SciELO - Scientific Electronic Library Online

vol.58 issue3No impact of Bt soybean that express Cry1Ac protein on biological traits of Euschistus heros (Hemiptera, Pentatomidae) and its egg parasitoid Telenomus podisi (Hymenoptera, Platygastridae)Low malathion concentrations influence metabolism in Chironomus sancticaroli (Diptera, Chironomidae) in acute and chronic toxicity tests author indexsubject indexarticles search
Home Pagealphabetic serial listing  

Services on Demand




Related links


Revista Brasileira de Entomologia

On-line version ISSN 1806-9665

Rev. Bras. entomol. vol.58 no.3 São Paulo July/Sept. 2014 



Thermal hygrometric requirements for the rearing and release of Tamarixia radiata (Waterston) (Hymenoptera, Eulophidae)



Mariuxi Lorena Gómez-TorresI; Dori Edson NavaII; José Roberto Postali ParraI

IDepartamento de Entomologia e Acarologia, Escola Superior de Agricultura "Luiz de Queiroz", Universidade de São Paulo, Avenida Pádua Dias, 11, 13418–900, Piracicaba-SP, Brasil.,
IILaboratorio de Entomologia, Embrapa Clima Temperado, BR 392 km 78, 96010–971, Pelotas-RS, Brasil.




Thermal hygrometric requirements for the rearing and release of Tamarixia radiata (Waterston) (Hymenoptera, Eulophidae). Tamarixia radiata is the main agent for the biological control of Diaphorina citri in Brazil with a parasitism rate ranging from 20 to 80%. This study investigated the influence of temperature on the development, fecundity and longevity of adults of T. radiata and the effect of relative humidity (RH) on their parasitism capacity and survival rate in the pre-imaginal period. The effect of temperature was assessed in the range between 15 and 35 ± 1ºC, 70 ± 10% RH, and a 14-h photophase. The RH effect was evaluated in the range from 30 to 90 ± 10%, temperature at 25 ± 1ºC, and photophase of 14-h. At 25ºC, circa 166.7 nymphs were parasitized, the highest parasitism capacity observed compared to other treatments. The longest longevity of females was observed at 25ºC, although the rate did not differ in the 20–30ºC temperature range. The threshold temperature (TT) was 7.2ºC, and 188.7 degrees-day were required for the development (egg-to-adult period). The parasitism rate and longevity were higher at 50 and 70% of RH. This shows that temperature and RH may affect the parasitism capacity of T. radiata on nymphs of D. citri, which can explain the great parasitism variation for D. citri observed in citrus groves in São Paulo State, Brazil.

Keywords: Biological control; capacity; fecundity; longevity; psyllid.



Diaphorina citri Kuwayama, 1907 (Hemiptera, Liviidae) is one of the most important pests for citrus producing areas in Brazil. It is the insect vector of the bacteria "Candidatus Liberibacter americanus" and "Candidatus Liberibacter asiaticus", that are associated to the main citrus disease in the world, the Huanglongbing (HLB) also known as Citrus Greening Disease (Martinez & Wallace 1967; Teixeira et al. 2005). Although D. citri is considered an exotic insect and it was first detected in Brazil at the end of the 1930's (Costa Lima 1942; Parra et al. 2010), only after the occurrence of HLB that its control started to be systematically carried out with chemical pesticides (Belasque Júnior et al. 2010). Chemical pesticides, therefore, became the main control management procedure for D. citri in citrus groves. The frequent use of pesticides has caused several problems. They kill natural enemies and other important insects for citriculture, compromising the Integrated Pest Management (IPM) in groves where the biological control was already implemented (Yamamoto & Parra 2005). Because D. citri is a widespread pest in Brazil and given its potential as a vector insect for HLB, it is feared that the Brazilian citrus industry be compromised, similarly to what has occurred in other countries (Fundecitrus 2004; Parra et al. 2010).

The search for alternative control methods of D. citri employs biocontrol agents such as the cenobiont ectoparasitoid Tamarixia radiata (Waterston, 1922) Hymenoptera, Eulophidae). This parasitoid has been widely used in several countries due to its high parasitism capacity, good establishment and field adaptation, constituting an important biological control agent of D. citri (Chien & Chu 1996; Hoy & Nguyen 2000; Étienne et al. 2001).

