Journal of the American Mosquito Control Association, 14(3):219-233, 1998Copyright O 1998 by the American Mosquito Control Association, Inc.

MOSQUITO VECTOR CONTROL AND BIOLOGY IN LATIN AMERICA-AN EIGHTH SYMPOSIUM

GARY G. CLARK1 AND YADIRA N. RANGEL, (ORGANIZERS)

ABSTRACT. The eighth annual Latin American symposium presented by the American Mosquito ControlAssociation (AMCA) was held as part of the 64th Annual Meeting in Sparks, NV, in March 1998. The principalobjective, as for the previous 7 symposia, was to promote participation in the AMCA by vector control spe-cialists, public health workers, and academicians from Latin America. This publication includes sumrnaries of29 presentations that were given in Spanish by participants from 10 countries in Latin America. Topics addressedin the symposium included results from chemical and biological control programs and studies, studies of insec-ticide resistance, and molecular and ecological studies of vectors of dengue (Aedes aegypti), malaria (Anophelesalbimanus and Anopheles aquasalis), and leishmaniasis (LutTomyia).

KEY WORDS Mosquitoes, mosquito control, Aedes, Anopheles, Culex, Lutzomyia, resistance

INTRODUCTION

The American Mosquito Control Association(AMCA) is dedicated to the study and control ofmosquitoes and other vectors and promotes coop-eration and interaction among professionals andstudents in this field. Although the majority ofAMCA members are from the USA, an increasingnumber of members and almost one half of the con-tributors to the Journal of the AMCA are from out-side the USA. To meet the needs of one segmentof the international membership, the first Spanishlanguage symposium was held at the AMCA An-nual Meeting in 1991 to provide a scientific forumfor scientists whose primary language is Spanish.In addition to addressing the language issue, thesession promotes interaction and esprit de corpswithin this group; interaction with control industryrepresentatives; and interaction with professionalcolleagues in the USA who are involved in mos-quito vector control, training, and research at theuniversity level, and state and federal governmentofficials.

This publication includes summaries of 29 pre-sentations that were given in Spanish by partici-pants from 10 countries in Latin America. Topicsaddressed in the symposium included results fromchemical and biological control programs and stud-ies, studies of insecticide resistance, and molecularand ecological studies of vectors of dengue (Aedesaegypti (L.)), malaria (Anopheles albimanus Wie-demann and Anopheles aquasalis Curry), and leish-maniasis (Lutzomyia). Summaries of 6 previoussymposia have been published (Clark and Suarez1991, 1992, 1993; Clark 1995, 1996: Clark andRangel 1997).

Financial support for this session was providedby the following sponsors and individuals: Arro-Gun (Dan Anaz), Superior Angram (Juan Angulo),

rDengue Branch, Centers for Disease Control and pre-vention, 2 Calle Casia, San Juan, PR OO92I-32C[..

2 Laboratorio Biologia de Vectores, Instituto de Zoolo-gia Tropical, Universidad Central de Venezuela, Caracas,Venezuela.

Van Waters and Rogers (Ed Meehan), Zanus Cor-poration (David Sullivan), Zeneca (Eduardo Mor-eira), the Pan American Health Organization, andthe Rockefeller Foundation. Funds from thesesponsors partially defrayed travel and meeting ex-penses of symposium participants. Enthusiasm andinterest for this symposium was high and this sym-posium will be a part of future meetings.

SUMMARIES

Colonization and release of aquaticpredators with Bactimos briquets forcombined control of mosquito larvae

C. Solis, H. Quiroz, V. A. Rodriguez, M. H. Badii,R. M. Gonzalez, D. Martinez. K. Arevalo and

L. O. Tejada

I-aboratorio Entomologia, Universidad Aut6nomade Nuevo Le6n (UANL), San Nicolas de los Garza,

Nuevo l-c6n, Mexico

In public health, more biorational approaches areneeded to manage mosquito populations. One ap-proach combines a specific chemical insecticide formosquitoes and one or more biological controlagents effective against these insects. The efficacyof2 aquatic predators (the dytiscid Laccophilusfas-ciatus, and the damselfly Archilestes califurnica)and Bacillus thuringiensis var. israelensis (B.t.i.)(Bactimoso) briquets were evaluated as 2 mosquitolarvae control strategies in plastic containers. Astudy design with 4 strategies was developed: Bac-timos added at a rate of one briquet per 9.29 m2,Bactimos plus 5 dytiscid adults, 5 dytiscids, and anunteated control. Natural oviposition of A. califur-nica occurred in the containers, and they were re-moved from the control. Larval population densi-ties were measured weekly by taking 10 dippersamples from each treatment. An analysis of vari-ance was used to detect treatment differences. Thetreatments with lowest larval densities were theB.r.i. alone when the pooled data of mosquito lar-vae were atalyzed and combined action of the bac-teria-aquatic predators when larval Culex spp. den-sities were analyzed,. No difference was found for

219

220 Jounr.r{- oF THE AMERIcAN Moseurro CoxrnoriAssoclATtoN VoL. 14, No. 3

the numbers of Anopheles pseudopunctipennis be-tween the 2 strategies and the untreated control. Weconcluded that the natural mortality caused byaquatic predators used with biorational products of-fered good results in short- and long-term vectorcontrol programs.

Influence of filamentous algae onAnophele s ps eudopunctip ennis

J. G. Bond-Compean, J. I. Arredondo-Jimenez andM. H. Rodriguez

Centro de Investigaci6n de Paludismo, InstitutoNacional Salud (INSP), Chiapas, Mixico and Cen-tro de Investigaciones Sobre de Enfermedades In-

fecciosas, INSP, Cuernavaca, Morelos, Mexico

Anopheles pseudopunctipennis Theobald is a pri-mary malaria vector in Mexico and breeds almostexclusively in association with filamentous greenalgal mats that occw in pools of receding rivers.To date, few experimental studies have investigatedthe ecological relationships between anopheline lar-vae and associated vegetation. The objectives ofthis study were to assess the effect of filamentousalgal coverage on An. pseudopunctipezzis survivaland predation rates and its influence on pupationtime and adult size; to assess the influence of thealgae in the selection of habitat for oviposition bygravid females, and to assess the role of algae as afood resource. The study was carried out from Junethrough September 1997 in a fish pond of Caca-hoata, Chiapas, Mexico. For the first objective, aseries of factorial experiments were undertakenwith floating enclosures, testing 4 algal coveragesand 4 fish predator densities (Gambusia affinisBaird and Girard). For the 2nd objective, a factorialexperiment with floating oviposition substrates withand without algae of 2 genera (Spirogyra and Cla-dophora) was carried out. To accomplish the lastobjective, the feeding habits of larval An. pseu.do-punctipennis were assessed by analyzing the larvalguts with 4'6-diamidino-2-phenylindole.2HCl(DAPI) staining.

Results demonstrated that Spirogyra aided thesurvival of An. pseudopunctipennis larvae by pro-viding shelter from G. afinis predations, with apositive correlation between number of mosquitolarvae and algal coverage and negative correlationwith respect to fish density. Where high densitiesof fish predators were provided, the pupation timewas increased and adults tended to be smaller insize. Significant differences occured in mosquitosurvival in treatments with and without algae (F :24.01, df : 3, P < 0.001) and between presenceor absence offish (F : 168.08, df: 3, P < 0.001).Similarly, the presence of algae favored ovipositionby females (F : 6.76, df : 1, P : 0.01), whereasno oviposition was observed in open water. No dif-ferences were observed in the selection of any algalspecies (Spirogyra or Cladophora) as oviposition

sites (F : 0.15, df : 1, P : O.7O). Larval gutcontent analysis demonstrated that filamentous al-gae (Spirogyra) may be very important as a foodsource because they comprised 47Vo of all particlesin the gut of 1,600 An. pseudopunctipennis larvaesampled. In conclusion, filamentous green algaeprovide oviposition substrates for An. pseudopunc-tipennis gravid females, as well possible as foodand shelter for the mosquito larvae.

