Evaluation of natural foci of Panstrongylus megistus in a forest fragment in Porto Alegre, State of Rio Grande do Sul, Brazil

Abstract

Introduction:
Panstrongylus megistus is commonly found in wild environments of the State of Rio Grande do Sul, Brazil. The aim of this study was to characterize the network of refuges used by triatomine in a forest fragment of Porto Alegre and to identify Trypanosoma cruzi infection, associated hosts and the epidemiological importance of both hosts and triatomines.

Methods:
Techniques including the spool-and-line method and active searching (transects) were used to identify natural foci.

Results:
The food source for each triatomine was determined using the precipitin test, and the infection of marsupials was determined by xenodiagnosis. A total of 33 adults (domestic environment) and 27 nymphs (wild environment) of P. megistus were found in addition to 43 Didelphis albiventris specimens. The infection rates of triatomine adults, triatomine nymphs and opossums with T. cruzi I were 64%, 73% and 69%, respectively. Birds, rodents and opossums were the main resources used by triatomine.

Conclusions:
This work presents the first characterization of a natural focus of P. megistus in Rio Grande do Sul. The natural characteristics of this focus and its implication in the transmission of T. cruzi are discussed.

Full text

575
Case ReportMajor Article
Revista da Sociedade Brasileira de Medicina Tropical 46(5):575-583, Sep-Oct, 2013
http://dx.doi.org/10.1590/0037-8682-0149-2013
INTRODUCTION
Address to: Dr. José Eloy dos Santos Jr. CPqRR/FIOCRUZ. Av. Augusto de
Lima 1715, Barro Preto, 30190-002 Belo Horizonte, MG, Brasil.
Phone: 55 31 3349-7704; Fax: 55 31 3295-3115
e-mail: santosjr_je@yahoo.com.br
Received 24 July 2013
Accepted 9 October 2013
Evaluation of natural foci of Panstrongylus megistus
in a forest fragment in Porto Alegre,
State of Rio Grande do Sul, Brazil
José Eloy dos Santos Jr[1], Mariana Gubert Viola[2], Elias Seixas Lorosa[3],
Evandro Marques de Menezes Machado[5], Antonio Leite Ruas Neto[5]
and Elio Corseuil[6]
[1]. Laboratório de Triatomíneos e Epidemiologia da Doença de Chagas, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, MG.
[2]. Programa de Pós-Graduação em Educação em Ciências: Química da Vida e da Saúde, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS.
[3]. Laboratório Nacional e Internacional de Referência em Taxonomia de Triatomíneos, Instituto Oswaldo Cruz, Fundação Oswaldo Cruz, Rio de Janeiro, RJ.
[4]. Departamento de Ciências Biológicas, Universidade Federal de Ouro Preto, Ouro Preto, MG. [5]. Unidade de Tapes, Universidade Estadual do Rio Grande
do Sul, Tapes, RS. [6]. Retired Professor, Free teaching in Entomology, Porto Alegre, RS.
ABSTRACT
Introduction: Panstrongylus megistus is commonly found in wild environments of the State of Rio Grande do Sul, Brazil.
The aim of this study was to characterize the network of refuges used by triatomine in a forest fragment of Porto Alegre and
to identify Trypanosoma cruzi infection, associated hosts and the epidemiological importance of both hosts and triatomines.
Methods: Techniques including the spool-and-line method and active searching (transects) were used to identify natural foci.
Results: The food source for each triatomine was determined using the precipitin test, and the infection of marsupials was
determined by xenodiagnosis. A total of 33 adults (domestic environment) and 27 nymphs (wild environment) of P. megistus
were found in addition to 43 Didelphis albiventris specimens. The infection rates of triatomine adults, triatomine nymphs and
opossums with T. cruzi I were 64%, 73% and 69%, respectively. Birds, rodents and opossums were the main resources used by
triatomine. Conclusions: This work presents the fi rst characterization of a natural focus of P. megistus in Rio Grande do Sul. The
natural characteristics of this focus and its implication in the transmission of T. cruzi are discussed.
Keywords: Panstrongylus megistus. Didelphis albiventris. Trypanosoma cruzi. Sylvatic foci. Rio Grande do Sul.
Chagas disease was discovered in 1909 and remains one
of the main neglected diseases in South America. In Brazil,
it is estimated that approximately 1.9 million individuals are
infected with Chagas disease, and an additional 23 million
are at risk of acquiring it1. Due to the success of the Chagas
Disease Control Program in 2006, Brazil was certifi ed as free
of disease transmissions by Triatoma infestans (Klug, 1834) and
blood transfusion2. Among the main species of autochthonous
triatomines of Brazil, Panstrongylus megistus (Burmeister,
1835) (Hemiptera, Reduviidae, Triatominae) is singularly
known for its high capacity to adapt to domestic environments
and wide distribution throughout the country, being present in
20 Federal Units3. Based on genetic and paleobiogeographic
characteristics, several authors have demonstrated that the
States of Bahia (BA), Minas Gerais (MG), Pernambuco (PE),
Rio de Janeiro (RJ) and São Paulo (SP) are probable areas
of species endemism4-6. Currently, some of these states,
particularly BA, MG, PE, SP and Alagoas, are areas of greatest
epidemiological importance for P. megistus7.
The presence of this triatomine in natural environments
represents possible sources of infestation and re-infestation of
residences and also contributes to the maintenance of the sylvatic
cycle of Trypanosoma cruzi3. Multiple authors have observed
cohabitation of P. megistus and small mammals in other natural
shelters since the beginning of the last century.
The fi rst occurrence of P. megistus in natural environments
was reported by Gomes8 in State of São Paulo (SP), citing the
capture of a female trying to feed from her captor in the forest.
In 1920, Oliveira9 presented evidence on possible relationships
among the triatomine and T. cruzi natural reservoirs in Rio
Grande do Sul (RS). However, he did not describe sylvatic foci.
The fi rst signifi cant sylvatic focus was reported by Leal et al.10,
who found numerous colonies in nests of opossums, rodents and
cuícas in bromeliad clumps as well as holes in trees on an island
in State of Santa Catarina (SC). In the same decade, Barretto
et al.11 found a large number of P. megistus nymphs and adults in
hollow trees, anfractuous agave clumps and roots of the fi g tree
Ficus euomphala in SP, recording a higher rate of infection in

