Joumal of the American Control Association, lg(3):l7g_1g5. 2002 Copyright A 20f.2 by the American Mosquito Control Association, Inc.

ENVIRONMENTAL FACTORS ASSOCIATED WITH LARVAL HABITATS OF VECTORS IN NORTHERN KYUNGGI PROVINCE. REPUBLIC OF KOREAI

DAVID M. CLABORN,',3PAUL B. HSHIEH,' DONALD R. ROBERTS,'TERRY A BRIAN C. ZEICHNER5 eNI RICHARD G. ANDRE,

ABSTRACT, The larval habitats of malaria vectors near the DemilitarizedZone of the Republic of Korea (ROK) were sampled from June through September 20O0 to determine larval abundance and to identify envi- ronmental factors associated with high larval density. Six primary habitats were identified: rice fields, irrigation ditches, drainage ditches, stream pools, irrigation pools, and marshes. Most habitats harbored similar densities of larvae until August and September, when population densities in rice fields declined and those in irrigation pools increased. The primary vector in the ROK, sinensis, occurred in water with a wide range of values for environmental factors, including pH, total dissolved solids, percent of surface covered with floiting vegetation, and nitrate and phosphate concentrations. No environmental factor or combination of factors were found that were predictive of high larval densities. This study suggests that larval Anopheles are capable of developing in a wide range of stagnant, freshwater habitats in northern Kyunggi Province, ROK.

KEY WORDS , Anopheles lesteri, Anopheles yatsushiroen.si.r, habitat, Republic of Korea, water quaf ity. vivax

INTRODUCTION One of the best-documented instances of local transmission in the USA occurred after the visit of After an absence of more than l0 years, malaria an American soldier to a Girl Scout camp in Cali- () has reemerged in the Republic fornia where vector Anopheles were abundant (Bru- of Korea (ROK) (Feighner et al. 1998). The number netti et al. 1954). of detected cases grew from I in 1993 to 3,719 in Until recently, malaria prevention policies for 1999. Of all cases diagnosed in the ROK through American soldiers in high-risk areas of the ROK 1999,83 (2.23Vo) cases occurred in American mil- relied exclusively on vector control and personal itary personnel or Korean augmentees to the U.S. protective measures (PPM). However, Strickman et Army. As of December 2OOO,16 confirmed cases al. (2001) reported acceptable vector and disease had occurred in American personnel stationed in control in areas treated with residual insecticides the ROK for that year alone (Preventive Medicine and command emphasis on personal protection. Directorate, l8th MEDCOM, personal communi- The U.S. malaria policy was modified in 1999 by cation). To date, the focus of the disease has been placing more than 6,000 American soldiers sta- in Paju County just south of the Korean Demilita- tioned north of the Imjim River on chloroquine/ ized Zone (DIN{IZ) that separates North and South primaquine chemoprophylaxis. Although these Korea. Large populations of military personnel, drugs are still effective against the strain of P. vivax both American and Korean, are stationed in this in the ROK, some concerns exist about relying on area. this method as the primary means of malaria con- The strain of P. vivax transmitted in the ROK is trol. For instance, soldiers stationed in areas south well adapted to a temperate climate and demon- of the Imjim River often train lbr several days in strates both short and long incubation periods. Long camps considered to be high-risk areas north of the incubation periods of greater than 6 months in- river. During 1999, soldiers training in high-risk ar- crease the risk of returning soldiers reintroducing eas were placed on chemoprophylaxis before the malaria to the continental United States, making exercise until the end of the malaria season in Oc- malaria control in Korea even more important tober. During the 2000 malaria season, only soldiers (Walter Reed Army Institute of Research 1999). residing north of the Imjim River were placed on chemoprophylaxis. Soldiers entering these areas I This manuscript reports original research and does not only for training depended solely on the use of necessarily reflect the policy of the Department of Defense PPM, including permethrin-treated uniforms, prop- or the U.S. Navy. er weerring of the uniform (pants tucked into the 'Uniformed Services University of Health Sciences, boots and shirt sleeves rolled down), and topical 4301 Jones Bridge, Bethesda, MD 20814-4799. repellents. 3 Present address: Navy Disease Vector Ecology and In addition to PPM, an area-based control meth- Control Center, Jacksonville, FL 32212-0043. od might provide better malaria conffol in those a Preventive Services Directorate, 1Sth Medical Com- areas with transient military populations. Strickman mand, Unit 15281, APO AP 96205-0054. 5 U.S. Army Center for Health Promotion and Preven- et al. (1999) suggested that Iarviciding in mosquito tive Medicine, Entomological Sciences Program, APG, habitats around American installations could be a MD 21010-5403. useful control method if such efforts were coordi-

