Journal of Stored Products Research 47 (2011) 306e310
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Journal of Stored Products Research
journal homepage: www.elsevier.com/locate/jspr
Susceptibility of stored product insects to high concentrations of ozone
at different exposure intervals
Marissa X. McDonough a, Linda J. Mason a, Charles P. Woloshuk b, *
a
b
Department of Entomology, Purdue University, 901 West State Street, West Lafayette, IN 47907, USA
Department of Botany and Plant Pathology, Purdue University, 915 West State Street, West Lafayette, IN 47907, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Accepted 14 April 2011
Ozone is a highly reactive gas with insecticidal activity. Past studies have indicated that ozone technology
has potential as a management tool to control insect pests in bulk grain storage facilities. The objective of
this study was to determine the efficacy of short periods of exposure to high ozone concentrations to kill
all life stages of red flour beetle (Tribolium castaneum (Herbst)) (Coleoptera: Tenebrionidae), and Indianmeal moth (Plodia interpunctella (Hübner)) (Lepidoptera: Pyralidae), adult maize weevil (Sitophilus
zeamais (Motsch.)) (Coleoptera: Curculionidae) and adult rice weevil (S. oryzae (L)) (Coleoptera: Curculionidae). Insects were treated with six ozone concentrations between 50 and 1800 ppm. The specific
objective was to determine minimal time needed to attain 100% mortality. The most ozone-tolerant
stages of T. castaneum were pupae and eggs, which required a treatment of 180 min at 1800 ppm
ozone to reach 100% mortality. Eggs of P. interpunctella also required 180 min at 1800 ppm ozone to reach
100% mortality. Ozone treatments of 1800 ppm for 120 min and 1800 ppm for 60 min were required to
kill all adult S. zeamais and adult S. oryzae, respectively. The results indicate that high ozone concentrations reduce the treatment times significantly over previously described results. Our results also
provide new baseline information about insect tolerance to ozone treatment.
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Ozone
Sitophilus zeamais
Tribolium castaneum
Sitophilus oryzae
Plodia interpunctella
1. Introduction
Insect infestation within stored product facilities is a major
concern to the grain industry. Losses due to insect damage and
mycotoxin contamination can exceed $500 million a year (Harein
and Meronuck, 1995). With limited control options and the
increased potential for resistance to insecticides (Zettler et al., 1989;
Zettler and Cuperusi, 1990; Benhalima et al., 2004), additional
methods are needed for management of insects in stored gains. One
candidate is ozone, which has proven effective against storage
insect pests (Erdman, 1980; Strait, 1998; Kells et al., 2001; Leesch,
2003; Zhanggui et al., 2003; Işikber and Öztekin, 2009). Although
the mechanism of action of ozone on insects is not completely
known, the insect’s respiratory system is a likely target (Tiwari
et al., 2010).
Ozone is highly reactive and a strong oxidizing agent, and is
classified as “GRAS” (Generally Recognized As Safe) by the United
* Corresponding author. Tel.: þ1 765 494 3450; fax: þ1 765 494 0363.
E-mail addresses: mfusco@purdue.edu (M.X. McDonough), lmason@purdue.edu
(L.J. Mason), woloshuk@purdue.edu (C.P. Woloshuk).
0022-474X/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.jspr.2011.04.003
States Environmental Protection Agency (US-EPA). Throughout the
world, ozone has been used to purify drinking water, kill bacteria,
sanitize food, deodorize, and decrease aflatoxin contamination
(Prudente and King, 2002; Sopher et al., 2002; Inan et al., 2007;
Tiwari et al., 2010; White et al., 2010). One advantage of ozone is
that it breaks down into atmospheric oxygen, eliminating the need
to store or dispose of hazardous chemicals.
Several studies have established that ozone treatments can kill
stored grain insects, including maize weevil, (Sitophilus zeamais
(Motsch), rice weevil (S. oryzae (L), red flour beetle (Tribolium castaneum (Herbst), confused flour beetle (T. confusum (Jacqueline du
Val), Indianmeal moth (Plodia interpunctella (Hübner), and Mediterranean flour moth (Ephestia kuehniella (Zeller) (Strait, 1998; Kells
et al., 2001; Leesch, 2003; Zhanggui et al., 2003; Işikber and
Öztekin, 2009). In each of these previous studies, the ozone
concentrations and the exposure times were limited. The present
study was undertaken to establish a better understanding about
the relationship between these two parameters on the mortality of
stored grain insects. The objective was to measure insect mortality
over a range of ozone concentrations and exposure to determine
conditions for achieving 100% mortality. The results provide the
baseline values needed to develop on-farm applications.