576
Santos Jr JE et al - Evaluation of natural foci of Panstrongylus megistus, Porto Alegre, Brazil
METHODS
triatomines associated with opossums (39%) than rodents (17%).
Work performed in the 1970s by Forattini et al.6,12,13 showed
P. megistus foci in hollow trees inhabited by bats, rodents, birds
and, mainly, opossums as well as in epiphytic bromeliads, pine
Cryptomeria sp. and palm Attalea sp. inhabited by opossums and
rodents. In a study at Horto Forest (Rio de Janeiro-RJ) in 1982,
Miles et al.14 found 12 nymphs infected by the parasite in a hollow
tree inhabited by Didelphis marsupialis, and in 1985, Schlemper-
Jr et al.15 detected an 84.5% rate of infection with T. cruzi in
P. megistus associated with opossums and rodents in hollow trees
and bromeliad clumps in SC. Grisard et al.16 found nymphs in
hollow trees in the same state. The most recent work, performed
by Santos-Jr et al.17 in the Bambuí, MG, municipality, reported
the fi rst fi nding of a P. megistus sylvatic colony in 69 years of
research. This colony was discovered in the hollow of a large
tree (Dimorphandra mollis Benth, popularly known as faveira).
Unlike the Northeast and Southeast Brazilian states, in RS,
P. megistus is predominantly sylvatic and rarely colonizes the
domestic environment. However, adult insect visits are common
in dwellings in close proximity with sylvatic areas during the
warmer months18. The fi rst reports of Chagas disease vectors in
RS were associated with the discovery of Triatoma rubrovaria
(Blanchard, 1843) in 1911 and T. infestans in 1913 in the Pelotas
municipality and surrounding areas19,20. Although vectors were
detected during the early 20th century, the fi rst acute case was only
described in 1939 at the boundary with Uruguay21. RS was one of
the last states to be certifi ed as free of Chagas disease transmission
(2005) in Brazil22. Recently 11 species of triatomines have
been recognized in the state; markedly, P. megistus, T. infestans
(currently in residual foci) and T. rubrovaria have notable T. cruzi
infection rates of 15%, 4% and 0.9%, respectively22-27. In Porto
Alegre, four triatomines species, T. circummaculata, T. oliverai
(both rarely), T. infestans (last observed in 1950) and P. megistus
(observed since 1931), have been reported and are found mainly
in the southern municipality region28-30.
In addition to elucidating the trophic relationships between
species, the identifi cation of the various natural habitats shared
by triatomines and their hosts allows for the characterization
of potential risk areas related to T. cruzi transmission, leading
to better Chagas disease control planning31. Various techniques
have been used to identify these habitats, including light
trapping, live-baited adhesive capture (Noireau trap)32, spool-
and-line methods33 and precipitin tests34.
This study aimed to characterize the network of refuges used
by P. megistus in an ombrophilous dense forest fragment of
Porto Alegre with an emphasis on T. cruzi infection, associated
hosts and the epidemiological importance of both hosts and
triatomines for the municipality.
The municipality of Porto Alegre, the largest City of Rio
Grande do Sul, Brazil, is situated at 100km from the Atlantic
Ocean and has a humid subtropical climate. Samples were
collected in an ombrophilous dense forest fragment (30o12’05’’S,
51o12’43’’W, 50m) of approximately 14ha i n the Ponta Grossa
neighborhood, located in the Guaíba lake basin (Figure 1A, 1B,