178 SBprslaesR.2002 MAI-Lnra Vscrons IN KYUNGGI PnovNce, KonEe t79 nated with Korean civilian authorities. One advan- the presence or abundance of vector populations tage of larviciding would be that protection could based on environmental factors by using logistic be conferred on all soldiers in the high-risk areas, regression (Savage et al. 1990), multivariate re- regardless of their length of exposure. However, the gression (Rodriguez et al. 1996, Grillet 200O, Mon- cost of such a larviciding program is unknown and cayo et al. 2000), and global information system would require significant information on the bio- techniques (Roberts and Rodriguez 1994, Roberts nomics of vector larvae in the malaria endemic ar- et al. 1999). Models of this type have the potential eas. In addition to the coordination of a larviciding to improve the efficiency of malaria vector control program with the surrounding Korean community, and surveillance programs in the ROK, but few if cost would be a major consideration. Cost compar- any have been developed for the indigenous malar- isons with other control methods, especially che- ia vectors. moprophylaxis, would be essential. The cost of larviciding is a function of the size Malaria vectors in the Republic of Korea of the larval habitats and the cost of treatment per unit area. However, the following 3 questions about In a review of Korea's of public the local vectors must be answered before the size health importance, Chow (1973) mentioned only 2 and location of larval habitats requiring treatments potential malaria vectors: Anopheles sinensis (Wie- can be determined. What larval habitats occur in demann) and Ano phe le s yat sus hi ro ensl.s Miyazaki. the high-risk area? What species of mosquito larvae Of these 2 species, Chow considered the fomer to occur in the local habitats? Which mosquito species be the most important vector. This conclusion gen- are proven malaria vectors? This paper describes erally has been considered correct by subsequent efforts to answer these questions and to develop a authors, but questions about the of An. quick and reliable means of surveying habitats in sinensis and others in the Hyrcanus Group have Korean malaria foci. caused significant confusion. In particular, the dif- Successful larval control requires the ability to ficulty of distinguishing An. sinensis from Anoph- identify larval habitats and to distinguish between eles lesteri Baisas and Hu is of some importance sites with high and low vector populations in a because the vector potential of the latter species is timely manner (Wood et al. 1991). Limited mos- unknown. Tanaka and others (1979) considered An. quito control assets in the ROK require prioritiza- lesteri, rather than An. sinensis, to be the probable tion of the areas in need of pesticide applications; primary vector of malaria in Japan, but little work this can be achieved with larval surveillance. One has been performed on this species on the Korean approach to surveillance is to identify key environ- peninsula. mental factors that predict the presence of vector In a survey of malaria vectors in Kyonbuk, Ko- populations, then to use these factors as markers to rea (Joo and Kang 1992), An. sinensis was the only predict the presence of significant larval densities species considered, even though it generally is con- either in an indirect survey or in a remote sensing sidered to be strongly zoophilic. Ree and others system. Many such environmental factors have (1973) reported only 3 Anopheles species caught in been identified for a variety of vectors. For exam- light traps (An. sinensis, Anopheles sineroides Ya- ple, throughout much of its extensive range, Anrsph- mada, and An. yatsushiroensis). Anopheles sinensis eles pseudopunctipennis Theobald was associated comprised 957o of the anophelines and l87o of the with green filamentous algae and aquatic vegetation total mosquito population. Strickman and others in sunlit freshwater stream pools (Manguin et al. (1999) caught the same 3 species in light traps in 1996b). In southern Mexico, high populations of the northern ROK, but also caught I specimen Wiedemann were linked to identified as An. lesteri. In that study, larval sur- flooded, unmanaged pastures at an elevation below veillance indicated densities of 0.02 to 3.1 larvae/ 25 m (Rodriguez et al. 1996). Positive associations dip in rice fields surrounding a military training between the density of An. pseudopunctipennis and area with a probable history of malaria transmis- the presence of filamentous algae or the plarfi Het- sion. A later collection (Strickman et al. 2OOl) eranthera spp. occurred during the dry season caught many more adult An. lesteri and An. yatsu- (Savage et al. 1990). In , Anopheles shiroensis, none of which were positive by enzyme- aquasalis Curry was collected most often in brack- linked immunosorbent assay for P. vivax circum- ish mangrove swamps and larval populations varied sporozoite antigen. In fact, only 2 out of 2,376 An. with the water salinity, whereas Anopheles oswal- sinensis tested for malaria infection were positive. doi Peryassu was associated with permanent fresh- Therefore, although An. sinensis is not considered water habitats (Grillet 2000). In Belize, the habitats to be an efficient vector, its relative abundance and of Root were characterized as the detection of P. vivax antigen only in this species being river margins with patches of floating debris implicate this mosquito as the primary vector in the (Manguin et al. 1996a). Vector populations also ROK. have been associated with pH and percent tree cov- This study investigated the distribution of larvae er (Rejmankova et al. 1998). of Anopheles among various larval habitats and at- Researchers have developed models to predict tempted to identify environmental factors that could 180 JounNlr- op ttrp AvpnrcnN Mosqurro CoNrnol AssocrlrroN Vor-. 18, No. 3

