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Bionomics of Aedes aegypti subpopulations (Diptera: Culicidae) from Argentina

2010; Wiley; Volume: 35; Issue: 2 Linguagem: Inglês

10.1111/j.1948-7134.2010.00083.x

1948-7134

Marta G. Grech, Francisco Ludueña-Almeida, Walter R. Almirón,

Insect Pest Control Strategies

Differences in biological features of immature and adult Aedes aegypti, as well as variability in vector competence, seem consistent with the existence of genetic variation among subpopulations and adaptation to local conditions. This work aims to compare the bionomics of four Ae. aegypti subpopulations derived from different geographical regions reared under temperate conditions. Life statistics of three Ae. aegypti subpopulations from the provinces of Córdoba, Salta, and Misiones were studied based on horizontal life tables. The Rockefeller strain was used as a control. The development time required to complete the larva and pupa stages varied from 6.91 to 7.95 and 1.87 to 2.41 days, respectively. Significant differences were found in mean larval development time between the Córdoba and Orán subpopulations. The larva-pupa development time was similar in all the subpopulations. However, survival values varied significantly between the Orán and San Javier subpopulations. The proportion of emergent males did not differ from females within each subpopulation nor among them. Adult longevity was similar among the subpopulations. The average number of eggs laid by each female was significantly different. The Rockefeller strain laid a significantly greater number of eggs (463.99 eggs/female) than the rest of the subpopulations. Moreover, differences in the demographic growth parameter Ro were detected among the four subpopulations. The differences obtained in larval development time, larva-pupa survival values, and net reproductive rates among the subpopulations might reflect underlying genetic differences as a result of colonization from different regions that probably involve adaptations to local conditions. Aedes aegypti (L.) has long been studied because of its importance as a vector of viral diseases such as dengue and urban yellow fever (Christophers 1960, Gubler 1988, Clements 1992, PAHO 1995). Governments and health organizations have invested considerable effort in eradicating Ae. aegypti from the Americas. Eradication programs initiated in 1915 achieved the apparent elimination of Ae. aegypti from most of the region by 1970, utilizing vertically-structured programs (PAHO 1994, Gubler and Clark 1994). By 1964, these mosquitoes were considered eradicated from Argentina (Kerr et al. 1964, Bejarano 1979, Gubler 1988, Nathan 1993). The first re-infestation of Argentina was reported in 1986 in the provinces of Formosa and Misiones, and in 1991 Ae. aegypti was detected in Buenos Aires province. In 1995, the Ministry of Health of Córdoba province reported the presence of Ae. aegypti and confirmed the establishment of this mosquito in Córdoba city (Carcavallo and Martínez 1968, Bejarano 1979, Campos 1993, Campos and Maciá 1996, Almirón and Ludueña-Almeida 1998, Coto et al. 2003). The re-infestation in Argentina might have entered the northwestern provinces from Bolivia and the northeastern provinces from Paraguay and Brazil (Rondán-Dueñas et al. 2009). In Argentina, the current geographical distribution of Ae. aegypti is wider than before its eradication (Curto et al. 2002). The distribution has expanded toward the west (Mendoza province) (Domínguez and Lagos 2001) and south (La Pampa province) (Rossi et al. 2006), in comparison to the historical distribution. Between April and November 1997, 19 cases of dengue fever by DEN-2 serotype were detected in the localities of Orán, Salvador Maza, Güemes, and Tartagal (Salta province) in northwestern Argentina (Avilés et al. 1999). In 1998, the first epidemic of dengue was recorded in Salta province with 359 confirmed cases by the same serotype and it was thought most likely to be linked to an under-reported outbreak of DEN-2 in Bolivia in 1996 and 1997 (Gianella et al. 1998, Van der Stuyft et al. 1998). During 2000, a second epidemic of DEN-1 serotype was recorded in Argentina, affecting the provinces of Misiones and Formosa (in the northeast). Both the 1998 and 2000 epidemics were associated with outbreaks in the neighboring countries of Bolivia, Brazil and Paraguay (Avilés et al. 2003). During 2002, 214 cases by DEN-1 were identified in Salta province, and the DEN-3 serotype was recorded for the first time in Misiones province. The emergence of DEN-3 in Salta province in 2003 (co-circulating with DEN-1 and DEN-2) was shortly followed in 2004 by significant outbreaks of DEN-3 there and in the neighboring