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M.X. McDonough et al. / Journal of Stored Products Research 47 (2011) 306e310
2. Materials and methods
2.1. Insects
All insects used in this study were obtained from colonies at
Purdue University, West Lafayette, IN. Colonies were maintained at
30 C 0.5 C and 60% rh.
Plodia interpunctella colonies were maintained on a diet made of
1000 g turkey mash (PurinaÒ chow animal feed), 72.4 g brewer’s
yeast, 150 mL water, and 150 mL honey. Adults (<48 h old) were
removed from the colony after gassing the rearing jar with CO2 and
placed in Petri dishes (100 mm 15 mm) to recover. Eggs were
obtained by placing adult moths in glass mason jars (w1000 mL,
inverted with a wire mesh lid into a plastic cup) and allowed to lay
eggs. Eggs were sorted and placed singly into 96-well plates (flat
bottom). After treatment, the plates were covered with ParafilmÒ
(Chicago, IL, USA). Larvae of different ages were collected from the
colony and placed into cylinders (10 larvae each) made by cutting
off the bottom of 50 mL CorningÒ centrifuge tubes. The ends of the
cylinders were covered with nylon mesh. Pupae were produced on
sections of cardboard roll placed in the colony jars. Ten pupae were
placed in a Petri dish. Petri dishes were covered with a modified lid
containing a hole (w6 cm diameter) with mesh fabric to allow for
gas exchange.
Tribolium castaneum were maintained on rearing media
(approximately 90% flour, 10% brewer’s yeast by weight). Adults,
larvae and pupae were collected from the colony on a #25 sieve
(710 mm USA standard ASTM E-11, Seedboro Equipment Co. Chicago, IL., USA). Ten insects of each life stage were placed in Petri
dishes. To obtain eggs of T. castaneum, adults were placed on
rearing media, and after 24 h, the adults were removed with a #25
sieve. The eggs were collected on a # 80 sieve (180 mm USA standard ASTM E-11; Seedboro Equipment Co.) and 10 eggs were placed
in each Petri dish.
Colonies of S. zeamais and Sitophilusoryzae were maintained on
whole kernels of corn and wheat, respectively. Adults were
collected on a #6 sieve (3.35 mm, USA standard ASTM E-11, Seedboro Equipment Co.) from the colony, and 10 insects were placed
into a Petri dish.
were scored in the same time period and manner as the corresponding ozone treatment. All treatments were performed in
ambient conditions (20 5 C).
Movement of adults and larvae was evaluated immediately after
ozone treatment. Then insects were placed in an environmental
chamber (Percival Scientific, Perry, IA, USA) (30 0.5 C and 60% rh)
for 24 h, and percent mortality was determined. Pupae and eggs
were placed in the environmental chamber after treatment and
examined daily for emergence. The experiment was terminated
when all insects of the control (no ozone) treatment had either
emerged or were determined to be dead. Once all control insects
emerged, the ozone-treated insects were scored for mortality and
discarded.
2.3. Data analysis
The data were analyzed based on mortality of the specific stage
of insect tested. A modified Abbott’s formula was used to determine
the corrected mortality. The modification was used to account for
control mortality (Rosenheim and Hoy, 1989; taken from
Subramanyam and Hagstrum (1996):
“Pcorr ¼ 1
ð1
TÞð1 KÞ
ð1 CÞ
and
K ¼
VarðCÞt2
ð1
CÞ2 nc
Var(C) is the variance associated with C, nc is the number of
replicates used for estimating C, t is the value of the t distribution at
nc 1 degrees of freedom (a ¼ 0.05).” T is the treatment mortality
(Subramanyam and Hagstrum, 1996). We calculated and compared
the mean concentration X time products (ppm-min) for 100%
mortality of all life stages between P. interpunctella and
T. castaneum. We also compared differences between 100%
mortality for life stages within species for P. interpunctella and
T. castaneum. Differences between 100% mortality of S. zeamais and
S. oryzae were determined. All data were analyzed for differences in
treatment mortality (a ¼ 0.05) using analysis of variance (ANOVA)
and Fisher’s least significant difference (LSD) test using general
linear model statistics (PROC GLM, SAS Institute, 2001).
