31 july 1981J. Med. Entornol. Vol. 18, no. 4: 266-278

@ 1981 by the Bishop Museum

CHAGAS' DISEASE IN THE AMAZON BASIN111.

Ecotopes of ten triatomine bug species (Hemiptera:Reduviidae) from the vicinity of Belém,

Pará State, Brazil

M. A. Miles,l A. A. de Souza2 and M. Póvoa2

Abstract. Tracking mammals to their nests or refuges, mi-crohabitat dissection, light traps and surveillance of humandwellings were employed to identify sylvatic triatomine speciesand their natural ecotopes in the vicinity of Belêm, Pará State,Brazil. Ten species were found: Rhodnius pictipes, R. robustus, R.paraensis, Panstrongylus lignarius, P. geniculatus, P. rufotubercula-tus, Eratyrus mucronatus, Microtriatoma trinidadensis, Belminusherreri and Triatoma rubrofasciata. AlI species except P. rufo-tuberculatus (only I specimen examined), B. herreri and T.rubrofasciata were infected with Trypanosoma cruzi; Triatoma rub-roJasciata was infected with Trypanosoma conorrhini. Natural eco-topes were defined principally in terms of specific habitats, suchas burrows and.hollow logs (P. geniculatus) , palro crowns (R.pictipes, R. robustus), large hollow trees (E. mucronatus, P. lignar-jus nymphs), small arboreal cavities (P. lignarius, R. paraensis),folded leaves within nests (M. trinidadensis), beneath loose treebark (B. hlirreri), and inside buildings (T. rubrofasciata). Only 4species showed signs of obligate dependence on particularhosts. E. mucronatus was extraordinary in that nymphs fed oninvertebrates in nature, as well as on the porcupine Coendouprehensilis. AIl species except P. geniculatus and M. trinidadensiscould be maintained experimentally. R. Pictipes and T. rubro-fasciata commonly invaded houses, while P. geniculatus and P.lignarius occasionally did so. R. PictiPes bugs entering houseswere usually heavily infected with T. cruzi. Light-attracted syl-vatic R. pictipes, originating from heavily infested mucajá palms(Acrocomia sclerocarpa), were considered the primary source oflocally sporadic and acute suburban cases of Chagas' disease.The incursion of domestic vectors into the Amazon basin fromknown endemic areas is considered to threaten the region withendemic Chagas' disease. Specific suggestions are given formeasures to prevent both new sporadic cases of sylvatic originand emergent endemic foci.

The first paper of this series (Lainson et aI. 1979)reviewed information on the prevalence of Try-panosoma cruzi-like trypanosomes in wild mammalsand mau in Pará State of the Brazilian Amazonbasin. Records were also given of adult triatominebugs that were known to have invaded houses inthe city of Belém between 1967 and 1977. Thepossibility that Chagas' disease would become en-demic in the Amazon region was also discussed.

1 Pepartment of Medical Protozoology, London School of

Hygiene and Tropical Medicine, Keppel Street, London WClE7HT. (Send reprint requests to this address.)

2 Seção de Parasitologia, Instituto Evandro Chagas da Fun-

dação SESP, 66.000 Belém, Pará, Brazil.

One of our specific objectives in the Amazon ba-sin has been to determine which T. cruzi zymo-demes are enzootic there, as described in the 2ndpaper of this series (Miles et aI. 1981a). In the pres-ent paper we are concerned principally with theidentification of sylvatic triatomine species andtheir ecotopes in the vicinity of Belém, and withevaluating their potential as domestic vectors of T.CruZl.

A variety of methods have been used in theAmericas to study sylvatic triatomine bugs as fol-lows. (1) Dissectionofmicrohabitats that might harbornests or refuges of vertebrate hosts. "Microhabitatdissection" is the carefu.l, systematic dismantlingand searching of a given microhabitat. This meth-od can be highly productive when mammal andbird refuges are easily recognized, e.g., in localitieswhere there are many freestanding palms, numer-ous large epiphytic bromeliads (Aechmea sp.), woodrat (Neotoma) lodges, or bird nests (Anumbius andPhacellodomus)'(Tonn et aI. 1976b, Miles 1979). Indense tropical rain forest, however, intensive laboris required to locate mammal nests or refuges andassociated triatomine bugs. (2) Attraction to lighttraps. Light traps have been extremely successful,especially in North America, for capturing Tria-toma protracta (Uhler) and T. gerstaeckeri (St.:.l),which commonly Ay considerable distances, and inPanama in open country where there are manytriatomine-infested palms (Wood 1950, Sjogren &Ryckman 1966, Pippin 1970, Tonn 1976a, Whitlaw& Chaniotis 1978). (3) Attraction to chicken houses.Forattini et aI. (1973, 1975) pioneered this ap-proach, which gives vital information in both hu-man settlements and established endemic areas onthe invasive and colonizing capacity of sylvatic anddomestic triatomines. Such investigations have em-phasized the extraordinary rapidity with which T.sordida (St.:.l), Panstrongylus megistus (Burmeister)and Rhodnius prolixus St.:.l disperse and form newcolonies (Forattini et aI. 1973, 1975, Otero et aI.1976, Miles 1979). (4) Attraction to resting-place traps

III.

1981

Miles et ai. Triatomine ecotopes in lhe Amazon basin 267

or animal-baited traps. Resting-place traps insidehouses are of value for population estimates (Scho-field 1979); outside houses, results have been dis-appointing for the amount of labor involved(Tonn et al.-1976b). (5) Surveillance by local resi-dents, with periodic house-to-house searches, tomonitor the incursion of adult bugs (Rodrigues &MeIo 1942).

af these methods, only microhabitat dissectiongives direct information on the natural ecotopes ofsylvatic triatomines. Identification of blood mealscan be used in combination with alI of these meth-ods to identify natural hosts, providing that thetest used is sensitive, includes antibodies to the fullrange of local hosts and incorporates rigorous con-trols (Miles 1979).