provinces of Jujuy and southwestern region of Formosa (National Ministry of Health 2006). During the summer-autumn of 2009, an unprecedented epidemic of DEN-1 affected the north-western provinces reaching central areas of the country, with autochthonous cases recorded for the first time in provinces such as Catamarca, Chaco, Córdoba, Corrientes, Buenos Aires, Santa Fe, Santiago del Estero, and Tucumán (National Ministry of Health 2009). According to Dègallier et al. (1988), the biological features of Ae. aegypti larva and adults seem to vary according to local conditions. Based on genetic studies of Ae. aegypti subpopulations from Argentina using molecular markers, great differences were detected in haplotype compositions between different regions of the country (de Sousa et al. 2000, 2001, Rondán-Dueñas et al. 2009). Tejerina et al. (2009a) reported differences in life statistics between subpopulations of Ae. aegypti from four localities from the north-east of Misiones province (Argentina). Moreover, variability in vector competence of these mosquitoes in several subpopulations has also been demonstrated (Beerntsen et al. 2000). At present, the best option to prevent epidemic outbreaks and reduce dengue transmission is to control the vector populations. Since there is autochthonous transmission of dengue virus and also the introduction of cases of dengue and yellow fever from neighboring countries (Bolivia, Brazil, Paraguay), it is necessary to study the ecological features of Ae. aegypti populations to understand the dynamics of viral diseases and be able to implement effective control programs. Therefore, the aim of this work was to compare life statistics of four Ae. aegypti subpopulations derived from different geographical regions (the provinces of Córdoba (Córdoba city), Salta (San Ramón de la Nueva Orán city –hereafter Orán), and Misiones (San Javier city)), reared under temperate conditions, because reinfestation is moving south. The Rockefeller strain provided by the CDC of Puerto Rico was used as a control for comparison with a strain highly adapted to laboratory conditions. Aedes aegypti subpopulations were from the cities of San Javier, Orán, and Córdoba (Figure 1). San Javier city (27º52' S; 55º08' W) is located in the southeastern region of the subtropical province of Misiones, along the banks of the Uruguay River opposite the Brazilian city of Porto Xavier. Annual average cumulative rainfall for San Javier is 1,948 mm and the mean maximum, average, and minimum temperatures are 27.6° C, 21.5° C, and 16.6° C, respectively. Entomological surveillance data were estimated for Misiones province (Colonia Delicia city) in an area in which a dengue outbreak took place. The House index (HI) was 51% and the Breteau index (BI) 106, recorded during 2000 (Masuh et al. 2003). In the city of Posadas, capital of Misiones Province, breeding sites found in the homes were buckets of 10 to 20 liters, tires, and containers up to one liter (Tejerina et al 2009b). Pupal surveys conducted in Formosa province (Clorinda city, northeastern Argentina) during the fall and spring 2007 showed that large tanks used for drinking water storage were both the most abundant and the most productive types of container (Garelli et al. 2009). Map of Argentina showing location of collection sites. Or = Orán city (Salta Province), SJ = San Javier city (Misiones Province), Co = Córdoba city (Córdoba Province). Orán (23º08' S; 64º20' W) is an endemic dengue area in the northeastern region of the subtropical province of Salta, 270 km north of Salta, capital city of the province. Annual average cumulative rainfall for Orán is 1,067 mm and the mean maximum, average, and minimum temperatures are 27.7°, 21.2°, and 16.4° C, respectively (National Meteorological Services 2009). The National Coordination for Vector Control of the National Ministry of Health recorded entomological data from Orán city, showing HI and BI of 6.46% and 8.9, 14.28% and 29.27, and 22.49% and 46.26 during February 2003, 2004, and 2005 respectively (Estallo et al. 2008). No specific information about Ae. aegypti breeding containers is available for the northwestern region of Argentina. Córdoba city (31º22' S; 64º12' W), in the central-western area of Córdoba province, is the main city and capital of the province, with a temperate climate. Annual average cumulative rainfall for Córdoba city is 869.9 mm. The mean maximum, average, and minimum temperatures are 24.03°, 17.28°, and 11.21° C, respectively (National Meteorological Service 2009). Entomological surveys were performed in Córdoba city during the summer months (December-February) of 1997–2000 and during February 2004, 2005, and 2009, in order to determine traditional