3. Results
2.2. Experiments
Air used for ozone generation was dried with a column of
anhydrous calcium sulfate (DRIERITE, Xenia, OH) prior to
being passed through an ozone generator provided by O3Co
(Aberdeen, ID, USA). A flow-through system was used to expose
insects to various concentrations of ozone. Ozone from the
generator was passed through three identical plastic boxes
(101.6 mm 101.6 mm 25.4 mm) connected in series by TygonÒ
tubing (12.7 mm 2.38 mm diameter). An ozone analyzer (INUSA
(Needham, MA, USA), L2-LC Model 040977) was connected to the
outlet of the box at the end of the series. In the first experiment,
treatments consisted of concentrations ranging from 50 to
1800 ppm ozone (50, 100, 500, 1000, 1500, and 1800) with two
treatment times (30 and 60 min). A pseudo-replicate consisted of
10 insects enclosed in a Petri dish, cylinder or multi-well plate.
Three pseudo-replicates were performed for each treatment
combination, and each treatment combination was replicated three
times. For all experiments, new sets of insects were used. If 100%
mortality was not achieved within the 60 min exposure at the
maximum ozone concentration (1800 ppm), a second set of
experiments was conducted in which the insects were exposed to
1800 ppm ozone until 100% mortality was reached. Additional
exposure times were increased in 30 min intervals. Control (no
ozone) treatments were tested with adults, eggs, and larvae and
In our first set of experiments, the objective was to determine
the level of mortality from treatments of 30 and 60 min at various
ozone concentrations. Table 1 summarizes the df, P-values and
F-values for all treatments. Mortality of the control (no ozone) was
0e1% for the adult insects, 21e30% for eggs, 7e13% for larvae, and
18e21% for pupae. These data were used in the Abbott’s formula to
determine the corrected mortality values for the various treatments. In general, all concentrations of ozone resulted in increased
insect mortality (Tables 2e4). Table 2 shows treatment mortality
(%) SE for the two treatment times (30 and 60 min) for all life
stages of P. interpunctella. Only the adult stage of P. interpunctella
were all killed within the original 1800 ppm and 60 min exposure
time (Table 2). Treatment mortality (%) SE for the two treatment
times (30 and 60 min) for all life stages T. castaneum is depicted in
Table 3. One-hundred percent morality was not reached for either
the 30 or 60 min treatment times. All S. oryzae adults were killed
after a treatment of 60 min at 1800 ppm (Table 4). Mortality of
S. zeamais increased during the treatments, but 100% was never
achieved for the 30 and 60 min exposure times.
In the second set of experiments, the objective was to determine
the exposure time needed to achieve 100% mortality at 1800 ppm
ozone. All life stages were killed within a 180 min exposure time.
Table 5 summarizes the results of the additional time intervals
above 60 min. Larvae of P. interpunctella and T. castaneum required
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M.X. McDonough et al. / Journal of Stored Products Research 47 (2011) 306e310
Table 1
ANOVA values for the 30 and 60 min treatment times.
Species
Life Stage Treatment Time (min) df
Plodia interpunctella
Egg
Larvae
Pupae
Adults
Tribolium castaneum Eggs
Larvae
Pupae
Adults
Sitophilus zeamais
Adults
Sitophilus oryzae
Adults
30
60
30
60
30
60
30
60
30
60
30
60
30
60
30
60
30
60
30
60
F value P-Value
53
1.5
53
9.1
53 39.4
53
6.8
53
6.4
53 17.4
26 89.1
26 67.8
53
7.3
53 15.7
53
8.3
53 56.2
53
0.9
53
7.6
53 37.8
53 35.7
53 17.8
53 81.6
53 110.2
53 192.1
0.2221
<0.0001
<0.0001
<0.0001
0.0001
<0.0001
<0.0001
<0.0001
<0.0001
<0.0001
<0.0001
<0.0001
0.4671
<0.0001
<0.0001
<0.0001
<0.0001
<0.0001
<0.0001
<0.0001
df, P-value, and F-values reflect mean treatment mortalities for the different
concentrations of ozone used for the 30 and 60 min treatments.
the least amount of additional time, reaching 100% mortality at
90 min (Table 5). Sitophilus oryzae was not included in this second
study because 100% mortality was reached after a treatment of
60 min at 1800 ppm. For 100% mortality of S. zeamais, 120 min at
1800 ppm was required. The eggs of P. interpunctella and
T. castaneum as well as T. castaneum pupae were most resistant to
ozone, requiring 180 min exposure. The larvae of the two species
were equally sensitive, both requiring only 90 min exposure to
reach 100% mortality (Table 5).