Most of the triatomine bugs reported from theI

Amazon basin havt: been adultsthat have flowninto houses. The species recorded are: CavernicolaPilosa Barber; Eratyrus mucronatus Stal; Microtria-toma trinida{iensis (Lent); Panstrongylus geniculatus(Latreille); P. lignarius (Walker); P. rufotuberculatus(Champion); Rhodnius brethesi Matta; R. paraensisSherlock, Guitton & Miles; R. PictiPes Stal; R. ro-bustus Larrouse; Triatoma maculata (Erichson); T.rubrofasciata (De Geer) and T. rubrovaria (Blan-chard) (Lainson et aI. 1979, Miles 1979). Ferreira& Deane (1938) have also recorded the presence,in houses, of the hematophagous hemipteran Cler-aJa apicicornis (Lygaeidae).

After intensive but almost fruitless attempts todetermine the ecotopes of sylvatic triatomines ina forest reserve near Belém, Miles (1976) devel-oped a sim pIe method of tracking trapped mam-maIs to their refuges. A crude spool-and-line ap-paratus was fitted to animaIs released at theircapture points, and the liDe that unwound fromthe device was traced subsequently. Twenty-threetriatomine bugs of 5 species were found with 3 of15 mammals retrieved, including 1 undescribedspecies, R. paraensis (Sherlock et aI. 1977), and 1species, M. trinidadensis, that was previously un-known in Brazil (Miles 1979).

Since 1977 we have developed an improved pre:-cision-wound spool-and-line tracking device (Mileset aI. 1981b) and used it on various mammalianspecies in our attempts to locate sylvatic triatom-ines; microhabitat dissection, light traps and housesurveillance have also been employed. Here we re-port on 10 species of triatomines that have beencollected near Belém, their natural ecotopes, thepresence or absence of infections with T. cruzi, at-tempts to establish laboratory colonies, and new

records of adult triatomines invading houses.Schofield (1979) has emphasized the need for suchinformation. We describe also a suburban focus ofT. cruzi transmission that was found following de-tection of the latest human acu!e case.

On the basis of our assessment of the origin ofcases in the Brazilian Amazon basin, and of thepotencial of the local vectors to adapt to houses, weoffer suggestions for the protection of the regionagainst the threat of endemic Chagas' disease.

MATERlALS AND METHODS

Study areas

lnvestigations were performed in the followingareas. (1) The Utinga Forest Reserve on the out-skirts of the city of Belém (Fig. 1). The reserveconsists of 3 sections used, respectively, by the citywater company (COSANPA), an agricultural re-search institute (EMBRAP A) and the Brazilianarmed forces. Our observations relate principallyto the areas surrounding Agua Pre1;a lake (COS-ANP A) and to the APEG (Area de Pesquisa Ecol-ogica do Guamá) portion of the EMBRAP A re-serve. Both dry ("terra firme") forest and swamp("igapo") forest borders lake Agua Preta, whileAPEG consists of dry forest descending to tidalswamp ("várzea"). (2) The Belém suburb <;>f Utin-ga, adjacent to the Utinga Forest Reserve (but notlinked to it by forest), especially those parts of thesuburb that are close to the Army firing range andthat feature a number of mucajá palm trees (Ac-rocomia sclerocarpa) and uncut scrub. (3) A smallpatch of secondary forest on Mosqueiro lsland, 70km N of Belém, together with nearby palms andscrub adjacent to a house where there had been arecent acute case of Chagas' disease (Silveira et aI.1979). (4) The Pirelli forest 40 km E of Belém onthe road to Castanhal. (5) Forest at a Dendê palmplantation (DENP ASA) 50 km NE of Belém on theroad to Mosqueiro.

ln addition, triatomines were collected by pri-vate individuaIs in various parts of the city of Be-lém; 2 other specimens were collected by a fieldworker in the Monte Dourado area of laTi Florestal(see Results section).

Mammal tracking

AnimaIs were captured in collapsible-wire orwooden-box live traps, using a bait mixture ofmashed banana, dendê oil, peanut butter and com-mercial animal feed, together with a small piece ofjack-fruit, pineapple or banana. Traps were set ap-

1981

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268 J. Med. Entorno!. Vol.18,no.4

BRASILIA2.058 km.

Microtriatoma trtnidadensis.. Eratyrus mucronatush. BelmiQus herreri*

proximately equidistantly along a transect butplacements were influenced by the availability oflikely animal refuges. Captured animaIs were oftenanesthetized (Sagatal@, May & Baker Ltd., Dag-enham) before being fitted with a mammal-track-ing device of appropriate size and range. Fivetypes of devices were available, with ranges of2300, 1350, 1300, 280 and 160 m. The construc-tion and use of these devices is fully described else-where (Miles et aI. 1981b). When mammal trackingtook place as part of field surveys to detect T. cruziinfections, animaIs were bled under anesthesia byheart puncture (0.2 ml for blood cultures) and/or

xenodiagnosis was performed before they were re-Ieased with a tracking device attached; animaIswere aIways given a fuII night of freedom beforebeing tracked. Refuges and nests were dismantIedand examined on the spot, in the same way as inroutine dissection of microhabitats (see beIow).Nests were then taken to the Iaboratory in plasticbags for more thorough investigation.

Microhabitat di5section

AlI areas in which mammal traps were set werealso reconnoitered for possible animal refuges(e.g., palms, hollow trees and hollow logs). Palms

FIG. 1. Collection sites of triatomine species in the Utinga Forest Reserve and suburb of Utinga, Belém, ParáState, Brazil. Urban areas are heavily shaded, forest is unshaded.