In Brazil, the occurrence of T. radiata was first registered in 2004 in the municipalities of Piracicaba and Jaboticabal, São Paulo State, immediately after the detection of HLB. In the period from March to July 2005, this parasitoid was registered in all citrus groves in São Paulo State, Brazil, showing natural parasitism rates between 27.5 to 80.0% (Gómez-Torres et al. 2006). This parasitism capacity has been greatly reduced due to the systematic use of chemical products for the control of D. citri (J.R.P. Parra, unpubl. data).

Complex hygrometric variables can affect the development, emergence and fecundity of this parasitoid, and temperature and RH are the most relevant (King et al. 1985). Thus, this study investigated the thermal hygrometric requirements of T. radiata for its production, mass rearing and establishment after releases in different sites as part of the IPM used for the psyllid control, vector insect for HLB.



Insect rearing. We used D. citri and T. radiata from the rearing of Laboratory of Insect Biology from the Department of Entomology and Acarology from Escola Superior de Agricultura "Luiz de Queiroz"– Universidade de São Paulo (ESALQ-USP). The rearing of D. citri was adapted from the methods described in Skelley & Hoy (2004) and Nava et al. (2007), using seedlings of the orange jasmine (Murraya paniculata (L.) Jack), 25–30 cm high, cultivated in a substrate of vermiculite and vegetal compound at 1:1 ratio, and maintained in incubators (30 ± 1ºC, 60 ± 10% RH, and a 14-h photophase). The plants with new sprouts were transferred to acrylic cages (34 x 34 x 40 cm) containing approximately 50 couples of D. citri to obtain eggs in a 24-h period. After oviposition, the plants were transferred to cages for nymph development (70 x 50 x 50 cm) and maintained in an incubators at 25 ± 1ºC, 70 ± 10% RH, and a 14-h photophase.

For the rearing of T. radiata, we used seedlings of the orange jasmine infested with nymphs in the 4th and 5th instars of D. citri (Chu & Chien 1991; Skelley & Hoy 2004; Nava et al. 2007). The seedlings of the orange jasmine containing nymphs in the 4th instar were placed for parasitism in cages (45 x 35 x 37 cm) for a 24-h period. Afterwards, the plantlets containing nymphs supposedly parasitized were placed in cages (60 x 50 x 52 cm) in an acclimatized room for T. radiata development. At emergence, 80% of the parasitoids were used for the experiments and the rest was used to maintain the rearing.

Effect of temperature on the parasitism capacity and survival rate of T. radiata. Twenty females of T. radiata, 24-h of age, mated, were placed individually in cylindrical plastic cages (15.5 x 5.5 cm), containing five holes at the top that were covered with voile fabric to allow aeration. The females were fed with a mixture of pure honey and pollen (1:1) (Chien et al. 1994), offered as fillets fixed to the inner sides of the cages. The nymphs of D. citri were exposed to T. radiata females for 24 h in incubators at 15, 20, 25, 30, and 35 ± 1ºC, 70 ± 10% RH, and a 14-h photophase.

For each female of T. radiata, we offered daily an orange jasmine plants with 30 nymphs of D. citri in the 5th instar, according to Chu & Chien (1991). After parasitism, the females were removed from the cages using a glass tube (12 x 75 mm). The nymphs of D. citri and the orange jasmine plantlet were placed in an acclimatized room kept under the same environmental conditions (temperatures, RH and photophase) during the parasitism period. We evaluated the number of nymphs parasitized daily, the accumulated parasitism rate, the total number of nymphs parasitized per female and longevity of females of T. radiata.

The experiment design used was a completely randomized with five treatments (temperatures) and four repetitions, containing four orange jasmine plantlets with 30 nymphs each.

Effect of temperature on the egg-to-adult period and thermal requirements. The orange jasmine plantlets with nymphs of D. citri in the 5th instar were offered to the females of T. radiata for a period of 24 h at 25 ± 1ºC, 70 ± 10% RH, and a 14-h photophase, in acrylic cages (15.5 x 5.5 cm) with holes on top covered with voile fabric to allow aeration. Afterwards, the females of T. radiata were removed from the cages and the orange jasmine plantlets with nymphs were placed in acclimatized rooms at 18, 20, 22, 25, 28, 30 and 32ºC, 70 ± 10% RH, and a 14-h photophase. Three hundred nymphs were observed at each temperature (treatments), they were divided into 10 repetitions (plantlets) with 30 nymphs per plant. Based on data from the egg-to-adult period, we calculated the threshold temperatures (TT), and the thermal constant (K) (Haddad et al. 1999).