Based on the experimental results presented in Tables 2e5, the
concentration X time products (ppm-min) (CT value) necessary for
100% insect mortality were calculated (Table 6). Differences were
observed in the susceptibility of all life stages of P. interpunctella
(df ¼ 35, F value ¼ infinity, P ¼ < 0.0001). There were no differences
in the susceptibility of eggs and pupae of T. castaneum as compared
Table 2
Percent mortality of all life stages of Plodia interpunctella treated with various
concentrations of ozone for 30 and 60 min.
Life Stage
Ozone Concentration (ppm)
Eggs
50
100
500
1000
1500
1800
50
100
500
1000
1500
1800
50
100
500
1000
1500
1800
50
100
500
Larvae
Pupae
Adults
Treatment Mortality (%) SE*
30 min
60 min
50.7 11.6 a,b
21.9 11.3 b
16.9 14.1 a,b
55.2 8.4 a
29.4 14.1 a,b
26.5 4.8 a,b
11.6 5.8 b,c
4.4 5.5c
1.4 7.1 b
14.3 6.4 b,c
90.0 7.1 a
84.5 4.7 a
15.9 7.3 c
46.5 9.6 b,c
56.5 11.5 b
69.0 13.1 a,b
57.2 10.9 b,c
95.8 2.9 a
4.4 2.9 b
15.5 6.6 b
90.0 4.4 a
24.9 12.5 b,c
12.7 7.9 b
36.5 6.9 b,c
24.7 11.6 b
88.5 4.4 a
66.9 11.7 b,c
6.31 4.2 d
9.7 5.9 c,d
47.9 13.4 a,b
37.7 12.1 b,c
64.1 9.6 a,b
66.9 11.7 a
16.8 8.4 b
24.9 10.6 b
74.9 11.9 a
89.9 2.9 a
89.2 5.9 a
92.9 3.7 a
15.5 5.5 b
22.5 8.2 b
100 0 a
*Different letters in each column and within life stage indicate significant mortality
differences between concentrations (P < 0.05, LSD test).
Table 3
Percent mortality of all life stages of Tribolium castaneum treated with various
concentrations of ozone for 30 and 60 min.
Life Stage
Ozone Concentration (ppm)
Treatment Mortality (%) SE*
30 min
60 min
Eggs
50
100
500
1000
1500
1800
50
100
500
1000
1500
1800
50
100
500
1000
1500
1800
50
100
500
1000
1500
1800
53.2 9.5 d
28.82 11.9 c,d
45.4 8.4 b,c
55.7 7.4 a,b
80.7 11.4 a
52.1 5.9 b,c
27.7 7.6 b
18.9 3.5 b
27.7 6.6 b
67.8 4.9 a
56.6 14.7 a
74.8 7.3 a
66.6 9.6 a
79.1 6.3 a
73.6 4.8 a
67.8 5.4 a
83.7 8.4 a
76.3 8.4 a
6.7 2.8 b
1.1 1.1 b
7.8 2.8 b
14.4 5.6 b
7.7 2.7 b
80.4 3.1 a
40.5 7.0 b
7.2 5.9 c
39.2 14.8 b
74.3 8.9 a
95.7 2.1 a
88.3 7.7 a
28.8 5.8 d
22.2 4.3 d
62.2 3.2 c
76.6 5.2 b
92.2 2.2 a
92.1 2 a
66.1 6.9 b,c
79.8 4.1 b
61.6 6.8 c
95.9 2.8 a
95.9 2.0 a
79.2 7.4 b
0.0 0.0 d
3.3 2.3 d
12.2 4.3 d
28.8 6.3 c
48.8 9.9 b
84.3 3.6 a
Larvae
Pupae
Adults
*Different letters in each column and within life stage indicate significant mortality
differences between concentrations (P < 0.05, LSD test).
to the other life stages (df ¼ 71, F value ¼ 85.59, P ¼ < 0.0001).