1C and 1D). This region is formed by three forest areas: Alto
Uruguay Subtropical Forest, Pines Forest and Atlantic Slope
Rain Forest35. The inclusion or proximity of forests in urban
areas make this region vulnerable to environmental changes.
Specimens were collected between October 2005 and
September 2006. Four residences near the 10m from the forest
were investigated monthly for the presence of triatomines. Searches
using fl ashlights and tweezers were performed in each domiciliary
environment, and a light trap was installed (mercury lamp focused
on a white wall) at a frequency of three nights per month. In the
peridomiciliary, two doghouses, one pigsty, one hennery, fi rewood,
tiles and two barns were searched. In the intradomiciliary, all rooms
were investigated. In the sylvatic environment, marsupial refuges
were investigated using transect and spool-and-line methods.
Transects were traversed from north to south in the studied area,
inspecting shelters up to 5m away from the stipulated line. For
the spool-and-line search, 20 wood traps (25 x 25 x 40cm) were
installed for capturing opossums (Figure 1E). The traps were
baited with banana for two consecutive nights/month. At each
capturing point, a mixture of codfi sh liver oil (Scott Emulsion™),
papaya, banana and water was spread to attract marsupials.
Captured opossums were subjected to xenodiagnosis (7 nymphs
of T. infestans/20min), marked and equipped with spool-and-
line (IBAMA: 02023.002615/05-25). The design of these spool
devices was adapted from that of Miles33; each device consisted
of a cylindrical plastic pot with a screw cap (4 x 7cm) and a
central axis (total weight: 54g). A yellow or orange colored 914m
polypropylene line was used (Kooban™). When refuges were found,
they were manually examined, and the internal materials were
collected (e.g., leaves and twigs) for screening in the laboratory.
Noireau traps were installed in inaccessible locations32. The distance
traversed by the opossums was verifi ed by the difference in the
spool-in-line weight after a night route. Refuges with nymphs,
eggs and/or exuviae were considered P. megistus foci. Refuges
with triatomines and/or opossums were characterized.
Fresh feces collected from triatomines for xenodiagnosis
were submitted to parasitological examination (diluting in
saline solution) to verify trypanosome infection under an optical
microscope (400x). Xenodiagnosis was assessed at 15, 30 and
45 days after exposure. Triatomine feces were stored on sterile
fi lter paper (air dried and stored at -20°C) for the molecular
characterization of T. cruzi (positive samples) and food source
analyses (all samples). T. cruzi samples were identifi ed by the
presence of a nontranscribed spacer of the mini-exon gene. For
deoxyribonucleic acid (DNA) extraction, round pieces of fi lter
paper (6mm) containing feces were boiled with 50μL of ult ra-pure
distilled water (Gibco™) for 10min36. The polymerase chain reaction
(PCR) amplifi cation and reaction conditions were performed
according to the protocol of Souto et al.37, which used the following
pooled primers: TC (5’ - CCC CCC TCC CAG GCC ACA CTG),
TCI (5’ - GTG TCC GCC ACC TCC TTC GGG CC) and TCII
(5’ - CCT GCA GGC ACA CGT GTG TGT G). Amplifi cation
products were subjected to polyacrylamide gel (8%) electrophoresis
and observed by silver staining. Parasites were classified
according to the procedures of Anonymous
38 and Zingales et al.39.

