrongylus tupynambai, P. lignarius, Rhodnius prolixus, Triatoma maculata, T. circummaculata, and T. rubrovaria. To reduce taxonomic identification errors, a guide was developed 33 and, more recently, a guidance manual for the surveillance of triatomines in Bahia, with identification keys, diagnosis, and distribution of vector species 26 .
This study describes changes in the triatomine fauna of Bahia between the analyzed periods more than 40 years after the implementation of the vector-control program in the state. During this period, we observed changes in the Chagas disease surveillance policies in Brazil due to decentralization. The centralized Brazilian control program was transferred to municipalities without corresponding training. The decentralization policy of the Chagas disease vector-control program aimed to bring the actions of the government closer to the communities, but true engagement by local communities has not been achieved. After decentralization, the efficacy of some municipalities in detecting triatomines has been reduced. We observed a reduction in the relative abundance and proportional spatial distribution of domestic and domesticated species of T. cruzi vectors P. megistus and T. infestans, respectively. In addition, we described an increase in the native species T. sordida, T. pseudomaculata, and T. brasiliensis species subcomplex and highlighted the important role of native triatomine species in T. cruzi transmission in the domiciliary environment. The observed changes in T. cruzi main vectors between the evaluated periods demonstrate the importance of reinforcing entomological surveillance actions. Furthermore, promoting and disseminating community-based scientific knowledge and health education actions on Chagas disease at a local scale will help mitigate surveillance challenges and control native triatomines.

Ethics
The procedures followed the ethical standards of the Research Ethics Committee of the Gonçalo Moniz Institute (IGM -FIOCRUZ, Bahia, Brazil). The consent form was waived since the analysis was performed using a database. However, to provide full anonymity to participants, no personal identification data were used. The research did not cause any physical, psychological, moral, intellectual, social, cultural, or religious risks to the residents or animals in the study areas. In addition, this study did not include endangered or protected species.

ACKNOWLEDGMENTS
We are grateful to the Epidemiologic Surveillance team of Bahia's State Health Service (SESAB-DIVEP) for the databases and all support with the triatomines and vectors information. We are also grateful to Rafaella Albuquerque Silva and César Augusto Cuba Cuba for their suggestions and comments. Also, we would like to thank Natalie Jennifer Hurrell for the help with the English language proofreading.