Table 1. Mosquito species taken from larval habitats in a rural environment. A high bluff overlooking the northern Kyunggi Province, Republic of Korea, and Imjim River bordered the southern perimeter of CP reared to adults (June-September 200O). Greaves. Rice farming was extensive on both sides Species of the river and continued from the camp into the (no. specimens) DMZ. Cattle and goat herds were located near the reared to Habitats where species eastem extension of CP Greaves. adult stage was found The area around CP Greaves was characterized Anopheles sinensis (442) Rice field, stream pool, drain- by low, wooded hills separated by shallow valleys. age ditch, irrigation ditch, The valleys usually were planted in rice, but also marsh supported crops of peppers, corn. onion, ginseng, An. Iesteri (42) Rice field. stream pool. irriga- and cabbage. Camp Casey was surrounded by tion ditch steeper, rockier hills that may have exceeded 300 An. yatsushiroensis (3) Irrigation pond, drainage ditch m in height. Other than the I large stream tran- Aedes vexans (277) Rice field. stream pool, drain- secting CP Casey, few natural streams occurred in age ditch, irrigation ditch, the study areas. Numerous irrigation ditches and irrigation pond, marsh ponds were interspersed between paddies, vagans (41) Rice field. stream pool. drain- the rice age ditch providing a variety of potential larval habitats. Cx. pipiens (79) Rice field, drainage ditch, irri- Six habitats were identified for sampling purpos- gation ditch, irrigation es: rice fields, irrigation ditches, drainage ditches, pond, marsh marshes, stream pools, and irrigation pools. The Cx. orientalis (36) Rice field. stream pool. drain- rice fields were typically small (less than 1,300 m') age ditch and were bordered by dikes ranging in height from Cx. tritaeniorhynchus Rice field, drainage ditch, irri- 0.3 to 2.0 m. Early in the season, the dikes were (30) gation ditch typically mowed and devoid of tall vegetation; Cx. mimeticus (7) Rice field, stream pool however, the degree to which the were Cx. hayashi (3) Irrigation pond borders Cx. bitaeniorhynchus Irrigation ditch, stream pool maintained varied, and some became overgrown (10) later in the season. The rice fields were transplanted Cx. rubensis (l) Marsh with seedlings in May through early June. Irrigation or flooding of the rice fields was inconsistent so that some were flooded whereas others at the same stage of plant development were not. Irrigation be used to efficiently locate the sites producing the ditches ranged in size from small troughs 0.3 m greatest number of malaria vectors. deep to large canals up to 2 m wide and 1 m deep. Drainage ditches also varied in size and were fre- quently lined with concrete or stone. Stream pools MATERIALS AND METHODS were observed only at CP Casey and were all up- The study was performed from June through stream of a wastewater treatment plant. Pools September 2000 in the northwestern part of Kyung- ranged in depth from a few centimeters up to I m, gi Province, ROK, about 60 km north of Seoul, and and in surface area from less than 1 m2 to about near the DMZ in the vicinity of Munsan, Paju 100 m'. They often supported heavy algal mats and County. This area was characterized by a temperate had sand or mud bottoms; large stones or concrete climate with 4 distinct seasons. Average tempera- sides surrounded most ofthe pools. Irrigation ponds tures ranged from -7 to 2'C in the winter and up were located nea.r the rice fields and provided water to 2O-29"C in the summer. Monsoons usually affect for irrigation. The ponds were usually about 1.5 m the area, with the heaviest average rainfall by sea- deep (range: 800-2,100 mm) with steep sides. Ear- son occurring in the summer months (mean : 258 ly in the season, few larvae were found in these mm./month). The summer of 2000 was unusually sites, but by August, large populations of larvae dry; however, typhoon-associated rainfall was sig- occurred among the algae growing along the edge. nificant and provided enough precipitation to flood Marshes were not common so only 2 were sampled many of the study sites, especially the streamside during this study. The marshes supported a wide pools. Sample sites were concentrated in the areas range of vegetation and were always flooded. The surrounding the Camp (CP) Casey complex and CP 2 marshes were included only for a general survey Greaves (both U.S. military bases). The CP Casey of the species present and were not included in sta- complex consisted of 3 military facilities that in- tistical analyses. cluded the contiguous CPs Castle and Hovey. A Data collection: Initlally,60 study sites were se- large, permanent stream that was prone to flooding lected and located with a Garmin III@ handheld transected the complex. The towns of Tongduchon global positioning satellite (GPS) unit (Garmin In- and Tokori surrounded the CP Casey complex on 3 ternational Inc., Olathe, KS). The GPS positions sides, but rice cultivation continued at the interfaces were collected for entry into a geographic infor- between the camps and villages. mation systems database to determine the size of The CP Greaves area was smaller and located in larval habitats in high-risk areas. Thirty sites were SepreNassn2002 M.e.La.nra.Vecrons tN KvuNccI PnovrNcp, Konpe 181

Table 2. Median number of larvae per dip (n, standard deviation, and range are given in parentheses) of Anopheles spp. collected in larval habitats of northern Kyunggi Province, Republic of Korea (June-September 20O0).r