larval indices. The infestation values during 1997–1998 were HI = 23%, CI = 16%, and BI = 47, but during 1999–2000 the infestation values were higher (HI = 47%, CI = 24%, and BI = 106). During 2004, 2005, and 2009, the HI, CI, and BI were 33%, 7%, and 77, 29%, 13%, and 70, and 25%, 7%, and 34, respectively (Almirón and Asis 2003, Estallo et al. 2009). Larvae and pupae of Ae. aegypti were found in a great variety of containers, with discarded tires, buckets, cans, and tins being those most commonly used as breeding sites according to entomological surveys performed in six neighborhoods of Córdoba (Almirón and Asis 2003). Aedes aegypti used in this study were obtained from laboratory colonies established from field immature stages collected in the cities of San Javier, Orán, and Córdoba; mass rearing techniques by Gerberg et al. (1994) and Domínguez et al. (2000) were followed. The source of colony material came from a large number of households and containers from each city. The material collected was transferred to Córdoba city, where the colonies from each locality were developed simultaneously in the laboratory of the Córdoba Entomological Research Center (National University of Córdoba). Larvae and pupae collected from artificial containers were laboratory reared. Larvae were kept in 750 ml water-filled trays and fed with liver powder (0.25 mg/larva/day). Water surface was daily cleaned with filter paper to avoid complications due to fungus and bacteria development. Pupae were put in plastic flasks and then placed in cardboard entomological cages (30 cm diameter, 50 cm high). The emerging adults were provided with 10% sugar solution soaked in cotton wick on 70-ml plastic flasks. Females were offered a restrained mouse for blood meals for two h twice a week. For oviposition, plastic flasks with filter paper on the inside wall and dechlorinated water were placed inside the cages. Eggs remained on moist filter paper for at least four to five days to ensure embryogenesis, and then were air-dried to store until the experiment started. When there were enough eggs from each locality, they were immersed in water for hatching. Four cohorts of 30 1st instar larvae were established and followed simultaneously for each subpopulation; experimental material was the first generation progeny from individuals that were field-collected as larvae. Each cohort was kept in a 750 ml water-filled tray, the larvae fed with liver powder and monitored daily, as previously described. Life statistics were estimated by horizontal life tables from these cohorts. Aedes aegypti mass rearing was conducted in Córdoba City, in the laboratory of the Córdoba Entomological Research Center (National University of Córdoba), between November 2006 and March 2007. During this period, daily mean temperatures ranged from 18.5° to 28° C, with a mean value of 22.9° C (mean maximum = 24.7° C; mean minimum = 21.1° C). The mean relative humidity was 59.1% and the natural photoperiod was approximately 13 h of light. The number of days spent in each preimaginal life stage and numbers of survivors in each stage were recorded daily. Survival was expressed as the percentage of individuals that reached the next instar/stage. The sex ratio of emerged adults was also estimated (Gómez et al. 1977). Adults that emerged from each cohort were placed in a cardboard cage (an average of ten males and eight females emerged per cohort). The number of living and dead males and females was recorded daily. Adults were provided with a 10% sugar solution daily. Females were offered a restrained mouse for blood meals twice a week, starting two days after the first female of the cohort emerged. Once the mouse was removed from the cage, a plastic flask with filter paper and dechlorinated water was placed inside the cage and replaced daily. Male and female longevity as well as fecundity were estimated. Daily fecundity was expressed as the number of eggs laid/female/day. Survival data (lx) and daily fecundity of females (mx) were used to calculate mean net reproductive rate (Ro) for each subpopulation. Net reproductive rate was expressed as , the summation of mean fecundity at age xtimes the probability of survival to stage xi. R0 expresses the per-generation growth rate of the population as the total number of female eggs a mosquito lays during her lifetime (R0>1.0 the population increases in size, R0= 1.0 no increase in population size and R0>1.0 population growth is declining) (Carey 1993, Service 1993, Irvin et al. 2004). The development time and survival in each preimaginal stage, adult longevity, oviposition time, and fecundity were analyzed with ANOVA followed by the Duncan's Test to detect significant differences and determine