EFFECTIVENESS OF SOME NATIVE DIATOMACEOUS EARTH AGAINST MAIZE WEEVIL, SITOPHILUS ZEAMAIS (COLEOPTERA: CURCULIONIDAE), UNDER CONTROLLED CONDITIONS

all DEs treatments; however, emergence was very low than control treatment. It is concluded that the native DEs (Ankara and Aydın) are very effective and promising against S. zeamais adults and might be used for its successful control in storage maize.

Dusts, such as diatomaceous earth, are known insecticidal or repellant activity products and effectively controls most species of insect pests of storage products (Ebeling, 1971;Sousa et al., 2013). Diatomaceous earth affects larvae and adults, stick to insect's body parts as they move on their surface or inside the treated grain mass (Kavallieratos et al., 2007;Rojht et al., 2010;Shams et al., 2011;Sousa et al., 2013). When dust contacts with insects, it causes the removal of the layer of wax from insect cuticle, bringing about their death by dessication (Rojht et al., 2010). Besides, this product is extremely stable and has low toxic effect on mammalian, little potential for insect resistance as well as a good residual effect (Eldridge,1964;Ebeling,1971;Korunic,1998). Turkey has a huge and good quality diatomaceous earth reserve (125 million tons) and most of them geological deposits occur in some regions (especially Center Anatolia, Aegean region) of Turkey (Çetin and Taş, 2012). The regional deposits of DEs in Turkey, their low cost, easy availability and use warrant their usage as grain protectants. Although abundant and high-quality DEs reserves exist in Turkey, these have not extensively been evaluated for their potential utility in controlling storage products pest.
Many studies have been conducted in Turkey and other countries to determine the insecticidal, biological and repellent activity of native and trade DEs against storage product pests (Korunic, 1998;Masiiwa, 2004;Ferizli and Beris, 2005;Athanassiou et al., 2004;Sousa et al., 2013;Doumbia et al, 2014;Ertürk and Emekçi, 2014;Wakil et al., 2015;Alkan et al., 2019;Şen et al., 2019;Kılıç and Mutlu, 2020). Whereas, biological efficacy and dose studies of various DEs have been conducted such as on wheat, rice and corn related to S. oryzae and S. granarius species in Turkey, there is no literature available related to efficacy of local DEs for S. zeamais on storage corn kernels. In this study, some native diatomaceous earth from Aydın and Ankara provinces were evaluated for efficacy against S. zeamais in the laboratory-controlled conditions in 2020.

MATERIAL AND METHODS Diatomaceous Earth (DE) formulations
Three diatomaceous earth (DEs) (two natives, i.e., Ankara, Aydın and imported, i.e., SilicoSec ® ) were used in the study. The DE formulations were given in Table 1.

Test insects
Sitophilus zeamais adults were obtained from cultures maintained in the Entomology Laboratory of Plant Protection Central Research Institute Ankara, Turkey. The adults were reared in sterile 2-L transparent plastic jars on maize grains in incubators maintained at about 25±1°C and 65±5% relative humidity and 16:8 h (L: D) in Insectarium of Plant Protection Department, Agriculture Faculty, Harran University. Maize grains were kept in the deep freezer at -18 o C at least 7 days to prevent contamination. The jars were not filled to the brim for allowing free air circulation and respiration. Approximately, 250-300 mixed-sex adult individuals were introduced in the jars. The adults on the maize grains were vacuumed after 24 hours for obtaining 1-day-old new adults later. The newly emergence adults were observed at ~35-40 days after. Emerged adults were collected with an aspirator and used in experiments when they were 1 to 7 days old.

Laboratory bioassay
Bioassay studies were conducted according to randomized parcel design with four replicates at two temperatures (25 and 30±1ºC), 50-65% RH. Three doses of local (Ankara and Aydın) and SilicoSec ® DE formulation (1000, 1500 and 2000 ppm (mg DE/kg maize grain) were used. Untreated maize grains were considered as the control. For each dose, 100 g maize grains were used in 1-liter volume plastic jars. The jars were sealed and shaken manually for 1 min to distribute DE equally. Thirty S. zeamais adults were introduced in each jar and the insects were kept in controlled condition insectarium rooms at 25 and 30±1°C. The numbers of living and dead adults were recorded after 2, 3, 5, 7, 14 and 21 days respectively.

Effect of diatomaceous earth on F1 progeny
The dead and living adult insects were removed 21 days after the initiation of laboratory bioassay. The same jars (infested maize grains) were kept at 25 and 30±1°C and 65±5% relative humidity condition to allow oviposited eggs to hatch and develop. Then newly emerged adults were counted. Inhibition rate was calculated according to the formula below. IR = (Cn -Tn) / Cn * 100 IR% : Inhibition rate Cn : Number of newly emerged insects in the untreated (control) jar Tn : Number of newly emerged insects in the treated jar.