June July August September

Rice paddies 0.1a O.4a 0-3a 0.0a (42,O.9, O-s.3) (42, O.9,O-4.1) (49,0.4, 0-1.9) (41,0.6, o-3.0) Ditches 0.1a O.4a 0.1a 0.1a (10,0.3,0{.9) (ll, 1.8,0-6.0) (17,0.3, 0-O.9) (14, r.4, O-4.7) Pools and ponds 0.0b O.7a O.4a 5.0b (8,0.0, 0-0.1) (12,4.3,0-14.7) (14, 1.4,0-3.0) (11,13.1,0-39:3)

'Median values within a column followed by the same letter are not significantly different (Kruskal-Wallis; P > 0.05). Irigation ditches and drainage ditches were combined into I category for purposes of statistical analysis. Stream pools and inigation ponds also were combined. located around each of the 2 camps, Greaves and (Lemna spp.) or algae was estimated visually. Three Casey. As the season progressed, other sites be- observers made independent estimates of the duck- came available and were added to the study for a weed or algal cover and the mean of the 3 estimates total of 93 sites: 50 rice paddies, 13 stream pools, was recorded. 12 irrigation ditches, ll drainage ditches,5 irriga- Rearing procedure.s: Because the species of Ko- tion pools, and 2 marshes. rean Anopheles are impossible to distinguish by lar- Larval sampling was performed around the pe- val characteristics, the larvae were reared to the rimeter of each study site with a standard 400-ml adult stage for specific identification. Mosquito plastic dipper. The circumference of each site was rearing was performed in an air-conditioned water- measured then divided by 30 so that 30 dips could testing laboratory at CP Casey. A plastic enclosure be equally spaced around the site. Initially, all was placed over a countertop and the interior was anopheline larvae and a sample of culicine larvae warmed continuously with two 100-W incandes- were removed with pipettes and stored in plastic cent light bulbs. Larvae were exposed to the light bags for transport to the central laboratory at CP continuously. The larvae were transferred to 16-oz Casey. When Anopheles spp. exceeded 50/dip, ap- plastic cups filled with tap water that had been al- proximately lOVo of the larvae were taken for rear- lowed to sit overnight to reduce chlorine residuals. ing. The plastic bags were filled with water from Larvae were fed finely ground Tetramin@ Tropical the study site then stored in an insulated chest for Flakes fish food (Tetra GmbH, Melle, Germany). transport to the laboratory for rearing. All collec- Each 4'h-stage larva was moved to an individual tions were performed between 0700 and 1200 h. plastic rearing vial. The 4th-stage larva and pupal With the exception of those added later in the study, exuvium were placed in SOVoethanol h a 0.25- each habitat was sampled once each month from dram glass vial with a rubber stopper for taxonomic June through September. After each additional site study. The adults were preserved according to pro- was added, it was sampled monthly through Sep- cedures of Belkin and others ( I 965), then identified tember. A value of 0 was recorded for sites that to species according to Lee (1998). Voucher spec- were dry, flooded with running water, or otherwise imens of adults and associated exuviae were de- incapable of supporting larval populations, as well posited at the U.S. National Museum of Natural as for suitable sites with no larvae in 30 dips. History, Smithsonian Institution, Washington, DC. In June and July, water analysis was conducted Statistical analyses: Mean and median densities on 1 of the 30 dips from each site. Nitrate and of Anopheles (number of larvae/3O dips) were de- phosphate concentrations were determined with a termined with the UNIVARIATE procedure in SAS pocket colorimeter (Hach Co., Loveland, CO); total (SAS Institute 2000). Medians were compared with dissolved solids and pH were both determined with the Kruskal-Wallis test. Data with similar coeffi- Hach@ Pocket Pal testers Model 46700-12. cients of variation were compared with Tukey's Percent of water surface covered by duckweed honestly significant difference test. Similar analyses

Table 3. Percentage of area covered with algae in 3 larval habitat types of northern Kyunggi Province, Republic of Korea (June-September 2000).

Standard Coefficient Mean Medianr deviation of variation Range Rice fields 50 7.6 0a 2O-l 2.6 0-80 Ditches2 20 18.9 0a 26-9 1.4 o-75 Pools' 15 19.4 0a 32.3 1.7 0-100

rMedims followed by the same letter are not significantly different (Kruskal Wallis test; P : 0.05). 'Imigation ditches and drainage ditches were combined into I category for statistical analysis. Stream pools and irrigation ponds also were combined. 182 JounNa.r-or rne Ar\4smcnNMclseurro CoNrnor-AssoctATroN Vor-. 18. No. 3

Thbfe 4. Percentage of area covered with Lemna spp. in 3 habitat types of northern Kyunggi Province, Republic of Korea (June-September 2000).