When comparing T. castaneum and P. interpunctella, the susceptibility to ozone is significant depending on insect and life stage
(df ¼ 35, F value ¼ infinity, P ¼ < 0.0001). Longer mean exposure
times were required to kill 100% of the eggs followed by pupae,
larvae, and adults (324,000; 270,000; 162,000; 123,000 ppm-min,
respectively). Eggs of P. interpunctella and eggs and pupae of
T. castaneum were the least susceptible to ozone, requiring a CT
value of 324,000 ppm-min (180 min at 1800 ppm) to reached 100%
mortality. Plodia interpunctella pupae, T. castaneum and S. zeamais
adults had a similar response to the treatments, with their required
CT value for 100% mortality at 216,000 ppm-min (120 min at
1800 ppm). Sitophilus oryzae required a smaller CT product
(108,000 ppm-min) (60 min at 1800) compared to S. zeamais
(216,000 ppm-min)to achieve 100% mortality (df ¼ 17,
F value ¼ infinity, P ¼ < 0.0001). The most susceptible insect and
Table 4
Percent mortality of Sitophilus oryzae and S. zeamais adults treated with various
concentrations of ozone for 30 and 60 min.
Species
Ozone
Concentration (ppm)
Treatment Mortality (%) SE*
30 min
60 min
Sitophilus oryzae
50
100
500
1000
1500
1800
50
100
500
1000
1500
1800
10.0 1.0 d
10.0 1.0 d
47.0 6.0 c
73.0 3.0 b
80.0 5.0 a,b
90.0 3.0 a
8.9 5.6 d
14.4 5.3 c,d
33.8 6.2 c
45.0 6.4 b
53.3 5.8 b
72.2 4.0 a
3.0
3.0
77.0
80.0
98.0
100.0
1.1
12.2
63.8
86.3
70.0
93.3
Sitophilus zeamais
0.5 c
2.0 c
2.0 b
5.0 b
0.05 a
0.0 a
1.1 c
3.6 c
5.6 b
6.2 a
5.3 b
2.3 a
*Different letters in each column and within species indicate significant mortality
differences between concentrations (P < 0.05, LSD test).
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M.X. McDonough et al. / Journal of Stored Products Research 47 (2011) 306e310
Table 5
Percent mortality of Plodia interpunctella, Tribolium castaneum, and Sitophilus zeamais treated with 1800 ppm ozone.
Species
ANOVA
Life Stage Treatment Treatment
Time (min) Mortality (%) SE*
Plodia
Eggs
interpunctella
Tribolium
castaneum
S. zeamais
30
26.5 4.8 b
Larvae
60
90
120
150
180
30
66.9 11.7 b
84.6 4.6 a
95.1 3.5 a
98.4 1.6 a
100.0 0.0 a
84.5 4.7 a,b
Pupae
60
90
30
66.9 11.7 b
100.0 0.0 a
95.8 2.9 a
Eggs
60
90
120
30
92.9 3.7 a
98.6 1.4 a
100.0 0.0 a
52.1 5.9 d
Larvae
60
90
120
150
180
30
88.2 7.7 a
94.5 3.4 a
97.0 2.0 a
97.1 1.9 a
100.0 0.0 a
74.8 7.3 b
Pupae
60
90
30
92.1 2.0 a
100.0 0.0 a
76.3 8.4 b
Adults
60
90
120
150
180
30
79.3 7.4 b
65.9 10.7 b
98.1 1.9 a
94.5 1.5 a
100.0 0.0 a
80.4 3.1 b
Adults
60
90
120
30
84.3 3.6 b
96.0 4.1 a,b
100.0 0.0 a
72.2 4.0 b
60
90
120
93.3 2.4 a
97.7 1.5 a
100.0 0.0 a
df ¼ 53, F ¼ 33.22,
P ¼ < 0.0001
Table 6
Calculated CT values to achieve 100% mortality of Plodia interpunctella, Tribolium
castaneum, Sitophilus oryzae and S. zeamais.