Community biological control of malaria usingBacillus thuringiensh v ar. israek nsrs<oconut in

Piura, Peru

P Ventosilla, E. Reyes, J.Yelez, H. Guena andP Novak

Instituto Medicina Tropical "Alexandre von Hum-boldt," Universidad Peruana, Cayetano Heredia,and Laboratorio Control Biol6gico de Vectores, In-

stituto Nacional de Salud, Lima, Peru

The number of malaria cases recorded by theMinistry of Health in Peru has been increasingsince 1990. In L996,208,543 cases of malaria wereconfirmed, of which 49,962 were Plasmodiumfal-ciparum. Most cases of both types were reportedfrom the states of Piura and Loreto. The purposeof this project has been to develop, test, and pro-mote a set of techniques using Bacillus thuringien-sis var. israelensis II-14 (B.t.i.) cultured in wholecoconuts for larval vector control in malaria en-demic areas. The goal is to eventually have thesedelivered in a sustainable fashion through directcommunity participation.

The intervention was implemented in the hamletof Salitral, District of Sullana in Department of Piu-ra state on the northern coast of Peru. Informationabout the community's knowledge and perceptionof malaria was developed to aid in the design of aneffective educational strategy. Malaria was notviewed by the community as a serious problem.With this information, educational modules and ap-propriate tools (comics, videos, theate! and so on)to convey the messages were designed. We foundthat there was limited community participation andthe school was the only dynamic entity in the com-munity.

The schools appear to be a good location for theproject to continue. A youth group, ABIMSA,which had been active in health education, formedthe project's field educational component. After theeducational intervention, a survey of 5O schoolchildren gave the following results: lNVo of t}rLebrigade members knew the effects of B.t.i. action,827o communicated their acquired knowledge totheir families (using direct observation techniques),60Vo h,new all stages of the B.r.l. cultivation pro-cedure, 54Va cortectly identified the malaria trans-mission cycle, and 25Vo werc able to distinguishbetween Anopheles and Culex larvae. Correctknowledge about malaria rose from tO to 32Vo. A

SepT el{sen 1998 Moseuno Vecron Svunosrul,r 221

positive correlation was found between accurateknowledge of malaria transmission and the pres-ence of a participating student in households. Morethan SOVo of the residents surveyed were in favorof continuing the experience with the students inorder to "better control the malaria problem" andto "share knowledge about the problem with therest of the community."

In the coconut-B.r.l. produced by the communi-ty, the average number of B.t,i. spores per coconutwas about 2.2 x l0s spores/ml. The average lar-vicidal effect in ponds was 1007o mortality of lar-vae at 6 and 12 h with a concentration of 2.2 x lUspores/rnl. One group of the ponds received 2 co-conuts and others received 4 coconuts and theAnopheles albimanus Wiedemann population de-creased after 96 h. The percentage of mortality wasbetween 80.7 and 89.4Vo. The anopheline larvae re-mained largely absent for 12 days after this period,whereas the larval population of the untreatedponds recovered normally.

The use of biological control of Anopheles vec-tors, supported by active community participation,could complement the National Malaria ControlProgram of the Ministry of Health, which focuseson case detection and limited use of chemical in-secticides.

Residual activity of organophosphatcinsecticides on five types of surfaces on

Anopheles aquasalis

D. Molina de Fernandez, E Saume and A. Gutierrez

Escuela Malariologia y Saneamiento Ambiental"Dr. Arnoldo Gabald6n," Ministerio de Sanidad yAsistencia Social, and Facultad de Agronomia,

Universidad Central de Venezuela, Maracay,Aragua, Venezuela

The surface where insecticides are applied canaffect their performance and persistence so it is im-portant to evaluate insecticidal activity on differenttypes of surfaces. We evaluated different surfacesthat are normally found in the field using the in-secticides malathion, fenitrothion, fenthion, andmethyl pirimiphos, and tested the surfaces and in-secticides with the mosquito species subject to con-trol. Residual activity of the organophosphorus in-secticides was measured under laboratory condi-tions on 5 types of surfaces for 180 days. The sur-faces (mud, mud with lime, cement, cement withlime, and painted cement) were sprayed with theinsecticides as wettable powder at a concentrationof 2 glm2. We tested adtit Anopheles aquasalisCurry from Yaguaraparo in Sucre State, Venezuela,following the method of the World Health Organi-zation. We used the effectivity index (EI) where theEI = number of dead insects/number of exposedinsects X time of exposure. Comparisons betweentypes of surfaces and insecticides were made withthe analysis of variance of Kruskal-Wallis. We

found significant differences between insecticides(H : 107.85; P : 0.00001; df = 9) and betweenthe types of surfaces (H : 9.658i P = 0.046). Thegreatest EI values were obtained for painted ce-ment, and among the insecticides the most effectivewas methyl pirimiphos (2.53), followed by fenitro-thion (0.52), fenthion (O.32), and malathion (O.22).Recent studies have found that populations of Az.aquasalis from Sucre State were resistant to DDTand pyrethroids but were susceptible to organo-phosphorus insecticides. These facts and the resultsof this evaluation show that methyl pirimiphos isthe best choice. It should be used in a plan of in-secticide rotation not only because of its persistenceand biological activity but also because it could beused to counteract pyrethroid resistance.

Residual activity of indoor application of teninsecticides against Anophelos darlingi in

Itupiranga, Para State, Brazil

I. A. Braga, A. K. R. Galardo, G. Calderon andM. Nelson

Fundacion Nacional Saude (FNS) and Organiza-ci6n Panamericana de la Salud (OPS), Brasilia,

Brazil

In order to determine the residual activity of in-secticides for the control of Anopheles darlingiRoot, the principal vector of malaria in Brazil, 9insecticides in l0 formulations were compared withDDT in Itupiranga, State of Para, Brazil, fromMarch 1995 until June 1996.

Standard World Health Organization susceptibil-ity tests with impregnated papers were performedwith female mosquitoes from the study area. Conebioassays were conducted on unpainted wood andmud panels and house walls sprayed with the fol-lowing formulations: organochlorines: DDT wetta-ble powder (WP) at 2 g active ingredient (Al/m'�;organophosphates: chlorpyrifos emulsifiable con-centrate (EC) at 2 g AUm2, fenitrothion WP at 2 gAVm2, malathion latex paint formulation at 2 g AUm2; carbamates: bendiocarb WP at 0.2 g AUrrP; py-rethroids: alpha-cypermethrin soluble concentrate(FW) at 40 mg AVm2, cyfluthrin WP at 50 mg AVm2, deltamethrin soluble concentrate (SC) at 25 mgAVfi;f , etofenprox WP at O.4 g AUmz, lambda-cy-halothrin microencapsulated at 3O mg AVm2, andlambda-cyhalothrin WP at 30 mg AUmz.

Mortality in the susceptibility tests was lNVo atthe concentrations and exposure times utilized, withthe exception of DDT and alpha-cypermethrin. Inthe bioassays on sprayed surfaces, etofenprox hada longer residual action than the other insecticideson wood panels (10O7o mortality for >12 months)and the longest residual action on house walls(>lOVo mortality for >12 months). Four other for-mulations (cyfluthrin WP, deltamethrin SC, lambda-cyhalothrin WB and malathion paint) showed mor-tality of 9O-lOOVo for >t2 months on wood panels,

222 Jouruqll oF TrG AMERICAN Moseuno CoNTRoL AssocrerroN Vor. 14, No.3

but none of them lasted even 9 months on housewalls. Results on mud surfaces were poor. Only cy-fluthrin and deltamethrin gave mortalities of >8OVofor I month, and after 6 months only etofenproxgave a mortality of >70Vo.