577
Rev Soc Bras Med Trop
46(5):575-583, Sep-Oct, 2013
0 250 500 1
50m
50m
Transects Traps
5m
Owellings
FIGURE 1 - Location of studied area: A: Brazil (arrow indicating the Figure 1B); B: Region of Patos lagoon and surroundings (arrow: Guaíba lake basin);
C: Ponta Grossa neighborhood (arrow); D: Study area (arrow); E: Croquis drawing of the study area showing the opossum traps, transects and investigated dwellings;
F: Investigation of a hollow tree with the aid of ropes; G: Panstrongylus megistus found inside the hollow; H: Nymph (N5) captured by adhesive tape.
AB
C
D
E
F G H

578
Food sources were determined by the precipitin test
according to the methods of Lorosa et al.40. Anti-sera from the
following species were used: armadillo, bird, cat, dog, horse,
lizard, opossum, ox, pig and human. Statistical analyses were
performed using an one-way ANOVA and the post-hoc Tukey
test for the evaluation of technique effi cacy (transects and
spool-and-line). The sex ratio of infection by T. cruzi in the
reservoirs was evaluated using the chi-square test with BioStat
4.0 software.
RESULTS
There were no colonies found inside residences or in
peridomiciliary areas, and no triatomines were obtained using
light traps. Peridomiciliary inspections revealed only the
presence of adult insects, which were found on outside walls
of residences visited between November 2005 and February
2006. The largest P. megistus captures occurred in December
(Figure 2). Among the 33 triatomines collected, corresponding
to 26 (79%) males and 7 (21%) females, 28 were analyzed,
and 18 (64%) were infected with T. cruzi. Molecular analysis
characterized the parasites as T. cruzi I.
A total of 27 nymphs were collected: 26 (96%) were
analyzed and 19 (73%) were infected. Different instars of
P. megistus were found among the nymph samples at different
times of the year: 26% were 2nd instars (7 of 27), 44% were 3rd
(12 of 27), 4% were 4th (1 of 27) and 26% were 5th (7 of 27).
However, 4th instars were only observed in March (Figure 3).
All negative nymphs were found in the same refuges (dead
tree hollow). Triatomine nymphs, eggs and/or exuviae were
found only in tree refuges between 10 and 205m from the forest
fragment edge near the residences (Figure 1F, 1G and 1H).
0
2
4
6
8
10
12
14
16
Number of ndividuals
Months/years
collected
infected
female
male
Mar/06
Oct/05Nov/05 Dec /05 Jan/06 Feb/06
FIGURE 2 - Capture of Panstrongylus megistus adults throughout the months
(October 2005 to March 2006) showing the number of infected individuals and
the sex ratio of specimens from Porto Alegre, State of Rio Grande do Sul, Brazil.
0
1
2
3
4
5
6
7
Sample months
N2 N3 N4 N5
Number of ndividuals
Dec/05 Feb/06 Mar/06 Jun/06 Jul/06 Aug/06 Sep/06
FIGURE 3 - Initial instars of Panstrongylus megistus (N2 to N5: 2nd to 5th instar)
captured in the sylvatic environment in Porto Alegre, State of Rio Grande do
Sul, Brazil. N: Nymph.
The Noireau traps captured two P. megistus nymphs (3rd and
5th instars) in opossum nests. The only adult triatomine obtained
in this environment was a female found among the branches of
a tree (in February 2006) ( Table 1). Using the transect method,
33 refuges were found, of which 18% (6 of 33) demonstrated
the presence of P. megistus. Utilizing the spool-and-line method
(with 27 opossums), 28 refuges were found, and this triatomine
was present in 14% (4 of 28). There were no significant
differences (p>0.05) between the sampling methods. However,
the spool-and-line method allowed for a better understanding
of the shelters used by marsupials. A total of 61 refuges were
observed and classifi ed into fi ve categories: 10 (16%) rock
shelters, 10 (16%) dead trees, 11 (18%) ground shelters,
13 (22%) roots and 17 (28%) live trees. Among the examined
trees, seven species revealed a high number of refuges for
P. megistus and D. albiventris, including Casearia sylvestris
(chá-de-bugre), Coussapoa microcarpa (fi gueira-mata-pau),
Ficus organensis (fi gueira-de-folha-miúda), Myrsine guianensis
(capororoca), Diospyrus inconstans (maria-preta), Luehea
divaricata (açoita-cavalo) and Trichilia elegans (pau-de-
ervilha). The spool-and-line results indicated that triatomine
foci occur between 5 and 893m from the animal capture/release
location. The opossum demonstrated a wide distribution in the
area. In six refuges (four hollow trees and two rock shelters),
only opossum nests were found (no triatomines foci) between
8m and 259m from the closest residences, representing a
potential focus of P. megistus. A total of 462 traps were installed
per night, resulting in 98 captures and recaptures of 43 different
D. albiventris. Among these animals, 39 (91%) were analyzed
(26 males and 13 females), and 27 (69%) were infected with
T. cruzi. There was no signifi cant difference ( p>0.05) between
the sexes that were caught. The parasites isolated from nymphs
and opossums were characterized as T. cruzi I. Opossum
characteristics, such as semi-nomadic habits, the use of the same
nest by different individuals and two females sleeping together
in a hollow tree, were observed in fi eld specimens.
Santos Jr JE et al - Evaluation of natural foci of Panstrongylus megistus, Porto Alegre, Brazil