Effect of RH on the parasitism capacity and development of the larval stages of T. radiata. The experiment was carried out in acclimatized rooms with regulated to 30, 50, 70 and 90 ± 10% HR, 25 ± 1ºC and, a 14-h photophase. Females of T. radiata, 24-h of age and mated, were placed individually in acrylic cages (35 x 35 x 45 cm) where they were fed with a droplet of pure honey and pollen (1:1) placed on the inner side of the cage.

An orange jasmine plantlet with 30 nymphs of D. citri in the5th instar was offered to each female of T. radiata. After 24 h, the females were removed from the cages using a glass tube (12 x 75 mm) and the nymphs were kept in acclimatized rooms under the environmental conditions described above.

We carried out daily observations to determine the rates parasitism and emergence of T. radiata. The parasitism rate was evaluated on the 5th day after offering the plantlets to the nymphs, when the parasitized nymphs had the mummy formation.

The experiment comprised a completely randomized design with five treatments (RH) and four repetitions, containing four plantlets with 30 nymphs each.

Data analysis. To determine the effect of temperature on the number of parasitized nymphs, the data were subjected to the analysis of variance (ANOVA) and the means were compared by the Tukey test (p < 0.05). Data on longevity were analyzed in the Kaplan-Meier estimator and later compared in the long-rank test using the R program (R Development Core Team 2011).

The temperature requirements were calculated in the hyperbole method (Haddad et al. 1999). To determine the effect of RH on rates of parasitism and emergence, the data were analyzed in the polynomial regression model. The analyses were preformed in the SAS program (Statistical Analysis System), version 9.2, 2002–2008 (SAS Institute 2002).



The total parasitism capacity of T. radiata greatly differed among the temperatures tested (F = 3.310; gl = 4; P = 0.039), and the highest parasitism capacity was observed at 25ºC followed by 30ºC. At 20, 35 and 15ºC, we observed a smaller number of nymphs parasitized, indicating that these temperatures are less adequate for the reproduction of T. radiata (Fig. 1). Regarding the daily parasitism, we observed that at 25ºC, the females parasitized a larger number of nymphs, followed by 30ºC (Fig. 2A).





In the other temperature conditions, the daily parasitism was lower, and at 15 and 35ºC, parasitism occurred on the 1st day of oviposition and did not surpass 10 parasitized nymphs (Fig. 2A). Furthermore, we observed that because these extreme temperatures (15 and 35ºC) negatively affected the life span of T. radiata, there was a shorter period of parasitism served at 20, 25 and 30ºC, respectively, which were mark- and 100% of parasitism was attained at these temperatures edly different from values found at 15 and 35ºC (Fig. 3). on the 6th day, while 100% of parasitism was attained between the 11th and 13th day at 20, 25 and 30ºC (Fig. 2B).



Longevity of females of T. radiata subjected to parasitism of D. citri was affected by the different temperatures (χ2 = 12.3; df = 4; P = 0.001). The longest life span was observed at 20, 25 and 30ºC, respectively, which were markedly different from values found at 15 and 35ºC (Fig. 3).

The duration of the egg-to-adult period of T. radiata was inversely proportional to the temperature rates studied, obtaining durations (± SEM) of 17.31 ± 0.13, 14.20 ± 0.12, 12.43 ± 0.13, 10.33 ± 0.13, 10.09 ± 0.11, 7.55 ± 0.19 and 7.59 ± 0.21 days for temperatures at 18, 20, 22, 25, 28, 30, 32ºC, respectively, significantly differing from one another (F = 2.761; df = 6; P = 0.025). Based on data on the egg-toadult period duration of T. radiata, we determined the temperature threshold of 7.2ºC and the thermal constant of 188.7 degrees-day (R2 = 0.9470) (Fig. 4).