Standard Coefficient Habitat Mean Median' deviation of variation Range Rice fields 50 40.2 30a 38.4 1.0 0-100 Ditches' t8 6.0 0b 74.1 2.4 0-50 Pools and ponds2 l5 2.O 0b o.7 0.4 o-10 'Medians followed by the same letter are not significantly different (Kruskal-Wallis test; P > 0.05). ' Irigation ditches md drainage ditches were combined into I category for statistical malysis. Stream pools md irigation ponds also were combined.

were used to compare environmental parameters by irrigation ditches, but this finding may be due to habitat, with both types of ditches combined into I the small number of specimens reared to the adult category and both types of pools similarly com- stage. Densities of larval Anopheles for most hab- bined. itats were similar in June and July (Table 2). A To develop a predictive model for sites with high trend toward greater larval densities in ponds and densities of larval Anopheles, each environmental stream pools started in July and this trend eventu- parameter was dichotomized at the median and also ally led to significantly greater densities in these at the 75th percentile in 2 different models to allow habitats in August and September when compared analysis by logistic regression. The LOGISTIC pro- to ditches and rice fields. The median number of cedure (SAS Institute 2000) was used to perform larvae per dip in rice fields was never great, al- stepwise logistic regression modeling of the prob- though as many as 50 larvae were collected in in- ability of a site being in the top quartile of larval dividual dips. Generally, distributions of larvae density as a function of water quality and amount were highly clustered within rice fields. of surface covered by vegetation. In September, many rice fields were drained in preparation for harvesting, resulting in the elimi- nation of larval habitats; however, RESULTS very large larval densities developed in the pools, especially irriga- Of the 14 putative species collected during the tion lagoons contiguous to the drained rice fields. study period, only 3 were anophelines (Table l). The pools supported the highest larval densities ob- These mosquitoes were all identified as adult spec- served during the 4-month study. imens by using morphological characteristics de- Tables 3-8 describe environmental factors for scribed in Tanaka et al. (1979) and Lee (1998). each of the habitats. The factors overlapped in their Adult characteristics used to separate An. sinensis ranges and no statistically significant differences and An. lesteri may not be reliable, so the relative were noted between habitats, with the exception of numbers of these 2 species must be considered with percent surface covered by floating duckweed some caution. Nevertheless, the majority of Anoph- (Lemna spp.). Rice fields had significantly greater eles reared to adulthood (81Vo) appeared to be A,?. coverage with this plant, although percent coverage sinensis, with An. lesteri representing only 18.8Vo. was not predictive of larval densities when using Only 3 specimens of An. yatsushiroensi.s were logistic regression techniques (P : 0.45). The sim- reared to the adult stage during the study, repre- ilarity in water-quality parameters is perhaps not senting less than 7Vo of rhe Anopheles. Other spe- surprising given that several of the habitats are in- cies that were identified in significant numbers were terconnected with a complex irrigation system that Aedes vexans (Theobald) and Culex pipiens Lin- moves water from streams. rivers. and wells to ir- naeus. rigation ditches to rice fields then back again. In Anopheles sinensis was collected in all 6 of the some fields, this movement was continuous and re- habitats sampled (Table 1). Anopheles lesteri was sulted in a slow movement of water through the only obtained from rice fields, stream pools, and paddies. Other fields were more stagnant and prone

Table 5. Mean nituate concentration (mg/Iiter) in 3 habitat types of northern Kyunggi Province, Republic of Korea (June-September 20O0).

Standard Coefficient Habitat Meanr deviation of variation Range

Rice fields 5l 0.9a 0.8 0.9 o-2.8 Ditches2 t6 l -+a 1.1 0.8 0 4.0 Pools and ponds' t2 7.2a o.7 0.6 o.4-2.6

'Means followed by the same letter are not significantly different (Tukey's honestly significant difference; P > 0.05). 'lrrigation ditches and drainage ditches were combined into I category for statistical analysis. Stream pools and inigation ponds also were combined. 183 Sep'rrvsen 2002 Mnumn Vr.crons tN KYUNGGI PnovrNcp, Konee

Table 6. phosphateconcentration (mg/liter) in 3 habitat types of northern Kyunggi Province, Republic of Korea (June-SePtember2000).

Standard Coefficient Habitat n Mean' deviation of variation Range

Rice fields 44 O.4a 0.5 1.25 o.1-2.4 Ditches' l8 O.7a 0.6 0.9 o.t-2.r Pools and ponds2 14 0.3a 0.3 t.o 0.0-1.3