Species
Life stage
Treatment
(min @ 1800 ppm)
CT product
(ppm-min)
Plodia interpunctella
Eggs
Larvae
Pupae
Adults
Eggs
Larvae
Pupae
Adults
Adults
Adults
180
90
120
60
180
90
180
120
60
120
324,000 a
162,000 c
216,000 b
30,000 d
324,000 a
162,000 c
324,000 a
216,000 b
108,000 b
216,000 a
Tribolium castaneum
df ¼ 26, F ¼ 4.98,
P ¼ 0.0156
df ¼ 35, F ¼ 5.00,
P ¼ 0.2150
df ¼ 53, F ¼ 18.73,
P ¼ < 0.0001
df ¼ 26, F ¼ 8.61,
P ¼ 0.0015
df ¼ 53, F ¼ 5.00,
P ¼ 0.0009
df ¼ 35, F ¼ 2.92,
P ¼ 0.0491
df ¼ 35, F ¼ 27.20,
P ¼ <0.0001
*Different letters in each column and within species indicate significant mortality
differences between concentrations (P < 0.05, LSD test).
life stage tested were adult P. interpunctella, needing a CT of
30,000 ppm-min (60 min at 500 ppm).
4. Discussion
In this study, we examined the effects of high ozone concentrations on four insect species belonging to three taxonomic families. The results indicated that 100% of the P. interpunctella adults
were killed within 60 min when exposed to 500 ppm ozone. The
same concentration and exposure time resulted in only 12%
mortality in T. castaneum. Işikber and Öztekin (2009) observed
similar differences between two other species from these same
insect families. Ephestia kuehniella (Zeller) and T. confusum adults,
which belong to the Pyralidae and Tenebrionidae, respectively,
were treated with a continuous ozone flow of 13.9 mg/L (6482 ppm)
for 2 h. Mortality for E. kuehniella was 100% compared with less than
10% for T. confusum. Leesch (2003) also noted this higher sensitivity
of moths to ozone than beetles. Similarly, our data suggest that
differences in ozone sensitivity may exist at the species level. While
100% mortality was observed for S. oryzae after 60 min exposure to
Sitophilus oryzae
Sitophilus zeamais
*Different letters within life stage for P. interpunctella and T. castaneum indicate
significant mortality differences (P < 0.05, LSD test). Differences for S. oryzae and
S. zeamais are between the two species (P < 0.05, LSD test).
1800 ppm ozone, which was similar to T. castaneum, only 93%
mortality was observed for S. zeamais under the same conditions,
and 100% mortality was not observed until 120 min exposure.
We also determined the treatment conditions required to obtain
100% mortality of the other life stages of P. interpunctella and
T. castaneum. In general, longer exposure times at 1800 ppm were
required for eggs and pupae to reach 100% mortality as compared to
larvae. Işikber and Öztekin (2009) also observed higher sensitivity
for larvae and pupae of E. kuehniella and T. confusum compared to
eggs. If ozone insecticidal activity involves the insect’s respiratory
system (Tiwari et al., 2010), the increased tolerance of eggs and
pupae to ozone may be due to lower respiration rates in these life
stages (Hoback and Stanley, 2001). It also is possible that the outer
layer on the eggs and pupae provide some additional barrier to the
ozone. Furthermore, these factors may be responsible for some of
the inconsistencies in overall trends in the mortality data for eggs
and pupae. The metabolic status and integrity of the outer layer
may vary, resulting in some individuals being more susceptible to
ozone than others.
The successful application of ozone technology requires a sufficient concentration of ozone for an appropriate amount of time. In
water treatment systems, models for the inactivation of microbes
with ozone have been developed with CT values, which are the
products of the ozone concentrations multiplied by the exposure
times needed to attain a desired outcome (Clark et al., 2002). There
are currently no CT models for ozone treatment of insects; however,
we calculated the CT values for ozone treatments required to achieve 100% mortality (Table 6). Based on the CT products for 100%
mortality derived from our experimental results, T. castaneum
requires longer exposure or higher ozone concentrations than
P. interpunctella (a mean exposure of 256,500 ppm-min and
183,000 ppm-min, respectively). We used these values to compare
our results with those previously published, which used different
ozone concentrations and treatment times.