Evaluation of deltamethrin, lambda-cyhalothrin,and etofenprox against Anopheles darlingi

using the Colombian curtain in Brazil

I. Braga, A. K. R. Galardo, G. Calderon, F Santos,R. Zimmerman, D. Bown and M. Nelson

Fundacion Nacional Saude (FNS) and Organiza-ci6n Panamericana de la Salud (OPS), Brasilia,

Brazil an"d Managua, Nicaragua

Our findings demonstrate that although delta-methrin, lambda-cyhalothrin, and etofenprox wereall found to produce good residual activity and suc-cessfully reduced indoor feeding, etofenprox wasthe most effective in reducing overall feeding andsurvival of fed mosquitoes. Although, overall mos-quito exit patterns from lambda-cyhalothrin- anddeltamethrin-sprayed houses indicated a lack of re-pellency, high exit rates produced by etofenprox to-gether with highly significant mortality rates, sug-gests an excito/irritance effect. Although vectorcontact with etofenprox was reduced, fed and unfedmosquitoes still maintained a higher degree of con-tact to produce a significant mortality, thereby re-ducing feeding success. Finally, these results showthat by an objective evaluation of vector--controlrelationships, a better understanding of the impactthat insecticides have on vector behavior and theirpotential role in malaria transmission and controlcan be achieved.

A two-year field evaluation of the impact ofdifferent resistance management strategies oninsecticide resistance in Anopheles albimanas

A. Rodriguez, P Penilla, A. Trejo, G. Fuentes,J. Hemingway and M. Rodriguez

Centro de Investigaci6n de Paludismo, InstitutoNacional Salud (INSP), Chiapas, M€xico; Schoolof Pure and Applied Biology, University of Wales,Cardffi United Kingdom; and Centro de Investi-gaciones Sobre de Enfermedades Infecciosas,

INSP, Cuemavaca, Morelos, Mexico

Since 1995, in southern Mexico we have under-taken annual rotations of unrelated insecticides(ROT), mosaic spraying (MOS) of 2 unrelated in-secticides in the same area, and traditional sprayingwith a single insecticide to compare the efficaciesof these strategies. The ROT scheme included anorganophosphorus (OP) during the first year, py-rethroid (PYR) during the 2nd, and a carbamate(CA) that is curently being applied during the 3rdyear. The MOS included the application of an OP

and a PYR in the same locations during the 3 years.Ttvo controls where traditional spraying with DDTand a PYR were included. Susceptibility tests(World Health Organization IWHOI) and biochem-ical assays have been used to monitor the suscep-tibility of Anopheles albimanus populations and theresistance mechanisms involved. High levels ofDDT resistance and low levels to OP, CA, and PYRinsecticides were detected by the WHO discrimi-nating dosage procedure before the intervention be-gan. The resistance to DDT was caused by elevatedglutathione-S-transferase (GST) levels. The DDTresistance decreased slightly after the first sprayingof OP in the ROT and OP and PYR in the MOS.However, DDT resistance increased again after the2nd year, when the shift to PYR in the ROT and a2nd year of constant use of OP and PYR in theMOS took place. The constant use of DDT andPYR in the controls increased PYR resistancequickly as compared to the ROT and MOS strate-gies. The higher increase in OP resistance in DDT-sprayed villages as compared to the ROT- andMOS-treated villages could be caused either be-cause GST is responsible for a secondary OP me-tabolism, or because of intensive OP use for agri-cultural purposes in these areas. Both of these pos-sibilities .re now being investigated. Changes inCA susceptibility reflected slight inhibition of ace-tylcholinesterase by biochemical assays.

Conventional and selective intradomiciliaryapplication of DDT and Bendiocarb for the con-trol of Anopheles pseudopunctipezzis Theobald

in southern Mexico

M. Casas, J. Torres, M. H. Rodriguez,J. I. Arredondo and D. N. Bown

Centro de Investigaci6n de Paludismo, InstitutoNacional Salud (INSP), Chiapas, Mdxico, and Or-ganizaci6n Panamericana de la Salud (OPS), Ma-

nagua, Nicaragua

The main objective of the study was to evaluatethe pre- and post-indoor feeding behavior and mor-tality of Anopheles pseudopunctipennis followingcontact with conventional and selective applicationsof insecticide on indoor sprayable surfaces of ex-perimental houses. The fully treated house (FTH)as well as the selectively treated house (STH) con-tained standard dosages of bendiocarb (0.4 g activeingredient [AIVm). The selective application con-sisted of 2 0.8-m wide horizontal swaths of insec-ticide, the first starting at 0.85 m from the base ofthe walls and extending up to 1.65 m in width andtlre 2nd starting at the base of the inner roofs ex-tending upwards. Residual activity on wood re-mained >9OVo in both the FTH and STH for 3months. Monality due to contact with tin remained)75Vo for 2 months in the FTH. whereas in theSTH monality could be observed only I month af-ter insecticide spray. Vector-human contact in the

SnpreMsrn 1998 Moseurro VEcroR Svlrposruv

FTH significantly increased nearly 6-fold outdoors(49.2 bites/man/night [bmn]), while remainingnearly the same indoors (8.3 bmn) as comparedwith the pretreatment and untreated house (UH),respectively. In the STH, a significant increase ofnearly 4-fold occurred in the indoor bmn (30.7) andmore than a 3-fold increase occurred in the outdoorbiting rate (56.2 bmn) in relation to the UH. Nosignificant differences in mortality rates were re-corded in outdoor and indoor biting mosquitoes,which ranged from 25.5 to 38.tVo in both FTH andSTH. Mortality in insecticide-treated houses wassignificantly higher than pretreatment and controllevels (12.5-15.6%o). Three hours after release inthe curtained houses, 81.3 and 84.67o of An. pseu-dopunctipennrs had exited the FTH and STH, re-spectively, compared to 77.7Vo in the UH. Despitelow proportions of fed mosquitoes exiting housesbetween 2100 and 24OO h, significant differenceswere found in mosquito feeding success from FTHand STH (2.3 and l.OVo, respectively), as comparedwith UH (8.3Vo). Prefed and postfed mosquitoescontinued to land within the preferred resting heightrange on sprayed surfaces. After mosquitoes had ablood meal, reductions in total resting time onsprayed surfaces were observed with respect to UHin the FTH and STH, from 1.4- to 2.9-fold, respec-tively. Finally, the present data demonstrated that acombination of the feeding status of the mosquito,the type of insecticide application, and the effectproduced by the insecticide are complementary fac-tors that are instrumental in determining mosquitocontact with insecticide. This was different for ben-diocarb and DDT

Cross-resistance to Malathion in Cuban Czlerquinquefascialzs induced by larval selection

with deltamethrin

J. A. Bisset, M. M. Rodriguez and A. Soca

Laboratory of Toxicology and Genetics, Depart-ment of Vector Control, Institute of Tropical Med-

icine "Pedro Kourt," Havana, Cuba

A strain of the southern house mosquito, Culexquinquefascialzs Say, from Cuba was subjected todeltamethrin selection to evaluate the usefulness ofthis pyrethroid insecticide for mosquito control. Ahigh level of resistance developed after 4 genera-tions of selection. Little or no cross-resistance wasobserved to other pyrethroids (lamMa-cyhalothrinand cypermethrin), a carbamate (propoxur), and 2organophosphate insecticides (chlorpyrifos andmethyl pyrimiphos), but high cross-resistance wasfound to the organophosphate Malathion. The highresistance to deltamethrin observed after only 4generations of selection indicated that this insecti-cide was not a good candidate for long-term mos-quito control in Cuba. This conclusion was basedon the widespread Malathion resistance existing inCuba and the cross-resistance between deltamethrin

and Malathion observed in the curent studv. Pos-sible resistance mechanisms responsible ftr thisphenomenon are discussed.

Cyfluthrin for the chemical control ofAedes aegypfi in Buga, Colombia

M. E Suarez. C. A. Morales and R. I. Nino

Escuela de Salud Priblica, Universidad del Valle,Cali, and Unidad Ejecutora Saneamiento, Buga,

Colombia

The incidence of dengue hemorrhagic fever is onthe rise in Colombia. Vector control is the mostimportant means of reducing dengue risk. Adulti-cide with chemicals using ultra-low volume (ULV)application has been the most widely used methodfor dengue control. The following were results ofthe ULV application of Cyfluthrin (SOLFAC@ EC50) with Maxipros@ and with Stihlo backpackequipment. We carried out a presurvey that wasthen compared to the postsurvey after Cyfluthrinapplication. Results showed a reduction of 2O.4Vowith the Maxipros treatment and 94.4Vo with thebackpack. The average number of mosquitoes perdwelling in the Maxipros-treated area was reducedfrom 2.4 before treatment to 2.1 after treatment. Incontrast, with the backpack treatment, the averagenumber of mosquitoes per dwelling was greatly re-duced from 2.8 to O.1. The bioassays in both areashad the following monality rates results: 24Vo withMaxipros and IOOVo with the backpack. These re-sults indicate that the backpack method (internaltreatment) is more effective in reducing adult Aedesaegypti density when used in the dwelling.