'Means followed by the same letter ile not significantly different (Tukey's honestly significant difference; P > 0.05). ponds also 2 lrrigation ditches ind drainage ditches were mmbined into I category for statistical analysis. Stream pools and inigation were combined. to intermittent drying. Environmental factors also between population density of An. sinensis and the were similar for infested and noninfested sites (i.e., pH and temperature of rice field water. However, a fields with and without detectable larval popula- positive correlation was found between the number tions). The range of each factor is reported for in- of immatures and the NH4-N concentration in that fested and noninfested flelds in Table 9. The results study that was not reflected in the nitrate concen- indicate that anopheline larvae in the ROK are pre- tration analysis of our work. sent in habitats with significant variations in water In our study, larvae were collected from a variety quality and plant coverage. of habitats and, at least early in the growing season, Stepwise logistic regression analysis revealed no showed limited identifiable preference for habitat factors that were predictive of high larval densities type. The larvae seemed capable of exploiting most (P > 0.05). The most likely candidate for predict- stagnant or slowly moving water habitats in the ing highly infested fields was percent duckweed study area. However, nearly 26Vo of the sites sam- coverage adjusted for nitrate and phosphate con- pled in July did not support detectable larval pop- : centrations in July (P 0.06), but the overall mod- ulations; the reason for the lack of larval popula- : el was nonsignificant (P O.24). tions in these habitats is unknown but may be The percent of each habitat in which at least I related to chemical fertilizers and pesticides applied larva was detected varied monthly. For rice fields, to the paddies at irregular intervals. Another pos- peak percent infestation (77Vo) was observed in sibility is that the number of potential oviposition July, but fell to 2OVo in September when most of sites exceeded the number of gravid females. the paddies were drained. In contrast, none of the Assuming that most of the Anopheles in the area in June, irrigation pools yielded anopheline larvae were An. sinensis, this malaria vector is capable of but by September, all of them supported significant development in areas with or without floating veg- larval populations. Larval populations in stream etation and in water with a broad range of pH, dis- pools and drainage ditches were variable, because solved solids, and nitrate and phosphate concentra- they were prone to heavy flooding from typhoon- tions. However, other environmental factors not associated rains. investigated during this study may serve as limiting factors. It is important to note that the sampling DISCUSSION method used during this study was designed for Larval Anopheles in the ROK were present in a study of large habitats that could be identified and wide range of the environmental factors investigat- analyzed remotely. A sampling technique that an- ed during this study. Because of the ability of the alyzes at the level of a microhabitat (i.e., algal mats larvae to survive in such a variety of conditions and vs. overhanging vegetation) might be more likely habitats, none of the water-quality or vegetation in- to identify environmental parameters that are pre- dices recorded in this study were reliable predictors dictive of vector abundance at a different scale. of high larval density. This conclusion is consistent The species collected during this study occurred with previous research by Ikomoto and Sakaki in different proportions than they did in a contem- (1979), who found a lack of consistent correlation porary adult trapping study in the same study site

Tabte 7. Values of pH in 3 habitat types of northem Kyunggi Province, Republic of Korea (June-September 2OOO).

Standard Coefficient Habitat Mean' deviation of variation Range Rice fields 38 7.5a 1.1 0.1 5.7-9.6 Ditches' 16 7.4a l.l o.l 6.3-10-0 Pools and ponds2 74 8.0a 1.2 o.2 6.2-9.6

'Mems followed by the sme letter ile not significantly different (Tukey's honestly significmt difference; P > 0-05). 2 lnigation ditches md drainage ditches were combined into I category for statistical malysis. Strem pools and inigation ponds also were combined. 184 Jouruvll op rnt AwentcaN Mosqurro CoNrnol AssocrerroN VoL. 18, No. 3

Table 8. Total dissolved solids (ppm) in 3 habitat types of northern Kyunggi province, Republic of Korea (June- September2000).

Standard Coefficient Habitat Meanr deviation of variation Range Rice fields 38 759a 80.5 0.5 o-375 Ditches, 16 2OOa 78.8 o.4 I l6-438 Pools and ponds, t4 1'l8a 55.8 0.3 106-285 'Means followed by the same letter are not sign.ificantly different (Tukey's honestly significant difference; p > 0.05). 'Inigation ditches and drainage ditches were combined into I category for statistical analysis. Stream pools and inigation ponds also were combined.

