Lethal effect of sulfuryl fluoride on eggs of different age of the Indian meal moth Plodia interpunctella (Hübner) – demonstration of the no constancy of the ct product for control

Letaleffekte von Sulfurylfluorid auf Eier unterschiedlichen Alters der Dörrobstmotte Plodia interpunctella (Hübner) – Nachweis der Nichtkonstanz des ct-Produkts für die Bekämpfung

For one day exposure and the tested SF concentrations, the lethal effect increased with increasing concentration and age, with the exception of three day old eggs and concentrations above 4.19 g/m³, when they were more tolerant than younger and older eggs. After exposure to 1 g/m³ of SF, the one, two, and three day old eggs were with 2% mortality more tolerant than the four day old eggs with 18% mortality. On increasing the SF concentration towards 4.19 g/m³ and 5.24 g/m³, one day old eggs possessed the highest tolerance of all egg stages at this exposure period. At 5.24 g/m³ and 6.24 g/m³, three day old eggs were more tolerant than one day old eggs. Meanwhile, four day old eggs showed the highest susceptibility at all applied concentrations. Only after exposures of one day, percent mortalities of two and three day old eggs ranged between those of one and four day old eggs.

After two days exposure, the percent mortalities increased clearly and reached even 100% at the two highest concentrations for all eggs of ages of three and four days. The corresponding ct products of 299.5 gh/m³ and 251.5 gh/m³, respectively, are given in [ ] in the line below (see numbers below the according 100% values in Table 1). One day old eggs were always least susceptible.

After increasing the exposure period to three days, even 1 g/m³ was enough to kill 80 – 90% of all eggs of all four ages with one and two day old eggs being most tolerant. The highest tested concentration of 6.24 g/m³ obtained full control of all egg stages tested within three days of exposure [ctp₁₀₀: 449.3 gh/m³]. This data point with 100% mortality is based on investigations of four age groups with 50 eggs each and three replications amounting to 600 eggs in total. Four day old eggs required the highest tested dose.

The full lethal effect occurred already at 1.99 g/m³ and four days of exposure with a ct product of 191 gh/m³ for altogether 450 tested eggs. Four days of exposure were not carried out with four day old eggs, since hatch of larvae from the untreated eggs occurred already during this period. Therefore, these results are based on 450 eggs instead of 600 eggs. Two day old eggs were most tolerant at concentrations of 1 g/m³ and 1.48 g/m³, respectively.

Fig. 1 presents all mortality results together in one graph without indicating the standard deviations. Generally during the various experiments, the increase of mortality followed very clearly for all egg ages and tested exposure periods the increase in concentration of SF. Four days exposure resulted already at fairly low con­centrations of 1.99 g/m³ in 100% mortality of all eggs of all stages [ct = 191 gh/m³], whereas 6.24 g/m³ achieved the same degree of mortality of all tested eggs of all ages within three days of fumigation at 27°C [ct = 449.3 gh/m³]. One and two day old eggs survived two days of exposure to 6.24 g/m³, the highest concentration tested, whereas eggs of all ages survived this treatment when they were exposed for one day only.

Fig. 1. Average mortality of eggs of P. interpunctella of different age (1d – 4d) after exposure to different concentrations of sulfuryl fluoride for different times of fumigation: 1 day exposure (1dex) to 4 days exposure (4dex); 27°C and 65% r.h., three replicates.

As shown in Table 1, the standard deviations of the three replicates of the mortality results remained nearly always below 5%; only in very few series of experiments the SD increased up to 8.4%. SD of the mortalities in the untreated reference samples varied between 0% and 1.6%. This low mortality in untreated samples can be expected from reported studies with eggs of this moth (Reichmuth, 1979; Reichmuth et al., 1999, 2003).

As example, Fig. 2 demonstrates for three day old eggs the magnitude of the SD in relation of the various aver­ages of the results. For one day exposure, the typical sigmoid function described the resulting mortality data for the increasing concentration. Four days of exposure at 1 g/m³ [ct = 96 gh/m³] or three days at 5.24 g/m³ [ct = 377.3 gh/m³] were sufficient for complete control of eggs of this age. This degree of control of these eggs was also achieved with 6.24 g/m³ [ct = 299.5 gh/m³] within two days of fumigation.

Fig. 2. Average mortality of three day old eggs of P. interpunctella after one to four days of exposure (1dex – 4dex) to different concentrations of sulfuryl fluoride; 27°C and 65% r.h., three replicates.