RH affected rates of parasitism and emergence (F = 167.26; P = 0.0001; and F = 64.87; P = 0.0001, respectively) (Figs. 5A and 5B). The highest parasitism capacity was observed at RH 70 and 50%, corresponding to 83.0 and 65.5%, respectively. At RH 30 and 90%, the parasitism rate was 29.8 and 37.3%, respectively. The optimal relative humidity estimated for the parasitism capacity of T. radiata was 61.95% where the parasitism rate was 79.53% (Fig. 5A). The best emergence rate occurred at RH 50 and 71%, corresponding to 57.4 and 82.3%, respectively, while at RH 30 and 90%, we observed a survival rate of 37.8 and 26.5%, respectively (Fig. 5B). The highest estimated emergence rate (74.58%) occurred at RH 59.1%.




Temperature and RH influenced the reproductive activity of T. radiata and they can be determinant factors for the success of its use in biological control, for its mass rearing in laboratory, as well as its establishment in the citrus groves.

The parasitism capacity of T. radiata on D. citri is related to other factors, such as food and developmental stage (Chu & Chien 1991; Chien et al. 1994). In addition, temperature exerts great influence on the developmental and reproductive activities, as demonstrated by Gómez-Torres et al. (2012), where the highest parasitism rate (77.3%) occurred at 26.3ºC, similar to the rate found in our study at 25ºC, where each female of T. radiata parasitized on 166.8 nymphs (Fig. 1). At the other temperatures, higher or lower than 25ºC, the parasitism rate was reduced. According to Chu & Chien (1991), at 25ºC, fecundity of T. radiata may range from 98 and 156 eggs/females, similar to the findings in our study (166.75) under the same temperature condition.

It has also been observed a quadratic relation between fecundity of the parasitoid and temperature in Eulophidae, due to the significant interaction between this abiotic factor with the embryonary development and with life span of females (Patel & Schuster 1991). The authors also report that at low temperatures, the females laid fewer eggs when compared with females subjected to temperatures between 20 and 30ºC. Hondo et al. (2006) evaluated the development, efficiency and reproduction of seven species of Eulophidae based on temperature tolerance and concluded that all species analyzed were adapted to high temperatures (25 and 30ºC).

The highest daily and accumulated rates of parasitism observed at 25ºC are probably related to the average life span of the females of T. radiata, given that at extreme temperatures (15 and 35ºC), the average life span was reduced by half, i.e., three days shorter than at 25ºC (Fig. 3).

The development of the egg-to-adult period of T. radiata at different temperatures was similar to that observed by Fauvergue & Quilice (1991), in which the authors determined for the constant temperatures of 20, 25, 27 and 30ºC, a duration of 16.8, 10.1, 9.1 and 8.5 days, respectively. For temperature requirements for the egg-to-adult development, our obtained data differ from threshold temperature of 11ºC and the thermal constant 165GD, shown by Chien & Chu (1996) for a population of T. radiata in Citrus sinensis and M. paniculata. This difference of 3.8ºC for the thresholds temperature and 23.7 degrees-day between the two studies is probably attributed to the geographic origin of the populations, once temperature requirements between species may vary according to the geographic sites (Honék & Kocourek 1990; Nava et al. 2010).

In our study, we observed that the parasitism rate of T. radiata on D. citri was higher at RH 70%; however, it is reduced at 30 and 90%. Likewise, Duale (2005) found similar results for another species of Eulophidae Pediobius furvus (Gahan, 1928), with high parasitism rates between RH 60 and 80%. Probably, extreme RH influences indirectly the parasitoids, once they are protected by the nymphs' exoskeleton. Unpublished data show that for D. citri RH below 30% is harmful to nymphs, therefore, the pre-imaginal development of T. radiata may have also been affected (J.R.P. Parra, unpubl. data).

The relation between a parasitoid and its host is highly complex, thus, it is essential to understand the responses of biological control agents to biotic and abiotic factors that may affect its reproduction, development and survival (Geden 1997; Almeida et al. 2002). Therefore, based on the data observed in this study, criteria must be used to obtain enough mass rearing of T. radiata to enable the use of this parasitoid for the control of D. citri in citrus groves. These criteria should be associated to temperature and hygrometric requirements determined in the current study, i.e., temperature between 25 to 30ºC and RH near 70%. These parameters show great adequacy for the reproduction, development and survival of T. radiata. Moreover, the release and adaptation of T. radiata to field conditions will depend on thermal hygrometric requirements, which even allow occurrence delineation or identification of the number of generations for the regions in São Paulo State, Brazil's major citrus producer.