R.pictipes .E.paraensis c

.!?.llgnarlus 8

'p'.genlculatu5 O

1981 MiJes et ai. Triatomine ecotopes in the Amazon basin 269

were climbed and examined thoroughly usinggloves and forceps, paying special attention tonestlike structures; attempts were made to captureany mammals in the palms. When palm trees werein areas to be cleared or in an area associated witha human case of Chagas' disease, they were cutdown. All fronds were removed and the entire treewas meticulously examined for triatomine bugs,which were collected.

Hollow trees that had cavities less than 5 m fromground levei could be found by tree-by-treesearches. The floor of such cavities and the basesof their trees were examined for animal feces, furor quills and for the conspicuous, identifiablescents of Prehensile~tailed Porcupines (Coendouprehensilis) or Collared Ante~ters (Tamandua tetra-dactyla). When possible, the:interiors of cavitieswere examined by flashlight:ébr probed to detectany bats. Live trees that had cavities were seldomdestroyed; windows were cut into the cavity withan axe, using climbing irons if necessary to scaleand grip the tree during this procedure; when thecavity was restricted to a single branch, this wasoccasionally cut orE. Mammals, their nests andloose wood from the lining of the cavity were re-moved and triatomine bugs were collected. If treeshad large internal cavities open at both ends, thecavities were entered from above by Tope; a facemask was worn for some protection against fetidodors, dust and possible contact with the agent ofhistoplasmosis. In early work, attempts were madeto locate tree cavities at canopy leveI by systemati-cally scaling alllarge trees with climbing irons. Thiswas exhausting and unproductive, however, andsuch cavities were located more readily by the trap-ping and tracking of mammals.

Hollow logs were examined by splitting themlongitudinally, by cutting windows or by crawlingthrough them with flashlight, forceps and a col-lecting vessel.

Initially attempts were made to dig out animalburrows, which are common in tropical rain for-ests. U nfortunately, many of these were uninhab-ited and extended for several metres; this ap-proach was abandoned except when an animal hadbeen tracked to the burrow and was known to beinside.

Light traPPingAlthough preliminary attempts to capture bugs

with a CDC light trap had been unsuccessful (Miles1976), reports by Whitlaw & Chaniotis (1978) ofhigh triatomine catch-rates from Panama prompt-~

ed us to reevaluate this method in the forest. Trapconstruction was as illustrated by Whitlaw & Chan-iotis (1978). Black- and white-light traps were setsimultaneously at canopy height or above the can-opy in the Utinga Forest Reserve on a 42-m scaf-folding tower. Traps were powered by a Honda300E generator and were run frc;>m dusk untildawn.

H ouse suroeillance

Dried specimens of Rhodnius PictiPes were dis-tributed to householders in the Utinga suburb ofBelém, which was known to harbor active foci ofT. cruzi transmission in palms withinlO m of earth-walled, palm-roofed houses. Householders wereasked to report the presence of bugs in houses toa local field worker. Approximately 50 houses inthe same area were searched for triatomines. Oth-er adult triatomines collected Eram houses in Be-lém were brought unsolicited to the Instituto Evan-dro Chagas by householders in the city, who werepossibly prompted by occasional preí1;s and televi-sion mention of their presence.

I dentification and examination of triatomines

Triatomine species were determined accordingto Lent & Wygodzinsky (1979); Professor Lent alsoconfirmed personally any of our equivocal identi-fications.

If numerous triatomines were collected on a giv-en occasion, a proportion was dissected to detectthe presence of T. cruzi infection. T. cruzi stockswere isolated for enzyme characterization by directculture of bug feces or mouse inoculation and sub-sequent culture of cardiac blood. If examples werefew or if the species was rare and was required forattempts to establish colonies, bugs were fed in in-dividual containers. Feces deposited were then ex-amined and treated in the same way as those ofdissected bugs.

A small number of blood meals Eram represen-tative triatomine species were precipitin tested(Boreham 1975). Blood meals were not deter-mined routinely, as frequently we did not wish tokill bugs immediately; furthermore, a full comple-ment of antisera to local- hosts was not availableand results were shown not to be entirelyreliable.We thus preferred to base our identification of thenatural hosts-,on the mammals with which the re-plete triatomines were found.

To establish colonies, bugs were placed in nylongauze-covered plastic buckets with material Eramtheir collection sites. Bugs were offered mouse

J. Med. Entornol. Vol. !.8, no. 4270

blood weekly or, if this was refused, rat or lizardblood. Established colonies were maintained inplastic buckets lined with filter paper and with fil-ter paper supports (Whatman No. 1). Wheneverpossible, welI-established colonies were fed on livechickens that were immobilized above the bucketsin a wooden cage. AlI colonies were housed in abug-proof cabinet or enclosure at 28 to 30 °C and80 to 90% RH.

b

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9

RESULTS AND DISCUSSION

Triatomine bugs collected (Fig. 2), their habitats(Fig. 3) and associated hosts are summarized inTable 1. By far lhe most common bug collectedwas R. PictiPes; 258 specimens were foond in syl-vatic locations on 35 different occasions-. N umbersof other species found in sylvatic ecotopes andnumber of occasions on which they were found (inparentheses) are as follows: R. robustus, 23 (3); R.paraensis, 4 (1); P. lignarius, 89 (19); P. geniculatus,9 (5); P. rufotuberculatus, 1; E. mucronatus, 55 (4);M. trinidade11\is, 50 (11); and Belminus herreri, 4 (2).T. rubrofasciata was never found in sylvatic habi-tats.

Most bugs were collected in lhe Utinga ForestReserve, although on Mosqueiro Island, 1 smallgroup of palm trees yielded 143 bugs (see below).The Pirelli and DENPASA forests yielded 15 R.PictiPes and 1 P. rufotuberculatus, and 1 P. genicu-tatus and 1 M. trinidadensis, respectively. Two oflhe 4 specimens of B. herreri were donated fromlhe Jari Florestal project in north Pará State.

Collection sites in lhe Utinga Forest Reserve andlhe species found there are shown in Fig. 1.