whether these life statistics varied among the four subpopulations studied. Sex ratio in each cohort was analyzed by the Student t-test for differences of proportions (Steel and Torrie 1988). Sex ratio among subpopulations was analyzed by ANOVA, following arcsin data transformation. All the data were checked for normality. Mean net reproductive rates were estimated by jackknife analysis of lxmx life table data and compared by using ANOVA followed by the Duncan's test. The jackknife method recombines the original data by omitting one of the n replicates from the jackknife sample and calculates pseudovalues of the parameter of interest for each recombining of the data. These new estimates form a set of numbers from which mean values and variances can be calculated and compared statistically. The jackknife method is well-suited for estimating variance for population growth statistics (Miller 1974, Meyer et al. 1986, Shao and Tu 1995, Krebs 1999). Significant differences were found in the mean larval development time between Córdoba (6.91 days) and Orán (7.95 days) subpopulations (F=8.90; d.f=(3;12); P=0.0022) (Table 1). The Córdoba subpopulation had a slightly lower development time for instars 1+2 (2.41 days), analyzed together, which differed significantly from the San Javier and Orán subpopulations (F=3.86; d.f.=(3;12); P=0.0383) (Table 1). The longest development time for the 4th instar (3.21 days) was recorded for the Orán subpopulation (F=5.28; d.f=(3;12); P=0.0149), differing significantly from the San Javier subpopulation and the Rockefeller strain (Table 1). Pupal development time did not differ significantly among the four subpopulations, ranging between 1.87 and 2.41 days (F=2.94; d.f.=(3;12); P=0.0761) (Table 1). Larvae and pupae completed their development in approximately 9.5 days, with no significant differences detected among the four subpopulations (F=1.71; d.f.=(3;12); P=0.2175) (Table 1). The lowest survival was observed for the Orán subpopulation, for the larval stage (73.3%) as well as for the larval and pupal stages analyzed together (59.2%), differing significantly from the San Javier subpopulation (95% and 85.8%, respectively) [F=4.94; d.f=(3;12); P=0.0185; F=7.59; d.f=(3;12); P=0.0042] (Table 2). The survival recorded for the pupal stage varied between 80.2 and 91.7% but no significant differences were observed among the 4 subpopulations (F=1.67; d.f=(3;12); P=0.2268) (Table 2). Pupal survival was higher than larval survival in the subpopulations of Córdoba, Orán, and the Rockefeller strain. No significant differences were detected between the mean number of males and females that emerged in each subpopulation (Table 3). Although no significant differences were found in the sex ratio from 1:1, more males were recorded in the four subpopulations (F=0.37; d.f.=(3;12); P=0.7748) (Tables 2[link] and 3). Mean of Aedes aegypti fecundity for four subpopulations studied. SJ = San Javier, CO = Córdoba, OR = Orán, ROC = Rockefeller. Mean male and female longevity varied from 15.7 days (Orán) to 33.8 days (Rockefeller strain), and 35.7 days (San Javier) to 43.8 days (Orán), respectively (Table 4). Longevity of adults was similar among the four subpopulations (Table 4), for both females (F=0.19; d.f.=(3;12); P=0.8995) and males (F=2.87; d.f.=(3;12); P=0.0804). Females lived longer than males in all subpopulations. The greatest oviposition time was recorded for females from the subpopulation of Córdoba (31.3 days), followed in decreasing order by females from the Orán (29.5 days) and San Javier (29.1 days) subpopulations, and the Rockefeller strain (26.74 days) (Table 4), but no significant differences were detected (F=0.12; d.f.=(3;12); P=0.9495). Significant differences were found in fecundity values (F=11.48; d.f=(3;12); P=0.0008) (Figure 2), with the Rockefeller strain females showing the highest daily fecundity (10.9 eggs/female/day) and fecundity values as well as total number of eggs laid. In decreasing order follow the subpopulations of San Javier (2.6 eggs/female/day), Córdoba (2.0 eggs/female/day), and Orán (1.4 eggs/female/day) (Table 4). Mean net reproductive rate was highest for the Rockefeller strain (R0= 134.22), differing significantly from the San Javier (R0= 27.04), and the group formed by the Córdoba (R0= 9.71) and Orán (R0= 10.33) subpopulations. Differences in the demographic growth parameter R0were detected among the four subpopulations (F= 380.26; d.f. = 3;12); P=0.0001) (Table 4). Larval development times obtained in this study (among 6.91 and 7.95) were lower than the 9.65 days at 20° C reported in an earlier study (Bar-Zeev 1958) under laboratory conditions. Domínguez et al. (2000) reported that under temperate semi-natural