Low-volume insecticide spray as an alternativemethod for the control of Aedes aegtpti

J. I. Aredondo-Jimenez, J. A, Martinez-Ibarra andR. Danis-Lozano

Centro de Investigaci6n Paludismo, Instituto Na-cional Salud (INSP), Chiapas, Mdxico, and Centrode Investigaciones Sobre de Enfermedades Infec-

ciosas, INSP, Cuernavaca, Morelos, Mexico

The resurgence of dengue fever and the rise inthe incidence of dengue hemonhagic fever in Mex-ico are in part a consequence of the failure of cur-rent measures to control the mosquito ve,ctor, Aedesaegypti (L.). Indoor residual spray of insecticidemight be a better tactic, but its feasibility is limitedby the high costs and timing of application. Theaim of this study was to optimize indoor residualspray of insecticide, applying it as a low-volumespray using a synthetic pyrethroid (lamMa-cyhal-othrin, OMS 3021, Icon@ l0 CS) against Ae. ae-gypti in Huixtla, Chiapas, Mexico (15'09'18'N,92o28'3O"W; population 45,000). Low-volumespray consisted of the 5O to 1o0-pm droplets at 25mg of active ingredienVm2 of pyrethroid on indoor

JounNel oF THE AMERTcAN Moseurro CoNrRor- AssocnrroN Vol. 14, No. 3

walls at a rate of 215 mVmin. Within the city, in-secticide was applied to houses on 25 blocks,whereas the houses on 66 surrounding blocks wereleft untreated.

The insecticidal effect was evaluated by its im-pact on larval aedine infestation indices as well asindices for adult mosquitoes and oviposition rates.The insecticide's residual effect on various wallsurfaces was also determined. During the 55 wk ofstudy, the first 34 wk allowed the assessment of thenatural fluctuations of Ae. aegyprl populations. In-secticide was applied during weeks 35-37, at a rateof 20 houses/sprayman/day. Each house was treatedfor an average of 10 min; including the actualspray, removal of furniture and house appliances,and protection of food. Insecticide residual effecton all wall surfaces tested caused mosquito mor-talities >75Vo for up to 19 wk. All aedine indiceswere significantly reduced in insecticide treated vs.untreated areas, respectively: house index for larvae(ix arcsin : 0.075 andO.297; r: 9.08, P : 0.001),Breteau index (i arcsin : 0.083 and 0.367; 1 :7.36, P = 0.001), house index for adults (I. arcsin: 0.04 andO.47; t : 9.81, P : 0.001), mean adultsper house (i arcsin : 0.007 and O.O42; t : 4.77,P : 0.00O1), and oviposition rates (x arcsin :0.192 and O.864; t : 13.74, P : 0.001), except thecontainer index (i arcsin : 0.04 and 0.03 in treatedvs. control areas, respectively, t : 1.54, P : 0.013).In conclusion, the pyrethroid applied by low-vol-ume spray had a significant impact on thLe Ae. ae-gypti population. This vector control strategy couldbe implemented in particular during dengue out-breaks, because it can suppress adult mosquito pop-ulations for up to 12 wk.

Integrated control of Culex quinquefasciatusfrom sewers in Cali, Colombia using

Bacillus sphaericus

M. E Suarez and C. A. Morales

Escuela de Salud Ptiblica, Universidad del Valle,Cali. Colombia

Storm sewers are the principal production sitesfor Culex quinquefasciatas Say and, Aedes aegypti(L.) mosquitoes in Cali, Colombia. Larvicidingwith chemicals has been widely used by the gov-ernment throughout the country for reducing adultmosquito abundance. However, insecticide resis-tance in some mosquito populations and environ-mental concern suggest the need for greater balancein the program. We present preliminary results con-cerning the susceptibility of Cx. quinquefasciatusto Vectolex@ and its impact on wild populations ofCx. quinquefasciatus. The baseline of susceptibilityfor Cx. quinquefasciatus to Bacillus sphaericus(sftain 2362'. Vectolex, Abbott Laboratories. Chi-cago, IL) was a median lethal concentration of0.015 mg/L. A pilot fleld trial using 20 sewers each

treated with 5 g of granular formulations of Vec-tolex showed the following residual effect of mos-quito mortality. Ttventy-four hours after Vectolexapplication, the larval mortality was 96Vo. After 48h and 8 days of treatment the mortality was 99Vo.We continued monitoring the residual effect of Vec-tolex that is particularly effective against Culexlar-vae.

Validation of the howe and Maya indices toestablish risk of Aedes aegypti populations

R. D. Lozano, J. I. Arredondo-Jimenez,J. A. Martinez Ibarra, J. G. Bond-Compean and

M. H. Rodriguez

Centro de Investigaci6n de Paludismo, InstitutoNacional Salud (INSP), Chiapas, Mdxico, and Cen-tro de Investigaciones Sobre de Enfermedades In-

fecciosas, INSP, Cuernavaca, Morelos, Mexico

Better aedine indices are needed to correlate en-tomological risk of dengue transmission in the sur-veillance and prevention of dengue. Among the al-ternative indices to assess risk of Aedes aegypti(L.), the Maya index (MI) and premise conditionindex (PC! have been recently evaluated. In these,the MI was less sensitive than the PCI to predicthouses with high risk of presence of larvae oradults of Ae. aegypti. In the present study, we val-idated the PCI in southern Mexico (Ciudad Hidal-go, Chiapas; 14%1'N, 92"09'W population 15,000)where dengue transmission is high, and tested amodification of the MI. The study was carried outfrom July to October 1996, collecting Ae. aegyptilarvae and adults between 0900 and 140O lrin7l7randomly selected houses. The indices assignedlevels of high, medium, and low risk of Ae. aegyptias follows. The PCI used the appearance of thehouse ("weariness" of roof and floor and openingson the wall), level of tidiness, and amount of shadeof the backyard. The MI categorized each house byits "hygiene," that is, the number of disposablecontainers, and its "breeding risk," that is, thenumber of controllable containers, dividing bothtypes of containers by the mean number of con-tainers of that type in the community. The weightedMI (MI*), was the same as the MI, except that bothcontainer types were divided by the mean numberof positive containers of that type in the commu-nity. For comparison, the Breteau index (BI) wascalculated.

The PCI probabilities for houses at medium orhigh risk of Ae. aegypri larvae were 4.36 (95Vo Cl: L.33-L4.29) and 15.19 (95Vo CI: 4.55-50.60)greater than houses with low risk. For adults theprobabilities for houses at medium or high risk andfor adults the probabilities for houses at medium orhigh risk were 1.69 (95Vo Cl = 0.85-3.3a) and 3.57(95% CI : I.75-:7.34), respectively. Although theMI could not associate risk of presence of mosquito

Spprerrangn Moseuno VBcron SYMPoSTM

larvae or adults, houses classified by the MI* atmedium and high risks were 2.35 (1.47-3.70) and2.33 (L.52-3.59) times at greater risk to have con-tainers withAe. aegypti larvae and for adults, hous-es classified at high risk were I.7l (1.17-2.50)greater risks than houses classified at low risk.Agreement for the PCI and MI* was slight (39Vo,K = 0.06, P < 0.05). The BI was 0.50. In conclu-sion, the validity of the PCI to predict the presenceof Ae. aegypri larvae or adults was demonstrated.In addition, the MI- provided a better approxima-tion for the risk of Ae. aegypri breeding in housesthan the unweighted MI.