We are grateful to FAPESP for granting a doctoral fellowship to the first author (Proceeding: 05/59769–5), and to Thermal hygrometric requirements for the rearing and release of Tamarixia radiata Antonio Lourenço Guidoni (in memorian), for help with the statistical analyses. This study was funded by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP), Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), and the INCT-Semioquímicos na Agricultura.



Almeida, M.A.F., Prado, A.P. & Geden, C.J. 2002. Influence of temperature on development time and longevity of Tachinaephagus zealandicus (Hymenoptera: Encyrtidae), and effects of nutrition and emergence order on longevity. Environmental Entomology 31: 375–380.         [ Links ]

Belasque Júnior, J., Yamamoto, P.T., Miranda, M.P., Bassanezi, R.B., Ayres, A.J. & Bové, J.M. 2010. Controle do Huanglongbing no estado de São Paulo, Brasil. Citrus Research & Technology 31: 53–64.         [ Links ]

Chien, C.-C. & Chu, Y.-I. 1996. Biological control of citrus psyllid, Diaphorina citri in Taiwan. International Journal of Pest Management 34: 93–105.         [ Links ]

Chien, C.-C., Chu, Y.-I. & Ku, H.C. 1994. Influence of food on longevity, egg production and population increase of the eulophid wasp, Tamarixia radiata. FAO Plant Protection Bulletin 36: 97–105.         [ Links ]

Chu, Y.I. & Chien, C.C. 1991. Utilization of natural enemies to control psyllid vectors transmitting citrus greening, p.135–145. In: Kiritani, K., Su, H. J. & Chu, Y. I. (ed.). Integrated control of plant virus diseases. Proceedings of the International Workshop TARI, Taichung, Taiwan, April 9–14, 1990.         [ Links ]

Costa Lima, A.M. 1942. Insetos do Brasil. 3º. Tomo. Homópteros. Escola Nacional de Agronomia, Série Didática No. 4, 324 p.         [ Links ]

Duale, A.H. 2005. Effect of temperature and relative humidity on the biology of the stem borer parasitoid Pediobius furvus (Gahan) (Hymenoptera: Eulophidae) for the management of stem borers. Environmental Entomology 34: 1–5.         [ Links ]

Étienne, J., Quilici, S., Marival, D. & Franck, A. 2001. Biological control of Diaphorina citri (Hemiptera: Psyllidae) in Guadeloupe by imported Tamarixia radiata (Hymenoptera: Eulophidae). Fruits 56: 307–315.         [ Links ]

Fauvergue, X. & Quilici, S. 1991. Etude de certains paramètres de la biologie de Tamarixia radiata (Waterston, 1922) (Hymenoptera: Eulophidae), ectoparasitoide primaire de Diaphorina citri Kuwayama (Hemiptera: Psyllidae), vecteur asiatique du greening des agrumes. Fruits 46: 179–185.         [ Links ]

Fundecitrus. 2004. Greening chega às regiões Norte e Sul do Estado. Re-vista Fundecitrus 124: 8–11.         [ Links ]

Geden, C.J. 1997. Development models for the filth fly parasitoids Spalangia gemina, S. cameroni, and Muscidifurax raptor (Hymenoptera: Pteromalidae) under constant and variable temperatures. Biological Control 9: 185–192.         [ Links ]

Gómez-Torres, M.L., Nava, D.E. & Parra, J.R.P. 2012. Life table of Tamarixia radiata (Hymenoptera: Eulophidae) on Diaphorina citri (Hemiptera: Psyllidae) at different temperatures. Journal of Economic Entomology 105: 338–343.         [ Links ]

Gómez-Torres, M.L., Nava, D.E., Gravena, S., Costa, V.A. & Parra, J.R.P. 2006. Primeiro registro de Tamarixia radiata (Waterston) (Hymenoptera: Eulophidae) em Diaphorina citri Kuwayama (Hemiptera: Psyllidae) no Brasil. Revista de Agricultura 81: 112–117.         [ Links ]

