Session 4: Evaluating chlorine dioxide and ozone as alternative methods for controlling phosphine-resistant insects in onfarm and commercial storages

biosecurity built on science

PB CRC 3038: Evaluating chlorine dioxide and ozone as alternative methods for controlling phosphine-resistant insects in on-farm and commercial storages

Bhadriraju Subramanyam, Xinyi EDepartment of Grain Science and Industry Kansas State University

Plant Biosecurity Cooperative Research Centre


What is the main message of this slide?

April, 2013: Signing of agreement. April-December, 2014: Preliminary laboratory experiments to

determine baseline data on chlorine dioxide and ozone dosages to control phosphine resistant insects.

February-June, 2015: Establishing baseline dosages of chlorine dioxide and ozone needed for controlling adults of phosphine resistant insects.

July, 2015-March, 2016: Full scale studies to establish realistic chlorine dioxide and ozone dosages.

April-August, 2016: Final report was prepared and submitted to PBCRC.

Milestones


What is the main message of this slide? Fumigant A fumigant is a chemical that can exist in the gaseous state in sufficient concentration to be lethal to a given target organism at a required temperature and pressure (Bond, 1989). It diffuses as individual molecules and can penetrate materials easily. Mists, fogs, and smokes are suspensions of liquids or solids in the air, and unable to penetrate the materials. Responsive tactic. Little or no residues. Uniform distribution (a challenge) in the treated commodities/spaces.

Rationale



Search for the new fumigants• Environmentally benign• High efficacy against all life stages of most species• No cross-resistance to existing fumigants• Effective suppression of progeny • Minimal impacts on grain quality• Cost-effective

Chlorine dioxide (ClO2) Water treatment: legionella, beverage and food industry, odor control, oil industry, municipal and waste water treatment. Surface and air treatment: food processing plants, biomedical devices, hospitality, residential.

Ozone (O3)Water treatment: cooling tower, bottling, hydroponics, aquaculture, drinking water. Soil remediation, odor control, industry laundries.

Rationale



Chlorine dioxide

Project summary


What is the main message of this slide?Species Strain County, state Collection year

Resistance status

Tribolium castaneum LAB Laboratory 1999 SusceptibleAB1 Dickinson, Kansas 2011 MediumPD Russell, Kansas 2011 MediumCF Washington, Kansas 2011 Weak

Rhyzopertha dominica LAB Laboratory 1999 SusceptibleCS Chase, Kansas 2011 WeakRL Riley, Kansas 2007 StrongFL Florida - Strong

Oryzaepilus surinamensis

LAB Laboratory 1999 Susceptible

AB2 Abilene, Kansas 2011 WeakSitophilus zeamais LAB Laboratory 1999 Susceptible

TX Texas 2011 WeakSitophilus oryzae LAB Laboratory 1999 Susceptible

TX Texas 2011 Weak



Post-exposure time (day)

1 2 3 4 50

20

40

60

80

100

2.02 g/m3, 6 h

Mean + S

E m

ortality (%)

0

20

40

60

80

1000.54 g/m3, 8 h

RFB-lab RFB-MN RFB-CF RFB-AB1 RFB-AB2

1.35 g/m3, 6 h

2.70 g/m3, 3 h

1 2 3 4 5

Mean + S

E m

ortality (%)

0

20

40

60

80

1000.54 g/m3, 8 h


1 2 3 4 50

20

40

60

80

1002.02 g/m3, 6 h

1 2 3 4 5

2.70 g/m3, 3 h

LGB-lab LGB-CF LGB-RL

1.35 g/m3, 6 h



Mean + S

E m

ortality (%)

0

20

40

60

80

1000.54 g/m3, 8 h


1 2 3 4 50

20

40

60

80

1002.02 g/m3, 2 h

1 2 3 4 5

1.35 g/m3, 6 h

2.70 g/m3, 3 h

STGB-lab STGB-PD STGB-AB2 STGB-CF

Mean + S

E m

ortality (%)

0

20

40

60

80

100RW-lab RW-TX MW-lab MW-TX

1.35 g/m3, 6 h 2.02 g/m3, 6 h


1 2 3 4 50

20

40

60

80

100

1 2 3 4 5

2.70 g/m3, 3 h 2.70 g/m3, 9 h


What is the main message of this slide?Species Strain

Exposure time (h) 24.8 °Cwith wheat without wheat

T. castaneum LAB 26 15AB1 26 15MN 26 15

O. surinamensis LAB 16 3AB2 16 3

R. dominica LAB 24 20CS 34 18RL 34 20

S. zeamais LAB 24 15TX 18 7

S. oryzae LAB 15 5TX 18 7

Exposure times required for complete mortality of adults of five stored-product insect species at 0.54 g/m3 (200 ppm) of chlorine dioxide based on mortality assessments made 5 d after the exposure.

Exposure times required for complete mortality of adults of five stored-product insect species at 1.40 g/m3 (520 ppm) of chlorine dioxide based on mortality assessments made 5 d after the exposure.

Species StrainExposure time (h) with wheat

24.8 °C 32.8 °C

T. castaneum LAB 12 8

AB1 12 8

CF 12 8

O. surinamensis LAB 4 4

AB2 4 4

R. dominica LAB 12 8

CS 12 8

RL 12 8

S. zeamais LAB 12 8

TX 12 8

S. oryzae LAB 12 8

TX 12 8


What is the main message of this slide?Progeny production (mean±SE) and percentage reduction (in the parenthesis) of laboratory and field strains of five insect species after exposure to 1.40 g/m3 of chlorine dioxide for various time periods in vials with wheat during October, 2015.