A study of the entomological risk factors(intra- and peridomiciliary resting habits of

Aedes aegypti) for the transmission of dengue

E. Lugo Villalba, M. Delgado, S. Valle,P Espinoza, D. Lopez and M. Lopez

Centro Nacional de Diagn6stico y Referencia,Ministerio de Salud, Managua, Nicaragua

Although Aedes alboplcras (Skuse) has beendocumented to have infested Guatemala and El Sal-vador, the principal vector of dengue in CentralAmerica and Mexico is Aedes aegypti (L.). The ob-jectives of this study were 2-fold: to study the inta-and peridomiciliary resting behavior of Ae. aegyptiin the absence of insecticide application, and tocompare the age structure and preferred restingsites of this mosquito.

Our results demonstrated that the Ae. aegypti(male and female) intra- and peridomiciliary restingactivity peaked between 1400 and 1600 h in bed-rooms and wash basins/barrels (peridomiciliary),respectively. Similarly, proportions of gravid fe-males (intra- and peridomiciliary) generally in-creased to more than 5OVo during the same period.Kitchens and hallways were found to provide theleast favorable resting sites during the sampling pe-riod, 0800-1800 h. These results illustrate that bymore effectively targeting vector behavior, an al-ternative to traditional control stratesies can beachieved.

Control of Aedes aegyptiz an educational stratesrinvolving the participation of medical students

from tfre University of Panama, 1992-1ry7

E. Quiroz and A. Ying de Ttrrner

Ministerio de Salud an^d Universidad de Panamd.Panamd City, Panama

For 5 consecutive years, the Microbiology De-partment in the Faculty of Medicine at the Univer-sity of Panama introduced a dengue prevention pro-tocol as an educational strategy to incorporate stu-dents' participation to help in solving a publichealth problem: the reinfestation of Panama with

Aedes aegypti (L.). Since 1985, the mosquito,which transmits dengue, has reinfested Panama.This report documents the activities carried out byl0 groups of participating medical students (60O)to improve the consciousness of the university stu-dents in the prevention of dengue and to eliminateactive Ae. aegypti breeding sites. Various types ofwater-filled containers were found to be positive forthe larvae or pupae of Ae. aegypti, including plasticvases, bottles, cans, air conditioner pans, and irreg-ulars structures in buildings; all were easy to elim-inate. Various methods were used to inform thecommunity, mostly involving speeches, brochures,illustrative boards. and seminars. Satisfaction andincreased awareness were evident at the end ofeachsemester, when the students presented their rec-ommendations to other groups.

Eradication of Aedes aegypti from the Island ofTaboga with community participation: the

diagnostic phase

A. Ying de Turner, G. Davila de Obaldia andC. Campos

Universidad de Panamd, Panamd Canal Commis-sion, and Ministerio de Salud, Panamd City,

Panama

In 1995, the dengue surveillance program in Pan-ama reported 14 cases of dengue fever caused bydengue-3 on Taboga Island. This situation was ofparticular interest to the National CommitteeAgainst Dengue. The epidemiologic implicationscreated a basis for creating a program with com-munitary participation against Aedes aegypti (L.)on Taboga Island.

With the objective of elaborating the necessarystrategies to completely eliminate Ae. aegypti onthe island, we conducted a diagnostic survey of thesanitary situation. During this period, we completedentomological, community, and sanitary surveys.We interviewed teachers, civil authorities, and per-sonnel of the National Institute of Aqueduct andSewerage.

T\e 625 existing landed properties on the TabogaIsland were evaluated and we found that the prin-cipal mosquito breeding sites were in water storedfor domestic use and inside summer houses thatwere closed for more than 1 week. The entomolog-ic indices (house, Breteau, pupal, and the differentdensity of the larval stages) reflected that breedingin unserviceable sites was a secondary problem.

The results of the community survey showed thatthe principal problem that the residents had was anundependable water supply. The strategy to elimi-nate Ae. aegypti breeding involves use of water re-serve tanks with covers. The community mustagree to acquire, use, and maintain those tanks. Theowners of summer houses who have water tanksindoors must agree to use tank covers indoors.

JounNaL or rns AusmcAN Moseurro CoNrnol AssocntoN Vor-. 14, No. 3

These strategies were based on the results of thefield survey.

The eradication of Aedes aegypti..returning to the past or looking to the future

H. Gomez-Dantes

C entro de Inv e sti gacione s Enfermedades Infe c cios -as, Instituto Naciorwl Salud (INSP), Cuernavaca,

Morelos. Mexico

The Americas are now facing the emergence ofdengue and its severe form, dengue hemorrhagicfever (DHF), which is rapidly becoming a majorpublic health problem in the hemisphere. Aedes ae-gypti (L.), the principal vector of dengue virus inthe Americas, is widely distributed in urban, peri-urban, and even rural areas throughout the region.In the early 1960s, this species was partially elim-inated from the hemisphere and its eradication isnow proposed as the solution for the transmissionof dengue and the elimination of the risk of DHFin the Americas. The emergence of dengue andDHF has prompted an unusual response amongpublic health institutions and the eradication pro-posals are justified based on the partial success oferadication in the past and the assumption of itsfeasibility in the present. The conditions that madethe elimination of Ae. aegypti possible in the pasthave changed, and the tools available for the con-trol or eradication ofthe species have not improvedor increased. Social issues that modifled the naturalniche of the vector have increased the vector's den-sity and dispersal. Urbanization, air travel, humanmigration, production of nonrecyclable goods,cornmerce, and poor public services in growingmetropolitan areas are among the most importantdeterminants of the expansion of Ae. aegypfi'srange and are factors impeding our ability toachieve eradication. The control strategy for thisvector is based on elimination of production sitesand larvicidal application in expanding urban areas.Clean-up campaigns do not always target the mostproductive or stable breeding sites, and their orga-nization is expensive, labor intensive, and relies onmunicipal initiatives. Larvicides have partial cov-erage, short impact, and demand many human re-sources. Insecticide spraying is expensive and hasa limited impact. Eradication is conceived as a ver-tical strategy focused exclusively on the vector andit will certainly compete for the scarce financial andhuman resources in related public health programs.The issues of priority, vulnerability, political com-promise, and costs involved in such a task as wellas the problems and obstacles are discussed in de-tail. If there are reasons to engage in a major tasksuch as eradication, there are also temptations andexperiences of the past that should caution us toconsider the potential consequences of such a pro-gam.

Larvicidal impact of two formulations ofBacillus sphaericus (strain 23622 Yerlolex CG,

Spherimos liquid) on Anopheles albimanuslarvae in natural habitats in Nicaragua in 1996

P. Rivera Mendoza

Centro Nacional de Diagnostico y Referencia,Ministerio de Salud, Managua, Nicaragua

A study to evaluate the larvicidal impact of 2formulations of Bacillus sphaericus (strun 2362,Vectolex@ [h5a5B, CG] and Spherimos liquid) wascarried out in Nicaragua. Six Anopheles albimanusWiedemann breeding sites in an extensive malari-ous zone of Managua near the International Airportand located parallel to Xolotlan Lake were selectedfor study. TWo of these sites were treated withSpherimos; one received the minimum dose (0.3mVm'�) and the other the maximum dose (0.6 mUm'�). Spherimos was applied using a Hudson X-pertsprayer with 8002 nazzle. TWo were treated byhand with Vectolex; one received the minimumdose (2 g/m'�) and the other one received the max-imum dose (5 g/m). The remaining 2 sites wereused as untreated controls.

The results indicated that both Spherimos or Vec-tolex can reduce larval An. albimanus populationsfrom 95.3 to IOOVo at72h after application. Duringthe study no significant differences were observedon the efficacy of dosage and formulation used. Thephysical and chemical factors of breeding placesappe,u to have no influence on larvicidal activityof these formulations.