The. most productive method of locating tria-tomine bugs in their forest ecotopes was lhe track-ing of mammals to their nests and refuges, al-though bugs were neve r very numerous inindividual nests or refuges. One large hollow tree(Fig. 3a) inhabited by 2 porcupines contained well-establishe~ populations of both E. mucronatus (41specimens) and P. lignarius (16 specimens) (Table1), and infested suburban palms aIs o sporadicallyyielded substantial populations. We did not at-tempt to assess seasonal fluctuations in lhe num-bers of triatomine bugs in natural ecotopes. Ourmost successful catches were in lhe dry season afterfruit had largely disappeared Eram lhe forest floor,simply because more animaIs were trapped. Tenwhite-light and 20 black-light trap-nights, usingfluorescent tube lights at canopy leveI in lhe APEGforest, yielded a single R. Pictipes adulto The lighttraps were set in April and further studies are

e

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a

iI

I

d

II

,h

FIG. 2. Examples of the 10 triatomine bug species cap-tured to date in the vicinity of BelélD, Pará State, Brazil.Species, sex and actual length (cm) of the specimens illus-trated are as follows: a, Eraryrus mucronatw, õ, 2.6; b, Pan-strongylw geniculatw, ~, 2.6; c, Rhodniw paraensis, ~, 1.2; d,P. lignariw, ~,2.7; e, R. pictipes, ~, 2.1; f, Microtriatoma trin-idadensis, ~, 0.7; g, Belminw herreri, ~, 0.9; h, Triatoma rub-rofasciata, ~, 2.4; i, R. robustw, õ, 2.4; j, P. rufotuberculatw,õ, 2.4; k, comparative sizes of 8 of the above specimens, pho-tographed together: (1) P. lignariw, (2) R. Pictipes, (3) P. ge-niculatus, (4) M. trinidadensis, (5) B. herreri, (6) E. mucronatus,(7) R. paraensis, (8) T. rubrofasciata.

clearly needed to establish their value, which marbe seasonal. No bugs at ali were captured by housesurveiliance.

T. cruzi infection

af the 10 species captured, 6 (R. PictiPes, R. ro-bustus, P. lignarius, P. geniculatus, E. mucronatus andM. trinidadensis) were found infected with T. cruzi

14 65

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b

c

d e

f

g

h i

a

j

k

1 2 3 4 5

6 7 8

1981 MiJes et aJ. Triatomine ecotopes in lhe Amazon basin 271

c..FIG. 3. Natural ecotopes of triatomines from Belém. a, large hollow tree colonized by Eratyrus mucronatu.5

and Panstrongylus lignarius feeding from 2 resident Prehensile-tailed Porcupines (Coendou prehensilil) (APEG re-serve, Utinga); b, excavated burrow of the Nine-banded Armadillo, Dasypus novemcinctus, colonized by P. genicu-tatus (pole in left foreground is at ground levei). Field assistant is in the burrow (APEG reserve, Utinga); c,mucajá palm, Acrocomia sclerocarpa (in a suburb of Belém), a common nest gire of the Common Opossum,DidelPhil marsuPialil, and frequently infested with Rhodnius pictipes; d, arboreal open nest (i.e., not enclosed in atree cavity) of the Common Opossum, D. marsuPialil; nest was infested with R. pictipes and Microtriatoma trini-dadensil (APEG reserve, Utinga); e, shallow burrow at the base of a fallen tree-a typical terrestrial refuge ofD. marsupialil (APEG reserve, Utinga); f, 'Pau a pique' house construction in a Belém suburb with a mucajápalm harboring R. PictiPes in the distance.

or T. cruzi-like organisms. Although the 4 speci-mens of R. paraensis recorded here were not in-fected, this species is a known vector of T. cruzi inthe same ecotope (Miles 1976, 1979). R. PictiPes, R.

robustus, P. lignarius and E. mucronatus were foundto be infected more commonly than noto M. trini-dadensis was rarely found infected and lhe infec-tions were extremely light; although lhe organism

a

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272 J. Med. Entorno!. Vol. 18, no. 4

TABLE 1. Preferred habitats and principal hosts of 536 triatomine bugs captured in the vicinity ofBelém.,* Pará State (June 1977-June 1980).

HABITATSPECIES

(No. SPECIMENS) HOST(S)

Rhodnius pictiPest (258; and 14adults flying into houses)

Didelphis marsupialis; othermarsupiais and rodents

D. marsupialis

UncertainRhodnius robustust (23)

Rhodnius paraensis*** (4)

Panstrongylus lignariust (89)

Echimys chrysurus

D. marsupiali5; Coendou prehensili

Palm crowns, especially Acrocomiasclerocarpa

In dense forest: "open" nests**

Palm crowns: A. sclerocarpa,Maximiliana regia

Canopy tree cavities

Tree cavities and hollow trees:adults on tree boughs andtrunks

Terrestrial, in burrows and hollowlogs

Uncertain

Hollow trees

Panstrongylw geniculatwt (9, and 2adults flying into houses)

Panstrongylw rufotuberculatw (1)

Eratyrus mucronatwt (55)

Dasypus novemcinctus; Philanderopossum

Uncertain

c. prehensili5; nymphs witharachnids (Amblypygi)

D. marsupiali5; rodent spp.

Lizards (Thecadactylus sp.)

Rattus rattus; man

Between folded leaves within nests

Beneath tree bark

Buildings

Microtriatoma trinidadensist (50)

Belminus herreri (4)

Triatoma rubrofasciata (27, flyiugiuto hous~s)

t Some infected with Trypanosoma cruzi.* Assuming major habitats and hosts (especially arboreal hosts not susceptible to trapping) were not overlooked; excludes

results published by Miles (1976).** i.e., not within tree cavities.