(Burkett et al. 2001). In particular, very few An. rice paddies were drained, larval densities in the yatsushiroensjs were obtained during this larval irrigation pools greatly increased. In fact, the great- study, whereas this species comprised nearly 48To est larval densities during the entire 4 months oc- of Anopheles captured with a Shannon trap during curred in September in the stream pools and irri- the same time in the same location. Several factors gation lagoons. A larval control program directed may contribute to the inconsistency between the toward pools and ditches late in the season would larval and adult studies. Differential mortality in eliminate the greatest concentrations of the vector the rearing process may have occurred that resulted population, although the relative size of the rice in the death of most of the larval An. yatsushiroen- field habitat would still make this habitat a major sis. Mortality was significant throughout the study, contributor to overall vector numbers. with less than 2OVo of the larvae attaining adult- Several aspects of the biology of malaria vectors hood. Alternatively, the distribution of this species in the ROK must be addressed in future studies. in rice paddies may be such that perimeter dipping First, the larval habitat of An. yatsushiroensis does not reflect the proportion of species present in should be identified. This species may play some the rice field habitat. Finally, another habitat may role as a malaria vector and thus might be important exist that is exploited preferentially by An. yatsu- in the epidemiology and control of this disease. possibilities. shiroensis. Of these 3 the 3rd seems Second, other environmental factors should be in- most likely. The adult trapping performed by Bur- vestigated as possible predictors of the presence kett and others was done near CP Greaves, an area and abundance of larval Anopheles. One factor that around which land mines remain from the Korean seems to have a substantial impact on larval density War. These mines prevented larval sampling in that was not addressed in this study is the pattern some areas, especially in wooded, hilly areas, and of flooding and draining in the rice fields. Many along the banks of the Imjim River. Habitats within fields were drained in the middle of the growing these areas or habitats within the river may serve season, perhaps accidentally. More attention should as the preferred larval habitats of An. yatsushiroen- be directed to the effect of flooding and draining (1979) sls. Tanaka et al. reported that this species on species composition and abundance within the is common in rice fields but that it is more common rice field habitat. Third, the in hilly or mountainous areas. in the ROK must be clarified before specific work The densities of anopheline larvae were similar on the biology and control of members of this ge- for most habitats in June. In Septembeq when the nus can be completed. In particular, the ability to distinguish between An. lesteri and An. sinensis by using adult morphological characteristics is ques- Table (range) 9. Mean of environmentalvariables in tionable. At present, numbers of specimens for study sites with and without larval Anopheles,Kyunggi these species from this study are tentative until fur- Province, Republic of Korea (June-September2000). No significant differenceswere noted between infested and ther systematic work to validate species identifica- noninfestedhabitats (Kruskal-Wallis; P : 0.05). tion is completed. This paper reports the lst part of a larger study Larval presence Environmental that used remote sensing to determine the size and factor Yes No quality of larval habitats of Anopheles within the flight range around 2 U.S. military bases in the Vo algal coverage r r.3(0-80) 13.8(0-100) 7a Lemna spp. cov- ROK. This information will be used to estimate the erage 31.7(0-100) 17.3(0-r00) cost of a larviciding program and to compare that Nitrates (mg/liter) 0.9(0-2.8) r.4(o.r-4.o) estimate to the cost of providing chemoprophylaxis. Phosphates (mg/ It should be noted that the U.S. government is not liter) o.4(o-2.4) 0.4(0.0s-2.0s) considering any unilateral larviciding efforts, and pH 7.6(s.9-9.9) 7.6(s.7-r0) would implement such a malaria control program Dissolved solids only with the support and concurrence of ROK (ppm) 164.0(0-375) 182.0(103-438) governmental authorities and local landowners. SsprevgEn 2002 MALARTA VEcroRs tN KvuNcct PpovINcs, Konse 185