Field evaluation of the lambda-cyhalothrin 107aWP to replace DDT for malaria control

in Mexico

J. Thurion-Icaza, J. A. Vazquez-Narvaez,J. E. Mendez-Galvan and J. L. Santos-Preciado

Sub s e cre taria de P rev encion de Enfe rmedad.e s,Secretaria de Salud, DF, and Secretaria de Salud,

Michoacan. Morelia. Mexico

The North American Regional Plan includes dis-continuation of DDT use for malaria vector control.Mexican authorities have therefore started evalua-tion of alternatives to DDT such as the pyrethroidinsecticide lambda-cyhalotbrin (ICON IOVo wetta-ble powder [WP]) which has already been adoptedfor house-spraying against malaria vectors in over30 countries. In an area of persistent malaria trans-mission in coastal Michoacan State, we sprayed ap-proximately 5,00O houses with lambda-cyhalothrin(30 mg/m'�in 1997). When interviewed after spray-ing,97Vo of housewives were pleased that the treat-ment was effective against cockroaches, scorpions,and other domestic pests, and all house-holders ac-cepted the regular spraying of houses with ICON10 WP Susceptibility tests with lamMa-cyhalothrin(O.lVo dragnostic concentration) showed >957o

Mosqurro Vecron Syuposruv 227

mortality of Anopheles albimanus Wiedemann andAnopheles pseudopunctipennis Theobald. Amongnearly 14,000 residents of sprayed houses, the ma-laria incidence was reduced by 37Vo during 1997compared to the previous year when DDT wassprayed.

Detection of resistance mechanisms inAedes aegypti from Cuba and Venezuela:

standardization of the methods

M. M. Rodriguez, J. Bisset, L. Mil6, D. Molina,L. Lauzan, E. Calvo and M. Raymond

Laboratory of Toxicology and Genetics, Depart-ment of Vector Control, Institute of Tropical Med-icine "Pedro Kourf," Havana, Cuba; Escuela deMalariologia, Ministerio de Salud y Asistencia So-cial, Maracay, Venezuela; and Institut des Sciences

de l'Evolution, Montpelier, France

Five strains of Aedes aegypti (L.) larvae, onefrom Santiago de Cuba, Cuba, and 4 from the Ven-ezuelan states of Apure, Aragua, Miranda, andTachira, were bioassayed for sensitivity to the or-ganophosphorus insecticides (temephos, Malathion,methyl pyrimifos, and chlorpyrifos) and the pyre-throids (deltamethrin, lambda-cyhalothrin, and cy-permethrin). DEF and peronyl butoxide (PB) syn-ergists were used to assess the possible involvementof esterases or oxidases as resistance mechanisms.This was the first time that biochemical assays wereused to test for resistance in Ae. aegypti from Cuba.We standardized the techniques to detect elevatedesterases and glutathion-^S-transferase. The Vene-zuelan strains showed low levels of resistance tothe organophosphate insecticides temephos andMalathion. Conversely, the Cuban strain showedmoderate and high levels of resistance to temephosand Malathion, respectively. All of the strainsshowed low resistance levels to the pyrethroid in-secticides. Polyacrylamide gel electrophoresis testswere performed to determine what kind of esterasewas involved in the resistance. A highly activestained band named A4 was found in all the resis-tant strains, but not in the susceptible strains.

Influence of highly active esterase in pyrethroidresistance in Culex quinquefosciafzs from Cuba

J. A. Bisset, D. Torres-Monet, M. Rodriguez,Y. O. Nffiez, E. Calvo, C. Diaz and A. Soca

Inboratory of Toxicology and Genetics, Depart-ment of Vector Control, Institute of Tropical Med-

icine "Pedro Kourt," Havana, Cuba

A strain (SP6) of Culex quinquefosciatus Saycollected from Santiago de Cuba, Cuba, was se-lected during 6 and 4 generations with lambda-cy-halothrin and deltamethrin. Different levels of re-sistance to those insecticides were observed whencompared to the susceptible reference strain (S-

Lab). Cross-resistance to Malathion was also de-tected. Lethal concentrations inducing 5OVo mortal-ity decreased when bioassays were conducted in thepresence of the synergist DEE an inhibitor of es-terase enzymes. This represents a decrease in theresistance factor of more than 907o, indicating thatelevated esterases are playing an important role inresistance to this pyrethroid. All SP6 mosquitoesshowed an identical esterase pattern after poly-acrylamide gel electrophoresis (B1, A6, 86). Cross-ing and back crossing experiments between SP6and S-Lab strains showed that B1 and B6 esterasesare coded by different genes, and that ,4'6 and 86esterases are linked, located just 6.7 recombinationunits apart. Resistance to pyrethroid was inheritedin a multifactorial and incomplete dominant mode,due mainly to the influence of B1 and A686, andprobably the action of other resistance mechanisms.lnhibition studies using the purified esterasesshowed this potential capacity to react with pyreth-roids, especially esterase ,4,6.

Population genetic structure of Aedes aegypbthe vector of dengue in Mexico, using the

RAPD.PCR

N. Gorrochotegui-Escalante, F Garcia-Franco,I. Fernandez-Salas and M. L. Munoz

Centro Investigaciones y Estudios Avanzados, In-stituto Pohtecnico Nacional, Mexico, DF, and Ia-boratorio Entomologia Medica, Universidad Au-tononta de Nuevo Le6n, San Nicolas de los Garza,

Nuevo Le6n, Mexico

In 1997, Mexico experienced nearly 50,0O0 casesof classical dengue and about 1,000 cases of den-gue hemorrhagic fever. This reinforced the impor-tance of understanding the role of virus serotlpesand mosquito vector population genetics, as a mo-lecular approach for explaining the changing epi-demiology of this emerging vector-borne disease.Vector control efforts and other environmental fac-tors have produced a reduction in the genetic vari-abihty in Aedes aegtpti (L.). Analysis of populationgenetics will provide information about the breed-ing structure of Ae. aegypri in Mexico. This willhelp us understand the relationship between thevector and dengue transmission, which will be in-valuable in vector competence studies. The purposeof this study was to define the local breeding struc-ture of Ae. aegypti in geographically separated ar-eas of Mexico using rapid analysis of polymorphicDNA-polymerase chain reaction (RAPD-PCR)methodology. Mosquito larvae were collected indengue endemic areas in 6 states, mainly from thePacific coast areas of Oaxaca, Chiapas, and Guer-rero, and adjacent to the Gulf of Mexico in Vera-cruz, Nuevo Leon, and Tamaulipas. Larvae werecollected from peridomestic containers (55-gallondrums, discarded tires, 5-gallon buckets, and oth-ers) in backyards at 3 or 4 cardinal points in each

JounNlr oF TrG ArlGRrclN Mosquno CoNTRoL Assocrenon Vor. 14. No. 3

city. Mosquitoes were reared to the adult stage andfrozen at -7O"C for DNA analysis. We tested 35primers for RAPD-PCR and selected 7 of them. Wefound 55 polymorphic loci in 15 individuals usingthe analysis in agarose gels. Our preliminary resultsshowed an Nm : O.019 + 0.019 (F.r) estimatedaccording to Lynch and Milligan, which was small-er than the Nm : 0.030 + 0.012 019 (F.r) esti-mated according to Wright for the northern andsouthern areas of Monterrey in Nuevo Leon. Theseresults suggest that a larger sample size is neededto estimate gene flow more accurately; therefore,we will analyze this sample using acrylamide gels,which will give a higher number of polymorphicmarkers.

Variability in mitochondrial DNA in populationsof Anophelcs albimanus in Guatemala

M. Palmieri, A. Molina, M. M. Yurrita. E. Molina.W C. Black IV and A. M. P Merida

Medical Entomology Research and Training Unit(MERTU-G), Guatemala City, Guatemala, and

Colorado State University, Ft. Collins, CO

Anopheles albimanus Wiedemann is consideredthe principal vector of malaria in Central America,parts of South America, and the Caribbean. Thespecies shows great variability in characteristics as-sociated with its vectorial capacity. The species isconsidered a poor vector of malaria because it ismore zoophilic than anthropophilic, more exophag-ic than endophagic, and more exophilic than en-dophilic in its behavior with humans. Morphologicvariants have been reported in all stages and theirfeeding behavior varies with season and geograph-ical location. Differences in their biting hours aremore related to the human activity that surroundsthem than to their natural circadian rhythms. It isnot known if their role as vectors is determined bythe high densities of mosquitoes or the existence ofa species complex, with some species having ahigher potential to transmit malaria. Little is knownabout the molecular genetics, population dynamics,and phylogeny of An. albimanus. Finding geneticmarkers that help identify malaria vectors is im-portant. Proper identification combined with epi-demiologic studies and transmission studies mayhave an impact on malaria control efforts.