579
DISCUSSION
TABLE 1 - Characteristics of hollows found in the sylvatic environment (October 2005 to September 2006) showing the different stages of
Panstrongylus megistus, Trypanosoma cruzi infection and associated food sources in Porto Alegre, State of Rio Grande do Sul, Brazil.
Methods Ecotopes * Height **Traverse ***Distance #Hosts *+P. megistus T. cruzi Food sources
Transect Dead tree 2.3 - 30 Hair opossum 2 eg/exv - -
3 N3#* Pos Opossum, rodent
1 N4 Pos Armadillo
4 N5#* Pos Rodent/bird,
opossum, bird
Transect Live tree 4.0 - 145 Fly feather 2 eg/exv - -
1 N2 Pos Opossum
Transect Live tree 2.2 - 186 - 3 eg - -
Transect Live tree 4.0 - 205 Rodent 4 eg - -
Transect Dead tree 2.8 - 36 Rodent 3 N3 Pos Rodent, opossum
1 N5 Pos Bird
Transect Dead tree 1.4 - 92 - 2 N2 Neg Bird
5 N3 Neg Rodent, opossum,
bird/opossum
Spool and line Live tree 3.8 322 148 - 7 eg/exv - -
4 N2++ Pos Bird
1 N3 Pos Rodent/bird
1 N5 Pos Bird
Spool and line Dead tree 2.5 835 140 - 5 eg/exv - -
Spool and line Live tree 1.2 167 10 Opossum exv - -
Spool and line Live tree 1.0 12 10 Opossum 1 N5 Pos Bird
Spool and line Live tree 3.3 5 59 Opossum ♀## Neg Rodent
P. megistus:. Panstrongylus megistus; T. cruzi: Trypanosoma cruzi. *height of hollows from the ground (m); **distance between the point
of capture/release and opossums found (m); ***distance between the hollows and nearest residence (m); #hosts found in hollows or traces
observed; +two female opossums; *+foci found: instars bugs found in the hollows, N2 to N5: nymphs of 2nd to 5th instar; eg: eggs hatched;
exv: exuviae; #*o ne individual caught with Noireau traps; ++one individual was not examined for T. cruzi and food source; ##female found in the
tree; Pos: (positive) and Neg (negative) for T. cruzi infection.
The precipitin test was conducted in 26 P. megistus nymphs
and 29 adult insects. The nymphs were associated with four
different hosts ( bird, rodent, opossum and armadillo) in a manner
that indicating that 88% used only one feeding source and 12%
used two feeding sources. The adults were related to seven hosts
(bird, rodent, opossum, armadillo, cat, dog and lizard) such
that 77% used one food source, 20% used two hosts and only
one (3%) individual used three different host species. The bird,
rodent and opossum showed the highest frequency of serving as
hosts in both environments (sylvatic and peridomiciliar). There
were no positive reactions for human blood (Table 2).
This paper reports the fi rst discovery of a P. megistus sylvatic
focus in RS. The occurrence of triatomines in residences during
the warmer months was reported in Porto Alegre25,29, with males
being more frequently present28. Even with the presence of
early instar nymphs throughout the year, as observed in natural
ecotopes, P. megistus showed only an annual life cycle, with
adults emerging in the last months of spring. This feature is
likely attributed to a prolongation of the 5th instar due to low
mobility and decreased hematophagism in the cold of winter.
With the nearing of summer, there is an increased activity of
individuals followed by a rise in the search for food, thereby
leading to the dispersion of adult triatomines41,42. Although
domestic P. megistus colonies were not found in the evaluated
houses, the proximity of natural foci to residences could
facilitate triatomine infestation15.
Although intradomiciliary triatomine foci were not found,
the Programa de Controle da Doença de Chagas (PCDCh)
has maintained continuous epidemiological surveillance.
Rev Soc Bras Med Trop
46(5):575-583, Sep-Oct, 2013