Statistical Analysis
All data were converted into percent mortality and subjected to arcsine scale followed by correction of cumulative mortality percentage for the corresponding control mortality (Abbott, 1925). Analysis of variance was conducted to assess the effect of concentration, time of exposure and their interaction with the insect mortalities. The differences among treatments were analyzed by means of Tukey multiple comparison tests (p<0.01). All statistical analyses were conducted with Jump (Version 7) package program.

RESULTS
The mortality varied within DEs and their doses. The highest mortality was recorded at higher DEs dose (2000 ppm/100g maize grains) at 25 and 30°C, while minimum mortality was obtained with lower dose. The lowest mortality after 2 nd day was 31.12, 36.09 and 13.33% with 2000 ppm dose of Ankara, Aydın and SilicoSec ® DEs, respectively at 25 °C (Table 2).  (Table 2). After 7th day, 2000 ppm dose of Ankara and Aydın DEs exhibited high mortality (96.60 and 93.25%), while mortality with the same dose of SilicoSec ® (77.50%) was lower. Insecticidal activity was >90% with 1000 ppm; however, 100% mortality was recorded with 2000 ppm dose of Ankara DE 14 DAT. However, SilicoSec ® did not reach 100% mortality with all concentrations. The mortality rate at 21 DAT was 100% with 1000 ppm doses of all DEs. The mortality with 1000, 1500 and 2000 ppm doses of all DEs at 30 °C was substantially greater than control at 3 DAT. A considerable increase in mortality was observed with increasing duration in control treatment but remained the lowest ( Table 2).
The mortality percentages of S. zeamais adults at 2, 3, 5, 7, 14 and 21-DAT at different concentrations of Ankara, Aydın and SilicoSec ® DEs at 30 °C are shown in Table 3. The highest mortality (97.3%) was recorded with 2000 ppm concentration of Ankara and Aydın DEs on 5 th and 7 th DAT, and reached 100% on 14 th DAT, while SilicoSec ® caused 58.67 and 78.44% mortality on 5 and 7 th DAT, respectively. The highest mortality (98.75%) for SilicoSec ® was noted 21 DAT and did not reach 100% with any application doses. Mortality in the untreated control remained constant after the insect exposure to untreated maize grains.
Different DEs and application doses showed different suppression effects on the progeny production of S. zeamais. After the long storage period (60 days), F1 adult emergence was recorded from all DEs and their all concentrations. The highest number of insects emerged from Aydın, Ankara and SilicoSec ® were 30, 26.5 and 15.5 adults respectively in the lower concentration treatment (1000 ppm) at 25 °C. The F1 adult numbers were decreased with increasing applications doses (1500 and 2000 ppm), which were 1, 2.5 and 7.8 adults for Ankara, Aydın and SilicoSec ® , respectively at 2000 ppm ( Figure 1).