There is great interest in developed countries indeveloping transgenic mosquitoes. To undertakethis type of manipulation, it is necessary to knowthe basic genetic structure and population geneticsof these mosquitoes. Guatemala has developed theinfrastructure and expertise to undertake these typesof studies.

Polymorphisms in mitochondrial DNA (mtDNA)are being increasingly exploited for the study ofgenetic relationships among closely related organ-isms. The simple and uniform organization, lack ofrecombination, maternal inheritance, and relatively

rapid rate of change make mtDNA a valuable toolfor the study of molecular evolution and phyloge-netic relationships. Numbers of maternal clones areindicative of founding population size and thus pro-vide evidence of the existence of severe populationbottlenecks. Maternal lineages can be traced andthe phylogenies of existing populations derived in-dependently of founder effects or adaptation. Thisevidence cannot be inferred from studies of nuclearmarkers that are biparentally inherited and recom-bine. Our ultimate goal in the mitochondrial DNAproject is to trace the historical establishment of An.albimanus throughout Latin America.

We studied the mitochondrial DNA in the ND5gene of An. albimanus from 3 malarious regions ofGuatemala. Mitochondrial DNA fragments wereamplified with primers designed from other insectsand examined for variability with a technique calledSSCP (Single Stranded Conformation Polymor-phisms). A total of 1,236 An. albimanu,s was ana-lyzed and 34 different ND5 haplotypes were iden-tified from the entire country. The southern regionof Guatemala presented 26 different haplotypes, thenorthern region presented24, and the eastern regionhad 18 different haplotypes. The Shannon diversityanalysis depicts the northern region of the countrywith the highest diversity (lI : 3.478), followed bythe southern (H : 2.9364) and eastern regions (I1: 2.5171). Analysis also indicates that the diversityof haplotypes among individuals within collectionsaccounted for most of the diversity in each region(89Vo for the eastern, 82Vo for the northern. and'l2Vo

for the southern region). The diversity amongcollections within a site contributed to the diversityof the region only by 8.6Vo rn the northern region,4.5Vo in the southern region, and O.3Vo in the east-ern region. These results suggest that collectionswithin a region were very uniform in diversity. Di-versity between sites accounted for L9Vo of the totaldiversity in the southern region, IlVo rn the easternregion, and.9OVo in the northern region. This sug-gests that more genetic variation occurred betweensites in the soutlern region compared to the otherregions of Guatemala.

Distribution of Anopheles aquasalis in theCaribbean region: vicariance or dispersal

y. Rangel

Instituto hologia Tropical, Universidad Central deVenezuela, Caracas, Venezuela

Arwpheles aquasalis Curry occurs from Brazil toNicaragua, Ecuador, Trinidad and Tobago, and theLesser Antilles. Studies of genetic structure carriedout in Venezuela, Trinidad, and Barbados showedthat mean heterozygosity per locus (If, number ofalleles per locus, and Wright's F statistics did notexhibit significant variation. However, the propor-tion of polymorphic loci (P) was significantly lower

SePTErusen 1998 Mosquno Vscron SYMPoSTM 229

for populations from Barbados (Il.l5%o) than forthose from Venezuela (34.42Vo) and Trinidad(32.5Vo). This finding and an absence ofrare allelescould be an indication that a major reduction in thegene pool occurred in the evolutionary history ofthe population from Barbados. This report presentsa hypothesis of historical reconstruction to explainhow the present patterns were formed. The MiddleAmerica-Caribbean region is extremely complexbut models of Caribbean evolution have attemptedto reconcile existing geological data with known orinferred tectonic plate motions. This model is a pre-liminary framework and it assumes a Pacific prov-enance for the Caribbean Plate. that the Lesser An-tilles were more closely tied to South America dur-ing their development from the Late Eocene to thepresent time, and Pleistocene climates and changesin sea level exerted a powerful influence on zoo-geographic patterns of distribution. The currentpopulation in Trinidad is suggested to be a frag-mented biota, or vicariated from Venezuela; where-as the population in Barbados required dispersalevents across intermediate islands by a steppingstone model.

Mosquitephytotelmata biogeography inVenezuela using parsimony analysis

of endemicity

J. C. Navaro. J. Luria and R. Barrera

Instituto hologia Tropical, Universidad Central deVe ne zue Ia, C arac as, Ve ne zue l.a

We studied the association between immaturemosquitoes and aquatic habitats in plants or part ofplants (phytotelmata) from 17 localities in nationalparks and natural monuments in Venezuela. A totalof 60 mosquito species was collected in a 3-yearperiod from 47 host-plant species. With 2 excep-tions, all other mosquito species collected were re-stricted to phytotelmata. Different degrees of mos-quito-plant associations were observed, with thehighest specificity existing at the genus and sub-genus levels. We ran a parsimony analysis of en-demicity (Wagners method; Henning86 andNONA) using a 17 (localities as taxa) X 60 (mos-quito species as characters) binary matrix. The finalstrict consensus cladogram (from 28 trees) showed2 main, well-supported groups named Guayanes(Guayana Shield) and Andino (Los Andes), and an-other group not as well supported called Cordiller-ano (north-central Cordillera). Alternatively, usinga semistrict consensus cladograrn we found 7groups, which can be split into 2 main groups:Guayanes and Cordillerano. The first group had 3monophyletic subgroups (Amazonico, Gran-saban-ico, and Guayanes), whereas the 2nd group had 4subgroups (Andino, Perijano, Caribeo, and Centro-oriental). These results support the hypothesis that

the Guayana Shield was the ancestral center of spe-ciation, followed by the Andean Region. The faunafrom the Central Cordillera is derived from the lat-ter and is evolutionarily more recent. This studyrepresents a first attempt to understand the curentdistribution and evolutionary history of this insectgroup in Venezuela and adjacent areas.

Characterizing anopheline embryogenesis:clarification and permeabilization of

initial assays

A. T, Monnerat. J. B. Pereira-Lima. M. J. Soares.M. Pelajo-Machado, B. S. Vale, H. L. Lenzi, M. G.

Freitas-Sibajev, R. Galler and D. Valle

Departamento Biologfa y Biologica Molecular, De-partamento Ultraestructura y Biologica Celular,Departamento Patologia, Instituto Oswaldo CruzFIOCRUZ, and U.S. Army Medical Research Unit,

Rio de Janeiro, Brazil

The construction of transgenic mosquitoes thatare refractory to a Plasmodium infection is an al-ternative to the control of malaria that has been thesubject of a great deal of effort. In this sense theknowledge of the vector's embryonic developmentis extremely important, although it has been sur-prisingly neglected. Tlansgenic mosquitoes are ex-pected to come from the injection of exogenousDNA into embryos and the consequent integrationof this material into polar cells. Hence, detailedknowledge of early embryogenesis, including timeand position of polar cell formation, is essential.Knowledge of wild mosquito embryogenesis as awhole will also be important to the characterizationof mutants that are currently being isolated and oftransgenic lines to be generated.

The recent availability of Neotropical anophelinecolonies has made the study of embryogenesis pos-sible: however. the melanized and sclerotized na-ture of the eggs of these mosquitoes still poses aproblem. Thus, different methods have been usedwith the goal of clarifying and permeabilizing an-opheline eggs.

The clarification method chosen is enabling thedefinition of the major morphogenetic movementsof this long germ band embryo, as well as the es-tablishment of a parallel with Drosophila melano-gaster. These analyses have been made with con-focal microscopy, because clarified anopheline eggsproved to be autofluorescent. Among differentmethods that are being tested for the permeabili-zation of anopheline eggs, the one classically usedfor tlre dechorionation and devitellinization of Dro-sophila eggs was not effective when applied to an-ophelines, as verified by scanning electron micros-copy.

This research was supported by Institute Oswal-do Cruz, CNPq, European Community.