*** A known vector of T. cruzi in the same ecotope, see Miles (1976, 1979).

in M. trinidadensis was indistinguishable from T.cruzi morphologically, its identity was not proven.As only 1 specimen of P. rufotuberculatus wasfound, we reei confident in stating only that, of the10 species collected, B. herreri, an apparently ob-ligate lizard feeder (see below), and T. rubrofasciatado not transmit T. cruzi in the vicinity of Belém.

Triatomine ecotopes

Rhodnius PictiPes (Fig. 2e). R. PictiPes commonlyinfested the mucajá palm (A. sclerocarpa) (Fig. 3c,1) and was also found in the inajá palm (M. regia).These palms did not always contain definite mam-mal nests or, at the time of examination, mammals.Mammals found in the palms were usually Com-mon Opossums, DidelPhis marsuPialis, the excep-tions being 1 murine opossum, Marmosa sp., 1Woolly Opossum CaluromysPhilander, and a few un-identified rodents that escaped capture. R. PictiPeswas not restricted to palm trees, however, and wasalso found within "open" (i.e., not in tree cavity)nests of D. marsupialis in dense forest. Two bloodmeal tests confirmed that R. PictiPes fed on opos-sums. It was notable that the coloration of R. pic-

tiPes blended with that of the dry bases of frondsin the crown of Acrocomia.

Rhodnius robustus (Fig. 2i). R. robustus cohabitedmucajá and inajá palm habitats with R. pictipes. Al-though the adults of these 2 species were readilydistinguished (Lent & Wygodzinsky 1979), thenymphs were less easily separated, so that thenumbers for R. robustus quoted in Table 1 mar bean underestimate. This species was found only inpalms on Mosqueiro I. The only suggestion of ahost for R. robustus is the fact that C. Philander wasfound in 1 infested inajá palmo It is not clear howR. pictipes and R. robustus compete for what ap-pears to be the sanie ecological niche. No hybridswere noted among specimens collected.

Rhodnius paraensis (Fig. 2c). R. paraensis has beenrecorded only in canopy, tree-cavity nests of theapparently rare Arboreal White-faced Spiny Rat,Echimys chrysurus. AlI stages from egg to adult werefound within nests, and eggs were laid on leavesof the nest (Miles 1976, 1979). ane nest colonizedby R. paraensis was coharnted by P. lignarius andwas used by both E. chrysurus and D. marsupialis;blood meal identification demonstrated that R.

1981

Miles et aI. Triatomine ecotopes in lhe Amazon basin 273

paraensis in this nest had last fed on the rodenthost. That this bug species was only recently dis-covered (Sherlock et aI. 1977) probably reflects thedifficulty in capturing E. chrysurus, which appearsto be entirely arboreal, living high in the canopy.

Panstrongylus lignarius (Fig. 2d). P. lignarius wasalmost invariably found on trees or within tree cav-ities. Adults, unlike E. mucronatus (see below), weremost commonly found on the surface of treeswhere their barklike dorsal coloration providedexcellent camouflage. When disturbed by fieldworkers climbing trees during the day, these bugsbecame extremely active and, though first movingtowards the source of the disturbance, subsequent-ly flew distances of up to~5 m to other trees. Treesharboring adults were fOund to be either holIowand occupied by Prehens~e-tailed Porcupines, C.prehensilis, or to have small cavities in which theCommon Opossum, D. marsupialis, was nesting. P.lignarius nymphs of alI stages were found amongcrevices in the lining of tree holes. On 2 occasionsa single nymph was found outside tree cavities,once in an open nest of D. marsupialis and once ina mucajá palm (Table 1). At canopy leveI E. chry-surus builds nests in tree holes; the nest structure isvery like that of D. marsuPialis. It was not surpris-ing, therefore, that 1 nest of this arboreal rodentwas also infested with P. lignarius. The tracking ofan adult D. marsuPialis, reported by Miles (1976),led to a nest inhabited by E. chrysurus that con-tained both R. paraensis (adults and nymphs) and2 P. lignarius nymphs. It appears, therefore, thatthe same nest mar be used by both mammalspecies. It is not clear, however, whether P. lignar-ius feeds on both Echimys and Didelphis. Of 4 bloodmeals that were identified, 3 were Eram porcupineand 1 Eram opossum.

Panstrongylus geniculatus (Fig. 2b). P. geniculatusis principalIy a terrestrial triatomine and is associ-ated particularly with armadilIo burrows (Miles1979). We have few records of P. geniculatus, un-doubtedly due to the difficulties of locating andexcavating burrows; alI records are Eram groundleveI or below. One burrow of the Nine-bandedArmadilIo, Dasypus novemcinctus, was infested; theburrow (Fig. 3b) was located by tracking the ar-madilIo, one of few that we trapped. This triatom-ine was also found in a nest of the Grey "4-eyed"Opossum, Philander opossum, in a holIow log on theforest floor; 2 unidentified nymphs, found byMiles (1976) in a nest of P. opossum in a cavity be-neath the roots of a tree, were also shown to be P.

geniculatus.

Panstrongylus rufotuberculatus (Fig. 2j). Only 1specimen of this species was found, in a tree-cavitynest of D. marsuPialis. Rodrigues & MeIo (1942)found this species with the Kinkajou, Patos flavus,in a hollow tree.

