First Report of Eratyrus cuspidatus (Hemiptera: Reduviidae) Infected with Trypanosoma cruzi in Peridomestic Environment in Chiapas,Mexico

Introduction

Triatomines (Hemiptera: Reduviidae) are bloodsucking insects with a wide distribution in the Americas. To date, 18 genera, 5 tribes, and 157 species have been described, most of which are associated with sylvatic ecotopes, but some are associated with domestic and peridomestic ones where they interact with humans (Alevi et al., 2021). Several triatomine species are of medical importance because they are vectors of the protozoan parasite Trypanosoma cruzi, which causes American trypanosomiasis or Chagas disease. Of the 6–7 million people worldwide infected with T. cruzi, the vast majority live in poor rural areas of the Americas where triatomines and humans frequently come into contact (WHO, 2022).

Mexico represents the second greatest diversity of triatomines in the Americas (after Brazil), with more than 30 species described (Ceccarelli et al., 2018). Most of them have been reported to be infected with T. cruzi, and 19 species are epidemiologically relevant because of their ability to invade and colonize human dwellings. Twelve species, including Eratyrus cuspidatus Stal, are considered strictly sylvatic (Ramsey et al., 2015). The role of sylvatic triatomine species as vectors of T. cruzi in Mexico remains to be fully understood.

The geographic range of E. cuspidatus extends from Southern Mexico to Northern Peru (Galvão et al., 1998). Studies conducted in South America showed that E. cuspidatus is a vector of T. cruzi (Dib et al., 2009; Moreno et al., 1996), but its role in the epidemiology of American trypanosomiasis in Mexico remains to be fully determined. In Mexico, E. cuspidatus was first reported in the southeastern states of Veracruz, Chiapas, Yucatán (Cruz-Reyes and Pickering-López, 2006), and more recently in Oaxaca (Ramsey et al., 2015) and Campeche (Tamay-Segovia et al., 2020) (Fig. 1A). In Chiapas, the first report of this species was made in Region XII in the municipality of Ocosingo (Bonampak reserve) and, subsequently, a second report was made in Region V in San Cristobal de Las Casas (GBIF 2023; Zarate and Zarate, 1985). E. cuspidatus collected in Chiapas were from sylvatic ecotopes, and for 30 years, its presence was not reported. In 2015, E. cuspidatus was collected again in a residential house in Region VI in the municipality of Villaflores, Chiapas (Fig. 1B) (Ramsey et al., 2015).

OPEN IN VIEWER

FIG. 1.

(A) Map of Mexican Republic showing the distribution of Eratyrus cuspidatus in the Southern region, states are highlighted in gray. (B) State of Chiapas divided into the 15 economic regions, dark gray circles showing previous record of E. cuspidatus, and light gray circle showing new record and first report of E. cuspidatus infected with Trypanosoma cruzi. (C) Environmental view of the collection site. (D) Dorsal view of a female specimen of E. cuspidatus, measuring 27 mm. (E) Microscopic visualization of T. cruzi in E. cuspidatus faces stained with Wright's dye in 100 × . (F) Amplification by TaqMan probe, Ct for positive control (I) was 17.32 and 25.33 for E. cuspidatus sample (II). Curve I shows positive control, curve II shows E. cuspidatus sample, and threshold line. (G) Mini-exon gene amplification shows a band in 350 pb that corresponds to lineage I. Ct, cycle threshold.

Despite vector control efforts, Chiapas is one of the Southeastern Mexican states with the highest number of diagnosed cases of acute American trypanosomiasis (Velázquez-Ramírez et al., 2022). In this context, special attention has been given to Triatoma dimidiata (Latreille) as the main vector of T. cruzi (Benítez-Alva et al., 2012), while sylvatic vectors such as E. cuspidatus that could play an important role in the transmission of T. cruzi in human dwellings have been overlooked. To the best of our knowledge, there are no reports of E. cuspidatus infected with T. cruzi in Mexico. The goal of this study was to present a new finding and report for the first time the presence of T. cruzi in E. cuspidatus in Mexico.

Materials and Methods

The state of Chiapas located in Southeastern Mexico (23.634501, -102.552784) is bordered to the north by Tabasco, east and southeast by the Republic of Guatemala, south by the Pacific Ocean, and west by the states of Oaxaca and Veracruz. The mean annual temperature varies depending on the region, from 18°C in the Chiapas Highlands to 28°C in the Coastal Plain (Fig. 1B). Our research on the epidemiology of Chagas disease in Southeastern Mexico involves sampling triatomines in rural settings (Velázquez-Ramírez et al., 2022). A triatomine was collected around 11:00 am (GMT-5) on the outside wall of a house in the town of Ignacio Allende (17.248995349653658, -92.14866717152924) in the municipality of Tumbalá, Chiapas (17.27866665418175, -92.31491107751356) (Fig. 1C). The floor and walls of the house were made of solid material (cement) with tin roof and no domestic mammals seen around, except for poultry sleeping on the periphery of the house. The triatomine was placed in a plastic container and taken to the laboratory at El Colegio de la Frontera Sur in San Cristóbal de Las Casas, Chiapas, Mexico, for analysis.