580
The few studies performed in recent years have reported that only
adults of P. megistus have been found in homes. Currently, the
PCDCh has four PITs (Ponto de Informação de Triatomíneos)
in various parts of the city and advises residents regarding the
presence of triatomines in houses near natural areas, the cleaning
of peridomiciliary environments (avoiding potential foci) and
the installation of screen doors and windows43.
In Bambuí State of Minas Gerais, Santos Jr et al.17
demonstrated that even with a low rate of infestation in sylvatic
habitats, the colonization of domiciliary environments has
been extensive during the last three decades. The presence of
triatomines in sylvatic or, occasionally, domiciliary environments
threatens to interrupt the progress of disease control measures
in many areas given the focus of control programs on resident
species 44. These programs should implement new strategies to
prevent human-vector contact using materials impregnated with
insecticide, such as curtains and mosquito netting, and create
alternatives for the proper management of the environment and
maintenance of peridomiciliary areas45.
The lack of housing foci found in this work corroborates
data from classic publications reporting that populations of
the species in southern areas have a lower tendency to occupy
artifi cial environments6,46,47. In an evaluation of populations in
the States of Bahia, Minas Gerais and Santa Catarina, Barbosa
et al.48 observed that the occupation of artifi cial environments
may be related to a greater effi ciency in resource utilization,
presenting a demographic cline where the lowest fi tness was
TAB LE 2 - Frequency of the hosts associated with sylvatic and domestic Panstrongylus megistus in Porto Alegre, Rio Grande do Sukl, Brazil,
as demonstrated by the reaction of precipitin.
Host found
Reactions observed Total frequency of the host by environment
nymphs adults sylvatic* peridomiciliary**
Bird 10/26 (38%) 7/29 (25%) 13/30 (43%) 13/36 (37%)
Rodent 6/26 (23%) 5/29 (18%) 9/30 (30%) 8/36 (23%)
Opossum 6/26 (23%) 2/29 (7%) 7/30 (24%) 4/36 (11%)
Armadillo 1/26 (4%) 3/29 (10%) 1/30 (3%) 3/36 (8%)
Cat - 2/29 (7%) - 3/36 (8%)
Dog - 1/29 (3%) - 3/36 (8%)
Lizard - 2/29 (7%) - 2/36 (5%)
Bird/opossum 1/26 (4%) 2/29 (7%)
--
Rodent/bird 2/26 (8%) 3/29 (10%)
--
Dog/cat - 1/29 (3%)
--
Bird/rodent/dog - 1/29 (3%) --
88%+
77%+
12%+
20%+
*nymphs and one female found in the natural environment; **only adults; +frequency (%) of one or two feeding source s identifi ed.
observed in SC. Additionally, genetic and paleogeographic
studies have demonstrated that the most recent geological
events may have acted differently on the various P. megistus
populations, leading to the observed behavioral differences in
these species49.
According to Forattini6, P. megistus have a less invasive
tendency of artifi cial ecotopes in more humid climates with rainy
winters and larger preserved natural areas. Aragão46 showed that
humidity followed by lower temperatures produces a greater
wet surface permanence in residences, making such habitats
unsuitable for these triatomines. However, the presence of
P. megistus in artifi cial habitats in the States of Santa Catarina
and Paraná demonstrates the species’ ability to colonize this
environment in the southern region of the country, even at a
low intensity50,51. In the State of Rio Grande do Sul, the fi rst
domiciliary species focus was described by Di Primio30 in the
municipality of Viamão. Here, colonies of the species were
occasionally found in artifi cial environments, maintaining its
sylvatic characteristics. According to Bedin et al.22, 88% of the
P. megistus found in RS between 1996 and 2008 occurred with
residences, and 96% of these were adults.
Although found in different types of habitats, P. megistus
demonstrates a preference for arboreal habitats11,13,18. This
preferred ecotope provides an opportunity for greater contact
between this triatomine and various species of rodents,
marsupials and birds31. According to Barretto et al.12, sylvatic
colonies of P. megistus may have a high rate of infection by
Santos Jr JE et al - Evaluation of natural foci of Panstrongylus megistus, Porto Alegre, Brazil