Fig. 3 reveals the increase of efficacy when the exposure period was expanded for up to four days. The increase of mortality from three days of treatment to four days – following the typical sigmoid shape – was not tremendous. Two day old eggs could tolerate best the four day treatment. Duplication of the concentration from 1 g/m³ to 1.99 g/m³ [ct = 191 gh/m³] led to full control of eggs of all ages as shown in Fig. 4 (see also Table 1).

Fig. 3. Efficacy of 1 g/m³ of sulfuryl fluoride towards eggs of different age (one day to four days) of P. interpunctella at 27°C and 65% r.h., three replicates per average point with 50 eggs each.

Fig. 4. Mortality of eggs of P. interpunctella of different age, 1 day (1d) to 4 days (4d) after exposure to 1.99 g/m³ of sulfuryl fluoride at 27°C.

As can be expected from Fig. 3, the prolongation of the exposure to 1 g/m³ beyond four days may lead to full control within few more days than four. This new concept of SF fumigation would on one side save plenty of fumigant and help to reduce emission of the gas during and after the treatment and lead to less formation of residues in treated goods. On the other side, prolongation of the exposure time can be costly in many situations (e.g. mill fumigation, ship fumigation).

At the tested temperature of 27°C, effective fumigation against P. interpunctella can obviously be carried out at fairly low concentrations within fairly short treatment periods. One day exposure would theoretically require about 13 g/m³ for 95% control [ctp₉₅ = 312 gh/m³] or about 16 g/m³ [ctp₁₀₀ = 384 gh/m³] for full control, if the corresponding mortality data were extrapolated for instance with formula 8076 of the TableCurve program (r² = 9656; a = 110.9866, b = 4.4049, c = –1.7145). The fiducial limits of this regression were fairly high and indicated the necessity to determine these data more precisely by further fumigation experiments. Even if – for practical reasons – the concentration during a fumigation in the field would be increased by factor two to three, as proposed by Reichmuth and Klementz (2008), full control of eggs of this species could be obtained with ct products below 1,500 gh/m³, as registered in many countries.

The nonlinear regression of all corresponding average results for 1.99 g/m³ with TableCurve (Equation 8076, logistic dose response model, r² = 0.9418), shown as line in Fig. 4, evaluated 3.89 days as the necessary exposure time for full control [ct = 185.8 gh/m³], confirming the result in Table 1 with four days being sufficient for 100% mortality of all tested eggs [ct = 191 gh/m³].

Table 2 and Fig. 5 try to highlight the dependency of the ct product for control from the age of the treated eggs and the exposure time. An obvious shift in susceptibility of eggs of different age from one day to three days of exposure leads finally to nearly the same mortality of one, two and three day old eggs, respectively, after four days of exposure. The explanation could be the change in membrane permeability for SF of the egg shells with age and during the exposure (Outram, 1967a, b) and the development of hardly susceptible eggs into fairly susceptible young larvae within the eggs during the exposure period. During four days of exposure, also very young eggs at the beginning of the treatment have presumably and obviously developed into more susceptible eggs.

Fig. 5. ct product as function of the exposure period for control of 95% of the eggs of different age of P. interpunctella with sulfuryl fluoride at 27°C, data see Table 2.

Since the source of some data in Fig. 6 were those concentrations and exposure times in Table 1 where full control of all exposed 150 eggs was achieved, the theoretical lethal dosage may be a bit lower due to the step width of concentration and exposure time. On the other hand, 150 tested eggs as total cannot lead to mortality data with much better than 1% precision as pointed out by Reichmuth (2007). In this case of ctp₁₀₀ – other than with ctp₉₅ –, nonlinear regression for determination of missing data for 100% mortality for short exposure periods was not applied due to very large fiducial limits, indicating the low precision of the calculated data. Still it was obvious that expectedly like ctp₉₅ also ctp₁₀₀ dropped at longer exposures than three days. For one and two days of exposure, ctp₁₀₀ data for were not produced and are missing in the Fig. 6. Table 2 contains these as extrapo­lated figures.

Fig. 6. ct products as function of the exposure period; data represent ctp values for control of the most tolerant eggs at each exposure time for control of P. interpunctella with sulfuryl fluoride at 27°C; data see Table 2; polynomial nonlinear regression of the dependency.