Eratyrus mucronatus (Fig. 2a). E. mucronatus wasalways found in hollow trees inhabited by C. pre-hensilis; both adults and nymphs were found. In 1large hollow tree (Fig. 3a), which was algo colo-nized by P. lignarius, Ist-3rd instars predominatedin the base of the internal cavity, while adults andlater instars were at the top of the cavity with 2porcupines. It was noted that nymphs from thebase of the tree were replete but did not containblood. They were bloated with a bluish, translucentfluid, suggesting that they had fed either on plantjuices or invertebrate hemolymph. Plant juiceswere not available in their immediate environmentbut there were severallarge arachnids (Order Am-blypygi) living in the base of the tree. One of thesewas taken to the laboratory alive and hungrynymphs of E. mucronatus fed avidly on its hemo-lymph. The E. mucronatus nymphs at the tree basedid not have easy access to the porcupines at 15 m,and mar have been there because eggs had beendisplaced from above. As far as we are aware, how-ever, this is the only record of a South Americantriatomine readily feeding on invertebrate hemo~lymph in a natural ecotope (Schofield 1979). Thissuggests to us that Eratyrus maybe an ancient bug;it differs rather strikingly from other triatomines,as is indicated by its placement in a distinct gentiscomposed of only 2 species. It is interesting that,like its natural porcupine host, Eratyrus has veryprominent surface spines. The identification of 2blood meals confirmed that E. mucronatus fed on

porcupines.Microtriatoma trinidadensis (Fig. 2f). This species

is known to be widely distributed in central andnorthern South America. As with some of the oth-er triatomine species considered here, once its nat-ural ecotope is known it becomes apparent that M.trinidadensis is adapted to a specific microhabitat.Both nymphs and adults are fourid in nests con-structed of leaves, especially nests of D. marsupialis.It is easy to overlook the presence of the bugs be-cause they are often found between the surfacesof tightly folded dry leaves. The species is dorso-ventrally flattened and is of almost precisely thesame color as its surroundings; were it not for itssmall size it would not have access to its restingplace, which presumably effectively protects itfrom its host or other predators. Identification of

1981

274 J. Med. Entorno!. Vol.18,no.4

a blood meal confirmed that M. trinidadensis fed :on opossum. M. borbai Lent & Wygodzinsky, a sim-ilar species, has beendescribed Eram the south ofBrazil, Paraná State, in bromeliads inhabited byopossums and rodents.

Belminus herieri (Fig. 2g). This species is previ-ously known only Eram the type locality in Panama,where it was collected Eram beneath tree bark. Onboth occasions when we collected B. herreri it wasassociated with a lizard, which in one case was iden-tified as Thecadactylus sp. The presence of B. herreriin Pará State greatly extends the species distribu-tion. The attachment of B. herreri to its lizard hostappears to be obligate (see below).

Triatoma rubrofasciata (Fig. 2h). T. rubrofasciata isa species that is cosmopolitan in the tropics andsubtropics, especially in maritime and riverineareas where it lives with the Black Rat, Rattus rattus.Twenty-five specimens collected between 1977 and1980 were alI adults that had flown into houses.

Experimental colonies

Both R. PictiPes and R. robustus survived quitewell in the laboratory and readily accepted mouseblood. Mortality of R. pictiPes was high initially af-ter removal Eram its natural ecotope, the mucajápalm, and had large numbers not been availablesome difficulty might have been encountered inestablishing a colony. As pointed out by Lent &Wygodzinsky (1979), R. pictipes is very adept atfeigning death when disturbed.

A small colony of R. paraensis was maintainedtemporarily and fed on mice but eventually de-clined; we were unable to collect any new speci-meus.

Small colonies of P. lignarius and E. mucronatuswere maintained for a period of 12 months, al-though there was considerable mortality amongthe P. lignarius bugs. The colonies were neglectedand could not be maintained during a 6 monthbreak in these observations; given continuous at-tention we would envisage no difficulty in estab-lishing colonies of both species.

P. geniculatus fed on mouse blood in the dark,but very reluctantly and seldom to repletion. Wewere unable to establish a cólony Eram the smallnumber of specimens collected. The natural host,D. novemcinctus, is difficult to maintain in captivityand was therefore not offered to P. geniculatus, nordid it occur to us to try what we suspect to be analternative host, P. opossum. Lent & Wygodzinsky(1979) state that 100% RH is required by P. genic-

ulatus colonies; this mar partialIy explain our lackof success.

M. trinidadensis fed readily on mice but, for rea-sons that are not clear, alI specimens died within15 days of colIection.

A pair of B. herreri was kept alive for 2 monthsby feeding on the common house gecko (Hemidac-tylus sp.), to which the bugs showed an avid re-sponse; no interest was shown in mice or opos-sums. No eggs were laid, however, and we wereobliged to discontinue our attempts to forro a col-ony. In a simulated natural ecotope with a labo-ratory colony Thecadactylus sr. there should be nodifficulty in producing an experimental colony ofthis bug species, which might have an interestingrole in the transmission of Hepatozoon and otherparasites of lizards (Osimani 1942, Rocha e Silva1975). Herrer et aI. (1954) colIected B. peruvianusfrom under tree bark and found a similar reluc-tance to feed on nonreptilian hosts.

T. rubrofasciata fed poorly on mice and at firstwe could not produce a health colony. When of-fered the laboratory rat, Rattus norvegicus, thisproblem was entirely resolved and the colony im-mediately thrived.

We conclude that only P. geniculatus, M. trini-dadensis and perhaps P. rujotuberculatus requirefurther study before adequate conditions for ex-perimental maintenance can be defined.

Ecological segregation (Table 1)The Amazon basin has a rich, diverse flora,

which is reflected particularly in its bird and insectfauna. Bat species are strikingly abundant but thenumber of other mammal species is not especialIyremarkable. The triatomine species in the Amazonbasin are not associated primarily with birds orbats, except for Cavernicola Pilosa, but with othermammals. It is not surprising, therefore, that bycomparison with the insect fauna in general, thenumber of known triatomine species is smalI.

Our detailed observations of the ecotopes of syl-vatic triatomines, in combination with attempts toestablish colonies in the laboratory, have helped usto understand their ecological separation in na-ture.