Intestinal material, by means of abdominal pressure, was obtained from the triatomine for fresh smears and staining by Wright's technique. The fecal sample preparation was examined under an optical microscope with 40 × and 100 × magnification for flagellated forms of T. cruzi. Another fecal sample was placed in an Eppendorf tube with 0.9% saline solution in a 1:1 ratio and kept refrigerated at 4°C. The triatomine was placed in 95% alcohol and stored at room temperature for identification using a taxonomic key (Lent and Wygodzinsky, 1979).

For molecular characterization, DNA from feces was extracted by Quick-DNA Miniprep Plus Kit from Zymo Research, and detection of T. cruzi was performed by quantitative polymerase chain reaction (qPCR) from T. cruzi satellite DNA (satDNA) amplification with the following primers: cruzi1c (5′TGAATGGYGGYGGGAGTCAGAG′3), cruzi2c (5′ATTCCTCCAAGMAGCGGAT′3), and probe cruzi3 (5′FAM-CACACACTGGACACACCAA-NFQ-MGB′3) under the following conditions: an initial 10 min passage at 95°C, followed by 45 cycles at 95°C for 15 s and 56°C for 1 min (Wehrendt et al., 2019).

Lineage identification was performed by amplifying the mini-exon gene with the mixture of three previously described oligonucleotides (5′-GTGTCCGCCACCTCCTTCGGGCC [TCI, specific for group 1], 5′-CCTGCAGGCACACGTGTGTGTG [TCII, specific for group 2], and 5′-CCCCCCTCCCAGGCCACACTG [TC, common to groups 1 and 2]) (Souto et al., 1996). A previously characterized strain was included for positive control (De Fuentes-Vicente et al., 2021). The negative control sample consisted of all qPCR components, except template DNA. The protocol was approved by the Research Ethics Committee of El Colegio de La Frontera Sur (CEI-06-07-2023).

Results

The triatomine collected was identified morphologically as an adult female E. cuspidatus. This species is characterized by its abundant pilosity, the anterior lobe of the pronotum with 1 + 1 short slightly acuminate disk tubercles, and the hemelytra do not cover tergite VII (Fig. 1D). It presents two tubercles in the anterior lobe of the pronotum and the humeral angle ending in a tubercle. The connexivum is particularly black, and the dorsal and ventral sides of the abdomen are light brown. The first two segments of the rostrum are elongated, and the pronotum is trapezoidal in shape. The scutellum is elongated, and the legs are robust and elongated. Metacyclic forms of T. cruzi were observed in the feces of this adult female E. cuspidatus (Fig. 1E). This was confirmed by qPCR (Fig. 1F). Amplification of the mini-exon gene showed that the T. cruzi infecting E. cuspidatus corresponded to lineage I (Fig. 1G), which is the predominant T. cruzi genetic lineage detected in Mexico.

Discussion

Despite the scarce records of E. cuspidatus in Mexico, the findings reported in this study confirm reports from South America on the vector competence for T. cruzi of this triatomine species. From the vectorial capacity, it is noteworthy that in this study, E. cuspidatus was collected in a residential home where the household members mentioned not having seen any other triatomine-looking bugs. Limited reports of E. cuspidatus may be due to its sylvatic nature as is the case for the majority of triatomine species in Mexico.

We are uncertain if the infection of E. cuspidatus with T. cruzi collected in Ignacio Allende, Chiapas, was established naturally. Beyond its apparent role in the enzootic cycle of T. cruzi, E. cuspidatus, where present, could transmit T. cruzi in and around human dwellings in rural environments (Dib et al., 2009). The domiciliation of sylvatic triatomines relates to the modification of their natural habitat, which forces them to search for a blood meal from hosts beyond wildlife, or may also be attracted by the light of peridomestic environments, which is an alternative for their establishment because food can be accessed more easily through domestic animals and humans (Cardozo et al., 2021).

We do not know the underlying mechanisms by which sylvatic triatomines may adapt to the periphery or interior of human dwellings, but since it is a gradual process, this phenomenon requires further research in rural and semirural parts of Mexico. For example, future studies involving sampling of E. cuspidatus in different environments across its geographic range in Mexico will help understand the adaptation of wild triatomines to human dwellings.

Conclusions

The adaptation of intrusive sylvatic vectors such as E. cuspidatus represents an important epidemiological risk in the transmission of T. cruzi (Dib et al., 2009). This highlights the need for vector surveillance research that includes sylvatic triatomine species as part of epidemiological studies in the context of global change (Gürtler, 2022). Adapting research findings to the national vector control program will enhance efforts to manage the health burden of acute and chronic American trypanosomiasis in the Southeastern region of Mexico (Secretaria de Salud, 2022).