DISCUSSION
The purpose of this study was to investigate the effect of two native diatomaceous earth (DEs) on mortality of S. zeamais adults under controlled conditions. The results of the study indicated that the efficacy of all DEs was considerably influenced by doses, time of exposure, temperature and type of DE formulations. Similar results have been reported by previous researchers (Korunic, 1998;Arthur, 2001;Arnaud et al., 2005;Vayias et al., 2006;Shams et al., 2011;Alkan et al, 2019;Şen et al., 2019;Kılıç and Mutlu, 2020). These studies indicated increased mortality of storage product pests exposed to inert dusts with increasing exposure time. Exposure time of two-day of DEs was not enough to obtain significant mortality with tested DEs in the current study. However, the mortality rate was increased at three DEs, as increased day of exposure after 5 th day and reached higher mortality rate on 14 th day at 25 and 30°C in the study. Similar result reported by Arthur (2001) who stated that mortality of Oryzaephilus surinamensis L. generally increased as exposure interval and temperature increased. Extended exposure time may be needed to increase mortality in adults, because more dust particles would be trapped by their bodies, losing more water and causing death by desiccation (Fields and Korunic, 2000;Arthur, 2001;Rigaux et al., 2001;Shams et al.2011).
The effectiveness of DEs increased with higher doses and temperature. This effect shows that the mortality of insects is dependent on the conditions in which the grains are exposed to DEs (Sousa et al., 2013). The particles and size of DEs have abrasive peculiarities but more importantly the ability to absorb lipid molecules from the cuticle of the insects and subsequently, in water loss, dehydration and death (Quarles and Winn, 1996;Korunic, 1998;Subramanyam and Roelsi, 2000). Altough the particles size of all DEs is very close, SilicoSec ® had moderate effect on S. zeamais adults. The results indicated that DEs from different locations vary in their efficacy against the maize weevil. This has also been explained by Korunic (1997) and Golob (1997) that DEs from different geological locations have different efficacies and DEs from marine areas are the most common but less efficacious.
The mortality was considerably increased with increasing temperature and was higher at 30ºC than 25ºC. The earlier studies on the influence of temperature on the efficacy of some local and traded DEs against storage pest (S. oryzae, T. confusum and R. dominica) indicated that increasing temperature resulted in increasing insecticidal efficiency against S. oryzae adults (Fields and Korunic, 2000;Arthur, 2002;Athanassiou et al., 2005;Vassilakos et al., 2006;Rojht et al., 2010Rojht et al., , Şen et al., 2019. The results of the current study agree with Vassilakos et al. (2006) who reported that insecticidal efficacy of SilisoSec ® against S. oryzae increased with increasing temperature. However, increased temperature would also increase feeding and therefore moisture replacement through the food and production of metabolic water. The synthesis of cuticular waxes may be faster at higher temperatures because of temperature effects on the biochemical pathways (Turnbull and Harris, 1986;Şen et al., 2019). High temperatures stimulate the movement of insects within the grain mass, allowing them greater contact with DEs. In addition, water loss is likely to be increased at higher temperatures (Arthur, 2001;Fields and Korunic, 2000).
The total mortality was influenced by DE types under both temperatures. The local DEs, Ankara and Aydın, showed >90% mortality 5 DAT, while SilicoSec ® caused ~60% mortality at 25°C. Smilar results were obtained at 30 °C which clearly indicated that the local DEs were effective than SilicoSec ® against the maize weevil. The differences between SilicoSec ® and local DEs can be explained by the differences in physical, morphological and chemical characters of DEs. Kılıç and Mutlu (2020) found the highest biological activity with local DE Aydın on Trogoderma granarium Everts larvae followed by Ankara DE, while SilicoSec ® had the lowest activity larvae at 30°C. However, Alkan et al., (2019) reported that the local DE Turco000 caused 100% mortality to Acanthoscelides obtectus Say. adults on chickpea at 1000 ppm 4 DAT and can be used as a valuable tool for bean weevil suppression in storage product pest management programs. Besides, complete mortality of T. confusum and S. oryzae can be achieved at lower concentrations ranging from 500 to 900 ppm local DE formulation (ACN-1) and this DE has potential to be used for control of storagegrain insects reported by Şen et al. 2019.
The study results showed that at least 1500 ppm dose at 30 °C caused >90% mortality 5 DAT. The results of the present study indicated that complete mortality of S. zeamais achieved at higher concentrations, ranging from 1500 to 2000 ppm. These results do not agree with Masiiwa (2004) who reported different results with local African DEs on the maize weevil. Insect mortality caused by DEs is attributed to the dehydration provoked by the abrasiveness of the small particles of this inert dust and by adsorption of oils in the body of the insect, which breaks the layer of wax on the epicuticle, exacerbating the fatal loss of water reported by Subramanyam and Roesli (2000). Therefore, at higher concentrations, the adsorption of wax and abrasiveness caused by the product occurs faster, causing death in a shorter time compared with low concentrations (Shams et al. 2011). In addition, the higher the concentration of DEs can be the more effective, because dust applied to cover containers and grain surface will be greater chances of the insect picking up the dust to cause enough damage (Masiiwa, 2004). Maize grain surface is rough, and adherence is greater than small grain like wheat, millet which are smooth surface have low adherence.
The number of emerged adults in progeny reduced with increasing temperature, reaching ~5% at 1500 and 2000 ppm doses of all DEs at 25 and 30°C. The high mortality was observed for 30°C 5 DAT. These results could be due to the fact that higher temperatures might reduce the rate of oviposition and limit the survival of insects and reduce the number of individuals in the progeny (Vardeman et al., 2006). Alkan et al. (2019) reported that there was no progeny production with local DEs, Turco000, Turco004, Turco020 at 200 ppm concentration at the end of 55-day incubation period on C. maculatus. Kılıç and Mutlu (2020) stated that 1000, 1500 and 2000 ppm of local DEs (Aydın and Ankara) applied to wheat resulted low mortalities of T. granarium larvae and higher doses did not prevent reproduction.
The results presented in this study suggest that 1500 ppm dose of Aydın DE with high temperatures can be recommended to control the maize weevil with 5 days exposure time.

CONCLUSIONS
The native diatomaceous earths obtained from local sources (Ankara and Aydın) showed high efficacy against the maize weevil under controlled conditions. The study also indicated that temperature had significant effect on the insecticidal efficacy of local DEs against the maize weevil, S. zeamais. Based on the results of the bioassays, mortality increased with increasing dose and the exposure time. In conclusion local DEs (Ankara and Aydın) have potential to be used for control of maize weevil.