In our opinion, the ecological distribution ofthese species is more dependent upon habitat pref-erences than upon availability of specific hosts(Table 1). Thus, we can identify a predominantlyterrestrial species, P. geniculatus, and several pre-dominantly arboreal species. Among the arboreal

MiJes et aJ. Triatomine ecotopes in lhe Amazon basin 275

species, R. PictiPes and R. robustus are principallypalm inhabiting, although in dense forest with fewpalms R. PictiPes is found in opossum nests (as longas they are of open consttuction). Hollow trees orlarge hollow trunks are favored by E. mucronatus,while nymphs of P. lignarius are equally commonin large and small tree cavities and R. paraensM isfound only in small tree cavities.

M. trinidadensM also has its specific microhabitat,but in this case the niche (between folded leaves ofnests) mar be within the more general ecotopes ofpalms and hollow trees or tree cavities.

Our observations imply that dependence on aspecific host plays a lesser role in segregation, with1 or 2 notable exceptions. R. PictiPes and R. robustusprobably feed from any malilmal, and perhapsfrom birds that live in palms. E. mucronatus prob-ably can feed on a wide rangeof hosts in nature,as the nymphs feed on invertebrate hemolymph,yet in practice its usual host is probably the por-cupine c. prehensilM, which occupies the same largehollow tree habitat. Host preference is seen withT. rubrofasciata in the dramatic improvement inexperimental colonies when they were fed on Rat-tus. P. geniculatus showed extreme reluctance tofeed on rodent blood. We suspect that R. paraensMmar have a degree of host specificity to E. chrysu-rus, and perhaps to other entirely arboreal spinyrats, for it is not found in tree cavity nests of D.marsupialM. B. herreri combines a specific habitat,the trunks of loose-barked trees, with obligate de-pendence on arboreal geckos.

The small numbers of bugs found in most nestsand refuges suggest that food mar not be a limitingfactor except perhaps in palm crowns, some ofwhich were heavily infested. R. PictiPes and R. ro-bustus would arrear to compete in palms as do P.lignarius and E. mucronatus in certain hollow trees.It is not entirely clear which factors regulate thiscompetition, whether host preference or predationby mammalian hosts or by other reduviids (whichare more abundant and diverse than the triatom-ines). We have observed that P. megistus bugs at-tack each other when competing for access to afood source. Interspecific competition mediated bypheromones cannot be ruled out. The mechanismsof dispersion are not fully understood but phero-mones mar partially regulate distribution amonghabitats. Adult P. lignarius respond rapidly to vi-brations caused by movements of field workers;other species mar not require a capacity to effi-ciently locate hosts if adherent eggs (Rhodnius) are

transported on fur or feathers or if refuges likelarge hollow trees withstand lhe use of several hostgenerations. Nonetheless, host seeking and disper-sion merit further study (Schofield 1979).

Species invading houses

We have recorded 3 triatomine species flyinginto houses: R. PictiPes, P. geniculatus and T. rub-rofasciata. One adult P. lignarius has also been re-ported previously (Lainson et aI. 1979) flying intoa house in the city of Belém.

P. geniculatus and P. lignarius arrear to invadehouses rarely in the vicinity of Belém; elsewhereP. geniculatus is well known to be attracted to lightin human dwellings (Lent & Wygodzinsky 1979).Adults of R. PictiPes and T. rubrofasciata were quitecommonly captured in houses by local residents.

Of 14 adult R. PictiPes invading 11 houses, 10were male, 3 were female and the sex of I was notrecorded. Eight of the 9 examined for T. cruziwere heavily infected. Of 27 T. rubrofasciata invad-ing 19 houses, 18 were female and 9 male. Noneof the 18 examined were infected with T. cruzi but5 carried T. conorrhini, which is clearly identifiablein infected feces by its morphology (Hoare 1972).

Experimental colonies helped to assess the in-vasive capacity of Amazonian triatomines and toexplain their absence from houses. Although theyinvade houses, R. Pictipes and P. lignarius requiredselection from a small population to overcome themortality that followed removal from their naturalhabitats. Most of the other species are either notattracted to houses or, as explained above, do notcross their given habitat barriers even if they readi-ly accept alternative hosts experimentally.

Sources of acute cases ofChagas' disease

In the first paper of this series (Lainson et aI.1979), it was suggested that sylvatic bugs inyadinghouses had flown several kilometres from local for-ests and that D. marsupialis, trapped near houses,also acquired T. cruzi infection in forests and pen-etrated the suburbs to forage. The latter supposi-tion was clearly erroneous. Active foci of T. cruzitransmission have now been found within 10m ofhouses, principally involving D. marsupialis and R.PictiPes in the mucajá palm (Fig. 3c, f). Apart fromT. rubrofasciata, R. Pictipes is the triatomine mostcommonly invading houses around Belém and isalmost always very heavily infected with T. cruzi.Our attention was brought to an acute case of Cha-~

1981

J. Med. Entomol. Vol.18,no.4276

gas' disease- diagnosed on Mosqueiro I (Silveira etaI. 1979); subsequently, 3 adult R. PictiPes flew intothe house in which the case had occurred. Over100 bugs were collected from 3 palm trees within30 m of the house, and almost alI those examinedwere infected with T. cruzi. We suspect that thevast majority of the sporadic cases of Chagas' dis-ease around Belém, and possibly in the Amazonbasin, are acquired from R. PictiPes. Of the 10 casesreported from the city, however, 4 were unusualsimultaneous infections with T. cruzi zymodeme 3,probably acquired by the oral route from eithereating an armadillo (D. novemcinctU5) or food con-taminated by P. geniculatU5 (Shaw et aI. 1969, Mileset aI. 1981a). One classic acute case, with Romanas'sign, in a 15-month-old infant occurred in July1980. We have been unable to impl.icate Cleradaapicicomis as a domestic vector of T. cruzi, althoughour household searches were very limited and thissmall hemipteran could easily have escaped detec-tion (Lent 1939).