conditions (Córdoba city), larval development time was 8.9 days at 22.1° C. However, under field conditions (26° C) in the temperate city of La Plata (Buenos Aires province, Argentina), Maciá (2006) obtained a larval development time that varied from four to 42 days for both sexes, with a mean of 8.3 days and 10.5 days for males and females, respectively. Under subtropical semi-natural conditions in Misiones province, a larval development time of 5.8 days at 25.6° C was reported by Tejerina et al. (2009a). Temperatures lower than 20º C, even for a few hours a day, interfere in the normal development of larvae (Bejarano 1956). Pupal development time obtained in this study was lower than the three days reported by Bar-Zeev (1958) and Domínguez et al. (2000). Tejerina et al. (2009a) also found a pupal development time of 2.5 days in the subtropical province of Misiones. The differences obtained here for the larval development time of Córdoba and Orán subpopulations may reflect underlying genetic differences that probably imply local life history differentiation. The Córdoba subpopulation had shorter development times, probably because it is better adapted to local environmental conditions in Córdoba city, where the duration of favorable temperatures permitting survival of adults and immature stages is shorter than in subtropical cities. Isozyme analyses conducted after the range expansion of Aedes albopictus in the United States were interpreted as indicating rapid local differentiation (Black et al. 1988). Rondán-Dueñas et al. (2009) reported different haplotype composition among the different geographical regions in Argentina. Moreover, high levels of genetic polymorphism were detected in Argentinian Ae. aegypti subpopulations, which suggests different origins of different subpopulations (de Sousa et al. 2000). Biber et al. (2006) found resistance values for cohorts from Córdoba similar to those observed for cohorts from Yacuiba (southern Bolivia, bordering with the NW of Argentina). Larval and pupal survival values were close to 100% in all subpopulations from Misiones province, including the San Javier subpopulation (Tejerina et al. 2009a). However, the same subpopulation reared here in Córdoba city under laboratory conditions has shown survival values close to 86%. This probably reflects that the San Javier subpopulation is well-adapted to local environmental conditions. Larval survival value obtained here was higher than the 76.75% recorded by Domínguez et al. (2000) for Ae. aegypti reared under semi-natural conditions during summer in Córdoba city, but pupal survival was similar (96.4%). The differences obtained in larval-pupal survival between Orán and San Javier subpopulations, both from subtropical regions, probably reflect differences in haplotype composition as a result of colonization from different regions of South America and also an adaptation to local conditions if appropriate selection existed. Previous studies reported that subpopulations of NW and NE Argentina were genetically differentiated, probably because they have received different samples of the gene pool of the species from Bolivia and Brazil, respectively, where Ae. aegypti was established much earlier than in Argentina (de Sousa et al. 2001). The current gene pool of the subpopulations of NW Argentina may be the result of remaining populations that survived the last eradication campaign and of a recent colonizing event that may have occurred in the northwestern provinces from Bolivia, whereas the genetic similarity between the subpopulations of NE Argentina, south Brazil, Paraguay, and Uruguay suggest the transportation of mosquito vectors between cities due to international travel and commerce (Rondán-Dueñas et al. 2009). As in this study, a sex ratio of 1:1 was also reported in Misiones province (Tejerina et al. 2009a) and for Ae. aegypti adults emerging from larvitraps in the temperate province of Buenos Aires (central-western Argentina) (Campos and Maciá 1996, Maciá 2006). However, a subpopulation from Córdoba city had a slight to high preponderance of females, but only when mosquitoes were reared during the summer months (Domínguez et al. 2000). Female longevity varied significantly among subpopulations from different localities of Misiones province, appearing to be longer in subpopulations where insecticide application is more frequent (Tejerina et al. 2009a). Females from the San Javier subpopulation (eastern Misiones province) reared in Misiones province showed less longevity (11.45 days) (Tejerina et al. 2009a) than the same subpopulations reared in Córdoba city (20.46 days). Under laboratory conditions, longevity was highly variable, with a minimum of one day and a maximum of 76 