Sousa et al. (2008) conducted laboratory experiments with
T. castaneum adults, treating these insects in plastic chambers
with 150 ppm ozone to achieved 95% mortality with a treatment
time between 23.35 h and 31.98 h (CT ¼ 210,150 ppm-min and
287,820 ppm-min, respectively). Even with the low ozone
concentrations and long treatment time, their CT values are
similar to our CT value of 216,000 ppm-min for 100% mortality of
T. castaneum adults. Kells et al. (2001) treated caged T. castaneum
adults buried in corn with 50 ppm ozone. After a 3-d treatment,
they obtained 92% mortality for a CT value of 216,000 ppm-min,
which also agrees with our findings. Zhanggui et al. (2003) achieved 82% mortality after treating T. castaneum adults for 28 h at
120 ppm (CT ¼ 201,600 ppm-min), and 100% mortality was
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M.X. McDonough et al. / Journal of Stored Products Research 47 (2011) 306e310
achieved after 50 h (CT ¼ 360,000 ppm-min). Leesch (2003)
treated T. confusum adults in a chamber with 300 ppm ozone.
One-hundred percent mortality was observed after an 18-h
treatment. This treatment equates to a CT value of 324,000 ppmmin, which is considerably higher (w33%) than the CT value
calculated from our data and comparable to the values of
Zhanggui et al. (2003) and the upper range of values recorded by
Sousa et al. (2008). The difference may be due to variation in
species sensitivity to ozone. Leesch (2003) also treated the insects
for 5 h and observed only 37% mortality (CT ¼ 90,000 ppm-min).
No results were reported for treatments between 5 and 18 h. It is
possible that 100% mortality occurred earlier in the treatment,
between 5 and 18 h.
For the treatment of moths, our results with P. interpunctella
differ substantially with those of Işikber and Öztekin (2009)
investigating E. kuehniella adults. They observed 100% mortality
of E. kuehniella after a treatment of 13.9 mg/L ozone for 2 h, which
calculates to a CT value of 777,840 ppm-min. We observed 100%
mortality of P. interpunctella at a CT value of 30,000, about 26 times
less exposure to ozone than Işikber and Öztekin (2009). Further
research is needed to determine if E. kuehniella is more sensitive to
ozone than P. interpunctella.
Our results indicated that S. oryzae adults are more sensitive to
high concentrations of ozone and would be easier to control than to
S. zeamais. At the same ozone concentration (1800 ppm), half the
exposure time (60 min) was required to achieve 100% mortality of
S. zeamais (120 min). Kells et al. (2001) achieved 100% mortality of
adult S. zeamais with a CT ¼ 216,000 ppm-min, similar to that found
in our study. Furthermore, de Sousa et al. (2006) achieved 100%
mortality of S. zeamais with a 48-h treatment time at 50 ppm ozone,
and Zhanggui et al. (2003) reported similar results with a treatment
of 28 h at 120 ppm. The CT value for the results of de Sousa et al.
(2006) is 144,000 ppm-min and 201,600 ppm-min for Zhanggui
et al. (2003), which differed from ours results by 1.5 and 1.1
times, respectively.
The effect of ozone differs with the life stage of the insect. Our
analysis suggests that, in general, longer mean exposure to ozone
was required for eggs to reach 100% mortality followed by pupae,
larvae, and adults with CT values of 324,000; 270,000; 162,000;
123,000 ppm-min, respectively. Kells et al. (2001) treated
P. interpunctella larvae with 50 ppm ozone. After a 3d treatment,
they obtained 95% mortality for a CT value of 216,000 ppm-min.
This value is 1.3 times the CT value we obtained (162,000 ppm-min)
for this insect.
Mendez et al. (2003) found that a 30-d treatment of a variety of
grains at 50 ppm did not affect the chemical properties of the grains
or their processing characteristics. A 30-d ozone treatment at
50 ppm has a CT value of 2,160,000, which, based on our data, is
well above the highest exposure (324,000 ppm-min) necessary to
attain 100% mortality. These results indicate that ozone treatments
required to reach 100% mortality for all life stages are below the
exposure concentrations and times that would impact the properties of grains that are important to end users. Although the results
indicate that increasing the ozone concentration reduced the time
needed to achieve 100% mortality, generating these levels in
a storage facility will be a major challenge. Alternatively, using
ozone in conjunction with modified atmospheres may increase
effectiveness. Leesch (2003) suggested that carbon dioxide may
force insects to open their spiracles, thereby increasing effective
contact with the ozone gas. More field research is necessary to test
this combination with CO2 and possibly other gases to determine
efficacy.
Acknowledgments
Support for this research was provided by USDA/NIFA, award
number 2005-51101-02358. We thank O3Co (Aberdeen, ID) for
supplying the generator and for technical assistance.
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Charles Woloshuk