581
T. cruzi, especially when associated with Didelphis sp. The
presence of nymphs in the reservoir nests implies a signifi cant
circulation of the parasite between vector and host, as observed
in this present study in which most of the sylvatic colonies
showed T. cruzi infection. The omnivorous habit of opossums
allows for the ingestion of insects as a nutritional alternative52.
T his insectivorous diet may act as an important route of
T. cruzi infection w hen associated with the triatomines present
in their burrows53, potentiated by semi-nomadic habits that
e nhance the possibility of encountering this vector in the natural
environment54. T. cruzi oral infection experiments demonstrated
a high infectivity toward marsupials, especially those that
ingested infected triatomines54,55.
According to Patterson et al.44, high infection rates might
be an indicator of close proximity to reservoir hosts and high
susceptibility to T. cruzi. In this paper, infection rates were
higher than in other studies conducted in RS (41%40 and
50%18 for P. megistus and 18%56 for D. albiventris). However,
the results presented herein are in agreement with studies in
other states (39-85% for P. megistus12,14 and 11-91.7% for
D. albiventris57-59). The parasite characterization performed
here allowed us to observe the involvement of D. albiventris
and P. megistus in the maintenance of the T. cruzi sylvatic
cycle. In contrast, Fernandes et al.5 6 demonstrated that although
P. megistus showed only Trypanosoma cruzi Z1 (TCI39) in Porto
Alegre, D. albiventris was found with Trypanosoma cruzi Z2
(TCII39), suggesting that the involvement of these marsupials
in domestic cycles may serve as a carrier mechanism for this
group in the natural environment.
The evaluation of food sources revealed that the main
resources used by adult and nymph triatomines were birds,
rodents and opossums. These three animals may use the same
shelters in trees, as observed by Barretto et al.11,60. Although
not analyzed in this study, the association of rodents with
infected triatomine colonies highlights their epidemiological
potential as loci, as rodents are considered potential reservoirs
in the T. cruzi sylvatic cycle. B irds, although refractory to
parasites, enable the maintenance of colonies and can act as
population growth vectors, thereby increasing the risk of disease
transmission, especially if the triatomines had previous contact
with an infected mammal61. In an analysis of populations in
the State of Minas Gerais Midwest, Villela et al.62 also found a
higher level of blood feeding on birds in triatomines captured in
peridomiciliary environments; however, the majority (90.4%) of
individual triatomines fed on only one source (bird and others
species). The same authors showed that 44% of infected bugs
had this source.
The adults showed three food sources that were lacking
nymphs: cats, dogs and lizards. Cats were found living only in the
woods due to the hostility of the local people; thus, dogs were the
only domestic animals used as a food source by triatomines. The
number of food sources indicated the eclecticism of P. megistus40,63.
The presence of anti-armadillo serum in a nymph found
in a hollow tree suggests a considerable mobility of these
triatomines in the natural environment. Forattini et al.64 observed
the mobility of 4th and 5th instar Triatoma arthurneivai nymphs,
which walked up to 10m between the studied habitats. A similar
behavior was indicated for P. megistus13. The lower occurrence
of dogs and absence of other animals, such as pigs, cattle and
humans, resulted in a low rate of infestation, confi rmed by the
absence of peridomestic colonies and a similarity of food sources
between adults and nymphs.
The forest fragment analyzed here represents the typical
arboreal species in the granite hills of Porto Alegre and,
considering its natural conditions, contributes to the maintenance
of the sylvatic characteristics of P. megistus. Thus, based on
the data presented in this study, it is suggested that sites with
similar characteristics in the city can present similar rates of
infection. Thus, sylvatic areas at greatest risk of urban effects
tend to connect humans and domestic animals to the sylvatic
cycle of the parasite. Despite the absence of colonization, the
high number of infected opossums and triatomines demonstrates
the potential risk of T. cruzi transmission in the municipality,
which suggests the importance of continued epidemiological
surveillance and management strategies in environmentally
important areas of the native city.
ACKNOWLEDGMENTS
The authors declare that there is no confl ict of interest.
CONFLICT OF INTEREST
FINANCIAL SUPPORT
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