230 Jourulel or ntg AvEttclrv Mosqurro CoNrnor_ Assocnrrorl Vo l . 14 , No.3

Population dynamics of Lutzomyia evansi andLutzomyia longipalpis (Diptera: Psychodidae) in

an ancient endemic focus of visceralleishmaniasis in Venezuela

M. D. Feliciangeli

Universidad Carabobo, Maracay, Aragua,Venezuela

Knowledge of the spatial distribution and behav-ior of sandfly vectors contributes to the understand-ing of the epidemiology of leishmaniasis. Infor-mation on the abundance and temporal distributionof vectors permits public health offlcials to predicttransmission seasonality and assists in planningcontrol strategies. Additionally, through the inte-gration of entomological data and epidemiologicaldata on humans and reservoirs from different de-fined ecotnres, an approach to the stratification ofthe leishmaniases may be attempted. Lutzomyiaevansl (Nunez-Tovar) allid Lutzomyia longipalpis(Lutz and Neiva) recently have been found to beinfected with Leishmania (Leishmania) spp. inGuayabita, Venezuela (20'10"N, 28'67'),1), an an-cient endemic focus of visceral leishmaniasis eco-logically situated in a premontane dry forest. In thislocality, I2.4Vo of humans were leishmanin positiveand I5.5Vo of the dogs were seropositive. Monthlycollections of phlebotomine sandflies were carriedout using a fluorescent light-baited Shannon trap(January 1993-June 1994) and Centers for DiseaseControl (CDC) miniature light traps (May 1993-June 1994). A total 3,230 males and 6,091 femalesfrom 1l phlebotomine sandfly species was collect-ed. Lutzornyia evansi was the dominant species(86.4Vo), l0-fold more abundant than Z. Iongipalpis(lO.67Vo). The 2 vectors markedly alternated sea-sonally: L. evansi peaked during the rainy season(May-November) and Z. Iongipalpis peaked duringthe dry season (December-April). Plots of theabundance of both species and abiotic environmen-tal factors (temperature, relative humidity, and rain-fall) were correlated to determine the weight ofeach of these covariates. The results are comparedwith observations made elsewhere.

This work was supported by FUNDACITE Ar-agua (Pasa 17), The Commission of EuropeanCommunity (S3-CT93-O247), and the University ofCarabobo.

Response of Leishmaniasis vectors to variouspesticides in Venezuela

M. Mazzari, M. D. Feliciangeli and M. Maroli

Division de Control de Vectores, Ministerio de San-idad y Asistencia Social and Universidad Carabo-bo, Maracay, Aragua, Venezuela, and LaboratorioParassitologia, Instituto Superiore di Sanitaire,

Roma, Italy

Cutaneous leishmaniasis (CL) and visceral leish-maniasis (VL) are endemic in Venezuela, where

Lutzomyia ovallesi (Ortiz) and Lutzomyia longipal-pis (Lutz and Neiva), respectively, are recognizedas vectors of these diseases in the countrv. Becauseboth species enter houses, a control program base.don the use of insecticides, indoor spraying, or im-pregnated curtains might be applicable. To deter-mine the susceptibility to different insecticides, alaboratory (S) and field strain of L longipalpis anda field strain of L. ovallesi were tested for theirsusceptibility to organochlorine (DDT 2Vo), carba-mate (propoxrx O.OLVo), organophosphate (Malathi-on 27o, fenitrothion l7o, arlrd methyl pirimiphosl%o), and pyrethroid (deltamethrin O.O6Vo, lambda-cyhalothrin O.06Vo, and permethrin O.2Vo) insecti-cides. Susceptibility to the insecticides tested wasevaluated in the field population and compared witha reference strain ofL. longipalpis. The 957o lethaltime (LT95) to propoxur and Malathion insecticidesfor the field population was lower than the LT95for the laboratory reference strain, showing highsusceptibility to these compounds. A low level ofresistance at the median lethal time (LT50) (<3-fold) was found for the fenitrothion, pirimiphosmethyl, and permethrin insecticides, but no resis-tance was detected at LI95. No significant resis-tance at LT50 and LI95 was detected for the py-rethroid insecticides. A preliminary field trial usinglambda-cyhalothrin (25 mg active ingredient [AI]/m2) sprayed in interior walls produced a reductionin the number of indoor sandflies for 2 months du-ration. Simultaneously, biological assays to test theresidual effect ofthis insecticide on wood, clay, andblock walls were made. Five, 69, and 131 days afterthe spraying, 3-6 batches of 20 laboratory-rearedfemale sand flies were exposed for 30 min tosprayed walls. An average of 99, 39, and 5Vo mor-tality was obtained, respectively. The highest mor-tality was observed among sand flies exposed tosprayed wooden walls. Further trials are phmned todesign an improved strategy of control.

Intra- and interspecific competition betweenAedes aegypti L. and Culex pipians L. (Diptera:

Culicidae) in the laboratory

J. L. Villanueva, H. Quiroz, V. A. Rodriguez,M. H. Badii, K. M. Valdez, E Gonzalez and

J. K. Olson

Laboratorio Entomologia, Universidad Aut6nomade Nuevo Leon, Nuevo Le6n, Mexico, and Depart-

ment of Entomology, Texas A&M University,College Station, TX

One component of population dynamics is com-petition. It is a very important factor in regulationof natural populations but often is forgotten in thecontrol of mosquito larvae. Competition has beenevaluated by studying larval mortality, growth, bio-mass, and fecundity. The objective of this studywas to determine intra- and interspecific competi-tion between Aedes aegypri (L.) and Culex pipiens

SppreMsen 1998 Moseutro Vecron SvrvIPosruv 233

generational time, intrinsic rate of increase, and re-productive value. Survivorship tables were pre-pared and the mortality increments were describedfor each instar. Total egg loss was 9l.2Vo at roomtemperature and 67.lVo in the incubator, due todeath of females, retained eggs, sterile eggs, andembryonic death. This analysis provides useful pa-rameters for handling colonies of these insects inthe laboratory and determining causes of mortality.

Operational experience on the use of twoformulations of Bacillus sphaericus

(strain 2632') in different types of breeding sitesof Anopheles albilnanus and

Culcx quinquefasciatus in Honduras

L. Rivera, C. Sherman, M. Ortiz and C. Tabora

Ministry of Health, Honduras

During April-August 1997, 2 formulations ofBacillus sphaericus, strain 2632, (Griselesf; at arate of 1O mVm'� were tested for the field control ofAnopheles albimanus Wiedemann and Culex quin-quefasciatus Say in the area of El Progreso, Hon-duras. Both species were susceptible to Bacillussphaericus. Data were analyzed to calculate thepercent reduction of larval indicators. Environmen-tal conditions did not permit monitoring of the Cu-/ex breeding sites longer than 10 days. Anopheles

breeding sites were monitored for up to 90 days.Breeding sites included fish farms, cattle drinkingtroughs, and drums. Both formulations achievedsatisfactory control of An. albimanus for up to 90days. Noticeable differences were found in lengthof control between the 2 formulations and betweendifferent types of breeding sites.

REFERENCES CITED

Clark, G. G. (organizer). 1995. Mosquito vector controland biology in Latin America-a fifth symposium. J.Am. Mosq. Control Assoc. ll:343-353.

Clark, G. G. (organizer). 1996. Mosquito vector controland biology in Latin America-a sixth symposium. J.Am. Mosq. Control Assoc. l2:460-471.

Clark, G. G. and Y. N. Rangel (organizers). 1997. Mos-quito vector control and biology in Latin America-aseventh symposium. J. Am. Mosq. Control Assoc. 13:t t3 - t26 .

Clark, G. G. and M. E Suarez (organizers). 1991. Mos-quito vector control and biology in Latin America-asymposium. J. Am. Mosq. Control Assoc. 7:633-645.

Clark, G. G. and M. F Suarez (organizers). 1992. Mos-quito vector control and biology in Latin Arnerica-asecond symposium. J. Am. Mosq. Control Assoc. 8:305-317.

Clark, G. G. and M. F Suarez (organizers). 1993. Mos-quito vector control and biology in Latin America-athird symposium. J. Am. Mosq. Control Assoc.9:441-453.