ACKNOWLEDGMENTS (Diptera: Culicidae) larval habitats in Belize' Central America. J Med Entomol 33:2O5-211. grateful to the soldiers of the We are extremely Manguin S, Roberts DR, Peyton EL, Rejmankova E, Pe- 702nd Preventive Medicine Section; the 2nd Infan- cor J. 1996b. Characterization of Anopheles pseudo- try Division; the 5th, 38th, and l54th Medical De- punctipennis larval habitats. J Am Mosq Control Assoc tachments; the l68th Medical Battalion (Area Sup- 12:619-626. port); and the 18th Medical Command for their Moncayo AC, Edman JD, Finn JT, 2000' Application of valuable assistance in conducting larval surveil- geographic information technology in determining risk virus transmission. lance during the field phase of this study. Special of eastern equine encephalomyelitis Mosq Control Assoc 16:.28-35- thanks go to McKinley Rainey, William Herman, J Am Ree HI, Self LS, Kong HK, Lee KW. 1973. Mosquito light Kenneth McPherson, and Alex Ornstein for their trap surveys in Korea (1969-1971). Southeast Asian.I in providing personnel during this study. support Trop Med Public Health 4:382-389. Kwan- Thanks also are due to Hung-chol Kim and Rejmankova E, Pope LO, Roberts DR, Lege MG, Andre woo Yi, who provided valuable technical assistance RG. Greico J. Alonzo Y. 1998. Characterization and in specimen identification. The manuscript was re- detection of Anopheles vestitipennis and Anopheles viewed by Richard Thomas, Tomoko Hooper, Su- puctimaculata (Diptera: Culicidae) larval habitats in san Langreth, and Art Lee. Funding was provided Belize with field survey and SPOT satellite imagery. "I by the Department of Defense, Global Emerging Vector Ecol 23:74-88. Masuoka Andre Infections System, U.S. Army Medical Research Roberts D, Vanzie E, Rejmankova E, B 1999. Use of remote sensing and geographic infor- and Materiel Command's Military Infectious Dis- R. mation systems to &[get malaria control measures in ease Research Program, and National Aeronautics Belize, Central America. SCOPE Malaria Research and (grant NAG5-8532). and Space Administration Policy Forum (Commentary Article, 15 December). of the re- This manuscript is in partial fulfillment Berkeley, CA: University of Califomia. http://www. quirements for the Doctorate of Public Health at the scope.educ.Washington. edu/research/malaria./ddt-ban/ Uniformed Services University of Health Sciences rcgertsl1999-12. by D.M.C. Roberts DR, Rodriguez MH. 1994. The environment' re- mote sensing and malaria control. Ann N Y Acad Sci REFERENCES CITED 74O:396-402. Rodriguez AD, Rodriguez MH, Hernandez JE, Dister SW, Aitken TGG, Shrick R' Belkin JN, Hogue CL, Galindo f; Beck LR, Rejmankova E, Roberts DR. I 996. Landscape Mosquito studies (Diptera: Culici- Powder WA. 1965. surrounding human settlem ents and Ano p heI es a I b iman- for the collection, rearing and preser- dae). II- Methods zs (Diptera: Culicidae) abundance in southern Chiapas, Contrib Am Entomol Inst (Ann Ar' vation of mosquitoes. Mexico. J Med Entomol 33:39-48. 7:19--78. bor) SAS Institute Inc. 2000. SAS/STAT@ user's guide, version Brunetti R, Fritz RR Hollister AC. 1954. An outbreak of 8 4th ed. Volume L Cary, NC: SAS Institute Inc. malaria in California, 1952-1953.Am J Trop Med Hyg Savage HME, Rejmankova E, Arrendondo-Jimenez JI, 3:'7'19-788. Roberts DR, Rodriguez MH. 1990. Limnological and bo- Burkett DA, Lee WJ, Lee KW, Kim HC, Lee HI, Lee JS, tanical characterization of larval habitats for 2 primary Shin EH, Wirtz RA, Cho HW Claborn DM, Coleman malarial vectors, Anopheles albimanus and Anopheles RE, Klein TA. 2001. Light, carbon dioxide and octenol- pseudopunctipennis, in coast areas of Chiapas State, baited mosquito trap and flight activity evaluation Mexico. J Am Mosq Control Assoc 6:613-620. againstmosquitoes in a malarious area of the Republic D, Miller ME, Kelsey LL, l-,en WJ, I-ee HW of Korea. J Am Mosq Control Assoc 17:196-2O5. Strickman Kim HC, Feighner BH. 1999. Evaluation of the Chow CY. 1973. Arthropods of public health importance l-ee KW Multipurpose Range Complex, in Korea. Korean J Entomol 3:31-54. malaria threat at the Korea. Mil Med 164:626-629. FeighnerBH, Pak SI, Novakoski WL, Kelsey LL, Strick- Yongp'yong, Republic of Kim HC, Wirtz RA, man D. 1998. Re-emergenceof Plasmodiumvivax ma- Strickman D, Miller ME, Lee KW, Roh 2001. laria in the Republic of Korea. Emerg Infect Dis 4:295- Perich M, Novakoski WL, Feighner BH, CS. Anop he - 298. Successful entomological intervention against Grillet ME. 20OO.Factors associatedwith distribution of les. sinensis limiting transmission of Plasmodium vtva-x Anophelesaquasalis and An. oswaldoi (Diptera: Culic- to American soldiers in the Republic of Korea. Korean idae) in a malarious area, northeast Yenezuela. J Med J Entomol 31:189-195. Entomol 37:237-238. Tanaka K, Misusawa K, Saugstad ES. 1979. A revision of (including Ikomoto I Sakaki l. 1979.Physico-chemical characters of the adult and larval mosquitoes of Japan the the water in rice fields in relation to their suitability for Ryukyu Archipelago and the Ogasawara Islands) and breeding of the mosquito larvae, Anopheles sinensis. Korea (Diptera: Culicidae). Contrib Am Entomol Inst Jpn J Sanit Zool 30:87-92. (Ann Arbor) 16:l-987. Joo CY, Kang GT. 1992. Epidemio-entomologicalsurvey Walter Reed Army Institute of Research. 1999. Addressing on malarial vector mosquitoesin Kyongbuk, Korea. Ko- emerging infectious disease threats: a strategic planfor rean J Parasitol 3:329-340. the Department of Defense Bethesda, MD: Division of Lee KW. 1998. A revision of the illustrated taxonomic Preventive Medicine, Walter Reed Army Institute of keys to genera and speciesoffemale mosquitoesof Ko- Research. rea (Diptera: Culicidae) Seoul, Korea: 5th Medical De- Wood BR, Washino R, Beck L, Hibbard K, Pitcairn M, tachment, 168th Medical Battalion, lSth Medical Com- Roberts D, Rejmankova E, Paris J, Hacker C, Salute J, mand,U.S. Army. Sebesta P, Letgers L. I 991. Distinguishing high and low Manguin S, RobertsDR, Andre RG, RejmankovaE, Hak- anopheline-producing rice fields using remote sensing re S. 1996a. Characterization of Anopheles darlingi and GIS technologies. Prev Vet Med 11:27'7-282.