Haddad, M.L., Parra, J.R.P. & Moraes, R.C.B. 1999. Métodos para estimar os limites térmicos inferior e superior de desenvolvimento de insetos. Piracicaba, FEALQ, 29 p.         [ Links ]

Hondo, T., Koike, A. & Sugimoto, T. 2006. Comparison of thermal tolerance of seven native species of parasitoids (Hymenoptera: Eulophidae) as biological control agents against Liriomyza trifolii (Diptera: Agromyzidae) in Japan. Japanese Journal of Applied Entomology and Zoology 41: 73–82.         [ Links ]

Honék, A. & Kocourek, F. 1990. Temperature and development time in insects: a general relationship between thermal constants. Zoologische Jahrbücher, Abteilung für Systematik, Ökologie und Geographie der Tiere 117: 401–439.         [ Links ]

Hoy, M.A. & Nguyen, R. 2000. Classical biological control of Asian citrus psylla. Citrus Industry 81: 48–50.         [ Links ]

King, E.G., Bull, D.L., Bouse, L.R. & Phillips, J.R. 1995. Introduction: biological control of Heliothis spp. in cotton by augmentative releases of Trichogramma. Southwestern Entomologist 8: 1–10.         [ Links ]

Martinez, A.L. & Wallace, J.M. 1967. Citrus leaf motlle-yellows disease in the Philippines and transmission of the causal virus by a psyllid, Diaphorina citri. The Plant Disease Reporter 58: 692–695.         [ Links ]

Nava, D.E., Gómez-Torres, M.L., Rodrigues, M.D.A., Bento, J.M.S. & Parra, J.R.P. 2007. Biology of Diaphorina citri (Hem., Psyllidae) on different hosts and at different temperatures. Journal of Applied Entomology 131: 709–715.         [ Links ]

Nava, D.E., Gómez-Torres, M.L., Rodrigues, M.D.A., Bento, J.M.S., Haddad, M.L. & Parra, J.R.P. 2010. The effects of host, geographic origin, and gender on the thermal requirements of Diaphorina citri (Hemiptera: Psyllidae). Environmental Entomology 39: 678–684.         [ Links ]

Parra, J.R.P., Lopes, J.R S., Gómez-Torres, M.L., Nava, D.E. & Paiva, P.E.B. 2010. Bioecologia do vetor Diaphorina citri e transmissão de bactérias associadas ao Huanglongbing. Citrus Research & Technology 31: 37–51.         [ Links ]

Patel, K.J. & Schuster, D.J. 1991. Temperature-dependent fecundity, longevity, and host-killing activity of Diglyphus intermedius (Hymenoptera: Eulophidae) on third ínstars of Liriomyza trifolii (Burgess) (Diptera: Agromyzidae). Environmental Entomology 20: 1195–1199.         [ Links ]

R Development Core Team. 2011. R: A language and environment for statistical computing. R Foundation for Statistical Computing. Available at: (accessed 11 July 2013).         [ Links ]

SAS Institute. 2002. SAS System – SAS/STAT computer program, version 9.2.         [ Links ]

Skelley, L.H. & Hoy, M.A. 2004. A synchronous rearing method for the Asian citrus psyllid and its parasitoids in quarantine. Biological Control 29: 14–23.         [ Links ]

Teixeira, D.C., Saillard, C., Eveillard, S., Danet, J.L., Costa, P.I., Ayres, A.J. & Bové, J. 2005. Candidatus Liberibacter americanus, associated with citrus huanglongbing (greening disease) in São Paulo State, Brazil. International Journal of Systematic and Evolutionary Microbiology 55: 1857–1862.         [ Links ]

Yamamoto, P.T. & Parra, J.R.P. 2005. Manejo integrado de pragas dos citros, p.729–768. In: Matos Jr., D., De Negri, J.D., Pio, R.M. & Pompeu Jr., J. (Org.). Citros. São Paulo, FAPESP.         [ Links ]



Received 19 December 2013; accepted 30 June 2014



Associate Editor: Adeney F. Bueno

Creative Commons License All the contents of this journal, except where otherwise noted, is licensed under a Creative Commons Attribution License