The fact that, over a ten-year period, 10 acutecases were brought to our attention suggests that,if undiagnosed or asymptomatic cases are allowedfor, there mar have been a few score in the city ofBelém during that time (Lainson et aI. 1979).

might present such a threat: R. prolixus-the prin-cipal vector north of the Amazon basin in Vene-zuela. It mar be foreseen that deforestation of theregion and subsequent increase in the number ofcultivated palms around human settlements marprovide an ideal prelude to the incursion of R.prolixus, which in Venezuela moves freely betweensylvatic and domestic ecotopes; P. megistus-theprincipal domestic vector in areas ofhighhumidityin littoral eastern Brazil. This species might be ex-pected to thrive in the Amazon basin, assumingthat it can cross the dry northeastern region; T.infestans-the most notorious of alI domestic vec-tors. It is known to be migrating northwards fromcentral Brazil (Barrett et aI. 1979), although it ap-pears to prefer less humid, and perhaps more tem-perate climates; T. sordida-a vector of Chagas'disease to mano Often peridomestic in chickenhouses or in the nests of the House Sparrow (Pas-ser domesticus), which has recently arrived in theAmazon basin (Fraiha 1977) and transports ad-herent T. sordida eggs on its feathers; T. maculata-peridomestic in Venezuela and now on the edgeof the Amazon basin in the region around the cityof Boa Vista (Territory of Roraíma); and T. bras-iliensis and T. pseudomaculata-characteristic of thevery dry northeastern Brazilian states and there-fore unlikely to have a significant impact on the

region.One or more of these domestic vectors could

spread rapidly into the Amazon region. Dispersionor importation of domestic triatomines becomesmore likely as highways, population and move-ment of people and cargo increase. The geograph-ical origin of any invading T. cruzi strains mar in-fluence the subsequent chronic manifestation ofChagas' disease, as distinct strains of T. cruzi ap-pear to be endemic to different regions (Miles etaI. 1980). We consider that implementation orstrengthening of the following measures mighthelp protect the Amazon basin against Chagas' dis-ease.

For sporadic cases oJ sylvatic origino (a) Periodic tele-vision, radio, and press reports and school educa-tion programs are needed to inform the popula-tion of the significance of triatomines, and to giveinstructions for their collection and the addressesof reference centers to which specimens should besubmitted. (b) Suburban palm trees should be sur-veyed, particularly A. sclerocarpa and M. regia, forthe presence of triatomines, with periodic insecti-cidal treatn1ent, or felliQg and insecticidal treat-ment of infested trees. (c) Further studies of in-

CONCLUSIONS

Chagas' disease is not at present endemic in theAmazon basin. There can be no doubt that this isdue to the absence among the sylvatic triatominesof species that can readily adapt to houses. Of thetriatomines recorded in the Amazon basin, wehave found alI species to be present around Belémwith the following exceptions: Cavernicola pilosa,which occurs in bat roosts only; R. brethesi; T. rub-rovaria; 2 new species yet to be described (Serra etaI. 1980a, h); T. maculata, which is peridomesticand restricted to the area adjacent to the Vene-zuelan border; and R. prolixus, the natural pres-ence of which is, in our opinion, dubious owing tothe difficulty in distinguishing small R. robustusEram large R. prolixus. Nonetheless, the latter 2species are quite valid; both occur in Venezuela,but whereas R. prolixus moves freely between syl-vatic and domestic habitats, R. robustus is fortu-nately not a domestic species (Lent & Wygodzinsky1979).

Although studies of autochthonous species arefar from complete, the main threat of Chagas' dis-eas~ clearly lies in the incursion of domestic vectorsEram present endemic regions. The following do-mestic vectors occur close to the Amazon basin and

1981 MiJes et aJ. 277Triatomine ecotopes in the Amazon basin

digenous sylvatic vectors and their ecology are

required.For emergent endemic foci. (a) Rigorous insecticidal

control of any incipient household colonies of localsylvatic species is essential. Submission of nymphs,rather than adults, to a reference center (which, asfar as we are aware, has not yet occurred) wouldimmediately imply the presence of such acolony.(b) Endemic Chagas' disease in areas around theAmazon basin should be controlled using estab-lished insecticides and resident-assisted surveil-lance and house improvement (self-reliant meth-ods) (Ferrer 1976, Rocha e Silva et aI. 1979). (c)The dispersion of known domestic vectors fromendemic areas should be monitored, particularlyalong new highways or where rural settlementcrosses natural geographical barriers. (d) Exami-nation and insecticidal spraying of the householdpossessions of immigrants from endemic areas isrequired. (e) Periodic serological surveys of schoolchildren should be conducted to detect emergentendemic foci. (f) Blood donors should be moni-tored or donor blood treated.

Although Chagas' disease is not currently a ma.,jor public health problem in the Amazon basin,between .4 and 7 million individuaIs are infectedelsewhere in Brazil. For an area undergoing rapiddevelopment there are clearly certain public healthpriorities, particularly in this case that of malaria,which is highly endemic and locally highly resistantto therapy. Nonetheless, reasonably strong mea-sures are indicated to protect future inhabitantsagainst the mortality and morbidity of Chagas' dis-ease.

Acknowledgments. We especially thank Fo S.Gomes and Ro N.de Almeida for their assistance in lhe field. We algo thank ourcolleagues at lhe Instituto Evandro Chagas, Fundação SESP, inparticular Dr Habib Fraiha for his encouragement and support,J. A. Nunes Lima for technical assistance and Dr P. D. Readyfor comments on lhe manuscript. Mrs J o E. Miles assisted withpreparation of lhe manuscript. We are grateful to lhe followingfor access to study areas: lhe Directors of lhe Companhia deSaneamento do Pará (COSANPA), Empresa Brasileira de Pes-quisa Agropecuária (EMBRAPA), Guamá Agro Industrial (Pi-relli), Dendê do Pará (DENPASA), Dr Miguel Brasil (Mosquei-ro); and lhe commanders of lhe 8th Military Region (Belém).We very much appreciate financial support from lhe WellcomeTrust (London) and Fundação SESP (Brazil).

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