Fig. 6 combines the results of Table 1 and 2 in selecting only those ct product values that were required to control the most tolerant eggs of the tested ages at each exposure time. The formula ctp₉₅ = 967 – 197 t – 395/t (r² = 0.99998987, for t = 1 to 4 days) describes the decline of the ct product with extended exposure beyond two days. In so far, it is demonstrated that the ct product for achieving 95% or 100% control is not constant for various exposure periods in the range of one to four days. Longer exposure required less SF for control than proportional would expected if the ct value would have been a constant.

As shown in Fig. 7, the ct product varied also as a function of the selected concentration and was not constant. Especially short exposure times required high concentrations for control.

Fig. 7. ctp₁₀₀ as function of the concentration of sulfuryl fluoride during one to four days of fumigation (1dex to 4dex) of eggs of different age (1d to 4d) of P. interpunctella at 27°C; at each exposure period, the larger symbols represent the older eggs; bold symbols derived directly from Table 1, some data (not bold symbols) determined with nonlinear regression with TableCurve (Equation. 8095).

The dependency of ctp₁₀₀ on the concentration of SF contains Fig. 7 with those data directly taken from Table 2. Non bold symbols are based on data calculated by non linear regression.

Fig. 8 describes the exposure time as function of the concentration of SF during a fumigation of eggs of different age of P. interpunctella at 27°C leading to 100% mortality of 150 eggs each at all data points. Values of ctp₁₀₀ for eggs of different ages are indicated with different symbols. Data that were taken directly from Table 1 are presented in bold symbols; the other calculated data are indicated as hollow symbols. The lines connect those data at each exposure time that required the highest concentration to be fully effective against the respective most tolerant age group of eggs. These boundary concentrations and corresponding values of the ctp₁₀₀ are pre­sented as figures in the frames. Exposure periods and concentrations with coordinates under and left of the lines would not be sufficient for full control of all eggs of all age groups. Appropriate values with coordinates right of and beyond these lines would lead to full control. A fairly simple polynomial regression y = a + bx + cx² with the regression coefficient r² = 0.9985 describes this borderline: y = lethal exposure time in hours; x = lethal concentration of SF in g/m³; a = 8.85, b = 0.07975 and c = –0.0015798611 are the constants of the equation.

Fig. 8. Exposure time and concentration of sulfuryl fluoride to obtain full control of eggs of P. interpunctella at 27°C; bold symbols directly taken from Table 1; data for other symbols by non­linear regression; square: 1 day old eggs, triangle: 2 day old eggs, circle: 3 day old eggs; diamond: 4 day old eggs; lines indicate necessary conditions for full control of eggs of all ages: Multiplication of exposure time and the indicated figures results in the ctp₁₀₀ for these conditions.

According to the presented data, fumigation for one day or four days seems to require the smallest ct products with short exposure of one day needing the higher concentration (9.8 g/m³) and longer exposure of four days only 1/5 (1.99 g/m³) of this high concentration. According to the nonlinear regression for ctp₁₀₀ at two days of exposure, this time or three days of exposure would require nearly the same ct product of about 450 gh/m³. Of course, the concentration for only two days of effective treatment had to be 1.5 times higher than for three days exposure.

The general question was, as to whether the ct product of SF remains a constant number over a range of concentrations and exposure periods or not. As found in this study in the contrary (see Table 1 and Table 2), the ct product for control changed for eggs of different age at each exposure period. The rank of the magnitude of the mortality varied between exposure periods and concentrations (see Table 3; data for the ranking taken from Table 1). In the four blocks with six concentrations and four different exposure periods, it can be seen that the ranks varied clearly, with most often rank 1 for egg age one day and rank 4 for egg age four days. The ranking was slightly distorted by some occasions when the same mortality occurred at two different data points. In this case the same figure was given several times. The lower line in Table 3 presents the sum of ranks for all ages and exposure times. It indicates the stated trend for most tolerant eggs of one day and least tolerant eggs of four days with some exceptions. In so far, it can be expected that also the ct products are not constant.

The lethal ct product of the most tolerant egg age clearly increased from one day to two or three days of exposure and decreased again strongly from two and three days of exposure towards four days of treatment (Fig. 8). The mortality data for one day exposure and the concentrations tested remained far below 100%. So, only extrapolated data with very high fiducial limits could be used for this type of consideration. Over the whole investigated and extrapolated range of concentrations and exposure periods, the lethal ct product varied and fluctuated between 96 gh/m³ and 450 gh/m³, depending on exposure time, concentration and egg age (Table 2). It can be concluded that at 27°C a ct product of 450 gh/m³ should be sufficient for control of all eggs of this moth in fumigations in the laboratory. All these data confirm the conclusion that the ct product to control eggs of P. interpunctella with SF at 27°C is not constant but strongly dependent on concentration, exposure period and age of the eggs.