days for both females and males, showing the genetic plasticity of Ae. aegypti and its potential to survive when environmental conditions are favorable (Muir and Kay 1998, Fouque et al. 2006). The mean number of eggs laid by Ae. aegypti females under natural conditions (La Plata city, Buenos Aires province) averaged 59.8 eggs/female, which were reared as larvae at low densities (Maciá 2006). Daily fecundity obtained in this study was lower than the mean value of 4.5 eggs/female/day for the San Javier subpopulation, reported by Tejerina et al. (2009a). The fecundity values reported in this study revealed that, in subpopulations from NE and NW of Argentina, there is a general trend that separates both subtropical region subpopulations. The reproduction pattern observed for Ae. aegypti populations from tropical and subtropical regions indicates that reproduction occurs during the whole year and their abundance may be either associated with rainfall regimens or not (Sheppard et al. 1969, Moore et al. 1978, Chadee 1991, 1992, Barrera et al. 1997, Micieli and Campos 2003). In temperate regions, winter temperatures affect mosquito abundance. A discontinuous reproduction pattern was reported in Ae. aegypti populations from Buenos Aires province as a result of low winter temperatures, and its abundance may be limited mainly due to temperature (Campos and Maciá 1996, Schweigmann et al. 2002). A similar reproduction pattern was reported for Córdoba province, and the abundance was associated with both temperature and rainfall regimens (Avilés et al. 1997, Domínguez et al. 2000). In northern Argentina, however, the reproduction of Ae. aegypti populations showed different patterns. High temperatures allow reproduction during the whole year in Salta and Misiones provinces (Micieli and Campos 2003). However, for Resistencia city (Chaco province, subtropical region) the reproduction pattern was similar to that observed in temperate regions (Stein et al. 2002). Coinciding with survival and fecundity values reported in this study, the San Javier strain had the highest mean net reproductive rate compared with Orán and Córdoba subpopulations (Table 4). Once again there is a trend that separates central and northwestern from northeastern subpopulations. Irvin et al. (2004) reported that the mean net reproductive rate was higher for the nontransformed laboratory Ae. aegypti Orlando strain (Ro= 87.23) when comparing results with the other transformed mosquito strains with Ro between 25.26 and 47.72. Environmental factors as well as genetic composition exert a strong influence on the response of mosquito subpopulations of the same species under different environmental conditions. The results of our study clearly show differences in Ae. aegypti life statistics among the subpopulations studied. We observed differences between temperate and subtropical subpopulations when they were reared in a common temperate environment, and these differences may reflect underlying genetic differences that probably imply adaptation to local conditions. Therefore, it would be interesting to repeat this experiment with the same subpopulations under subtropical conditions and see how the Córdoba strain performs under the conditions under which the Orán and San Javier strains normally exist. The subpopulation from the temperate region (Córdoba) presented a shorter development time than subtropical region subpopulations (Orán and San Javier). In the temperate climate of Córdoba province, this may contribute to the vector competence of Ae. aegypti since autochthonous dengue cases were reported for the first time in this province in 2009. It is also important to note the differences in larval-pupal survival reported here between the subtropical region subpopulations, one genetically associated with subpopulations from Bolivia and the other with subpopulations from Brazil. The Aedes aegypti subpopulation from Orán (northwestern Argentina) showed the longest larval development time, the lowest percentage of larva and pupa survival, and the lowest values of fecundity in a temperate region. However, the vector competence of this subpopulation in a subtropical region is still quite good since several dengue epidemics have been recorded in Orán. The authors thank Iliana Martinez for her helpful comments and suggestions that improved the quality of this manuscript. This work was supported by grants from FONCYT (Fondo para la Investigación Científica y Tecnológica), CONICET (Consejo Nacional de Investigaciones Científicas y Técnicas), SECYT (Secretaría de Ciencia y Tecnología), and UNC (Universidad Nacional de Córdoba). WRA is a scientific member of CONICET, Argentina. MGG is a scholarship holder of CONICET, Argentina.