Studies on various insect species showed that the egg is generally the most tolerant stage (TEAP report, 2011). Ciesla and Ducom (2009, 2010) in their studies confirmed the study of Reichmuth et al. (2003), that the dosage for an efficacious SF fumigation can be reduced “intelligently” for instance with increasing the temperature in the treated object or extending the treatment time. Even double treatment with low concentration may be an option to overcome the high tolerance of insect eggs towards treatment with SF. Karakoyun and Emekci (2010) showed in their results that the susceptibility of eggs of Carpophilus hemipterus to SF at 25°C varied according to age. One day old eggs of C. hemipterus seemed more tolerant than one to two day old eggs at all the temperatures tested. At 25°C, Baltaci et al. (2009) found that a complete mortality of one to four day old eggs of E. elutella was obtained after one day exposure at 21.3 g/m³ [ct = 511.2 gh/m³]. Extrapolated from the results after one day exposure of this study at 27°C, a ct product of only 235.92 gh/m³ (ctp₁₀₀) would have been sufficient for this degree of control for eggs of all age groups of P. interpunctella. In the presented study, a ct product of 449.3 gh/m³ achieved complete mortality of eggs of all ages of P. interpunctella at a concentration of SF of 6.24 g/m³ after three days of exposure at 27°C.

Bell et al. (1999) investigated the response of eggs of the Mediterranean flour moth Ephestia kuehniella to fumigation with SF. They observed strong dependency on the temperature and also on the age of the treated eggs. The necessary ct products for control even at 25°C were much higher than those determined for eggs of E. elutella (Baltaci et al., 2009) or P. interpunctella in this study. For effective treatment of eggs of a closely related pest moth, the rice moth Corcyra cephalonica, Barakat et al. (2009) needed 6.24 g/m³ (5.24 g/m³ and 4.19 g/m³, respectively) at 27°C within three (four and five) days of exposure [ctp₁₀₀: 449.28 gh/m³ (3d), 503.04 gh/m³ (4d) and 502.8 gh/m³ (5d)]. Compared to this study with P. interpunctella, the ctp₁₀₀ of 449.3 gh/m³ for three days of exposure was equal. Four days of exposure to only 1.48 g/m³ [ctp₁₀₀ = 142.1 gh/m³] and 1.99 g/m³ [ctp₁₀₀ = 191 gh/m³] were sufficient to control one and two day old eggs of P. interpunctella. Three day old eggs of C. cephalonica were not tested by Barakat et al. at this exposure period. In this study, P. interpuctella eggs of that age died already at ctp₁₀₀ = 96 gh/m³.

Walse (cited in TEAP, 2011) presented results for treatment over one day of eggs of P. interpunctella for control with SF at different temperatures. At 26.7°C, 559 gh/m³ achieved 99% mortality of several thousands of eggs in the test.

Several field and laboratory studies with SF included all developing stages of P. interpunctella in their groups of tested insects (Drinkall et al., 2003 [semolina mill, 22 h 15 min, 1101 gh/m³ – 1501 gh/m³, 28°C – 31°C], Ducom et al., 2003 [three laboratory studies, 24 h, 1125 – 1877 gh/m³, 25°C], Reichmuth et al., 2003 [three mill fumigations, 36 h to 48 h, 1860 – 2055 gh/m³, 22°C – 40°C]) and observed 100% control in all samples with this species including eggs. Some overdosing for this species can be assumed, since also more tolerant species like the flour beetles Tribolium spec. had to be controlled in the tests.

In general, it can be concluded that the relationship between the ct product obtained with SF and the mortality of eggs of P. interpunctella is complex. The influencing factors include the levels of concentration, the period of exposure and the age of the eggs. Therefore the ct principle cannot be used as the only guide for practical fumigation work also due to very specific conditions of the fumigations. Always either the concentration or the exposure period must be mentioned together with the respective ct product to obtain a full picture.

The results of this study support the presumption that for fumigating eggs of pest insects with SF, short exposures at relatively high concentrations are not as effective as long exposures at relatively low concentrations. At a given temperature and exposure periods between one and four days, the lethal ct product is not constant but varies considerably with the tendency to decline for longer exposures than two days.

In so far, the concept of the constancy of the product for control of eggs of insects as valid for methyl bromide does not hold for SF. This gas is for many applications a promising alternative for methyl bromide but the determination of the appropriate dosage for control is more complex and needs more consideration.