ExposureTime (h)

R. dominica S. zeamais S. oryzae

LAB CS RL LAB TX LAB TX

0 111.7±11.9 41.7±10.7 41±13.3 230±28.4 209±23.9 286.3±12.4 263.7±39.6

1 109.7±22.7(1.8%)

32.3±7.0(22.5%)

39±18.5(4.9%)

119.7±28(48.0%)

98.3±22.8(53.0%)

169.7±21.8(40.7%)

185.7±23.5(29.6%)

2 41.3±8.7(63.0%)

21.0±5.2(49.6%)

21.0±6.4(48.8%)

89.0±8.5(61.3%)

11.0±1.0(94.7%)

138.7±15.8(51.6%)

65.7±23.4(75.1%)

3 40.7±23.4(63.6%)

20.3±1.3(51.3%)

7.3±3.7(82.2%)

112.3±19.4(51.2%)

16.7±9.3(92%)

73.7±3.0(74.3%)

0.7±0.3(99.7%)

4 37.7±14.9(66.2%)

13.0±6.1(68.8%)

48.3±11.2(-17.8%)

4.0±3.5(98.3%)

8.3±5.8(96%)

25.0±16.8(91.3%)

2.0±0.6(99.2%)

5 11±2.9(90.2%)

7.7±1.9(81.5%)

15.0±14.5(63.4%)

27.3±12.5(88.1%)

1.0±0.6(99.5%)

_ _

6 19±6.4(83.0%)

4.7±2.3(88.7%)

12.7±4.5(69.0%)

12.3±10.3(94.7%)

0.7±0.7(99.7%)

_ _

7 13±6.4(88.4%)

5.0±2.1(88.0%)

8.0±4.6(80.5%)

2.7±1.8(98.8%)

24.3±24.3(88.4%)

_ _

8 3.3±2.0(97.0%)

2.0±2.0(95.2%)

11.7±9.7(71.5%)

1.3±0.9(99.4%)

0±0(100%)

_ _



• 800 ppm (2.16 g/m3) for 4 hours• 6 kg of wheat for each mini silo• Gas entered from the bottom and vented

from the top.• Insects were placed at the bottom, the

center and the top of the mini silo.• 25°C, 20 % r.h.

Species Strains% Mortality (Mean ± SE) at day 5

Bottom Center Top

T. castaneum

LAB 100 ± 0 100 ± 0 92.9 ± 3.6

CF 100 ± 0a 77.5 ± 1.9b 88.5 ± 4.1ab

MN 96.6 ± 1.7 91.2 ± 6.3 87.8 ± 6.1

R. dominica

LAB 100 ± 0 98.1 ± 1.9 94.3 ± 5.7

CS 100 ± 0 95.6 ± 4.4 97.8 ± 2.2

FL 96.7 ± 1.6 86.0 ± 6.3 95.2 ± 2.9

O. surinamensisLAB 100 ± 0 100 ± 0 100 ± 0

AB2 100 ± 0 100 ± 0 100 ± 0

S. zeamaisLAB 47.6 ± 6.5 36.2 ± 1.7 35.0 ± 1.5

TX 29.6 ± 3.7a 9.7 ± 2.3b 5.6 ± 5.6b

S. oryzaeLAB 91.7 ± 4.5 89.0 ± 1.6 78.2 ± 6.3

TX 37.9 ± 6.0 37.9 ± 6.0 30.2 ± 4.8

Means by species and strains followed by different letters was significantly different (P < 0.05) within a species and strain (by Bonferroni t-tests)


What is the main message of this slide?Strains

Column (% mortality at day 5) Box (% mortality at day 5)

50 g 250 g 500 g 250 g 500 g

RFB-LAB 100 ± 0a 66.7 ± 6.7b 13.7 ± 3.2c 85.0 ± 2.9ab 68.8 ± 8.5b

RFB-CF 100 ± 0a 67.4 ± 11.3b 26.6 ± 6.6c 100 ± 0a 100 ± 0a

LGB-LAB 100 ± 0a 23.6 ± 1.6b 0 ± 0b 98.1 ± 1.9a 73.2 ± 15.5aLGB-FL 100 ± 0a 9.8 ± 5.8c 6.7 ± 4.4c 81.2 ± 4.7a 42.0 ± 0.5b

STGB-LAB 100 ± 0a 100 ± 0a 70.0 ± 0b 100 ± 0a 100 ± 0a

STGB-AB2 100 ± 0a 100 ± 0a 72.3 ± 11.3b 100 ± 0a 100 ± 0a

RW-LAB 100 ± 0a 98.3 ± 1.7a 38.5 ± 11.3b 100 ± 0a 100 ± 0a

RW-TX 89.1 ± 7.5ab 60.0 ± 9.2b 59.9 ± 9.8b 96.6 ± 3.4a 96.7 ± 3.3a

Means by species and strains followed by different letters was significantly different (Bonferroni, P=0.05).


What is the main message of this slide?Ozone:

Ozone exposure time (h)0 5 10 15 20 25

Mean ± SE m

ortality (%)

0

20

40

60

80

100

0 5 10 15 20 25

A

C

B

D

0

20

40

60

80

100

LAB CS FL

A Mortality assessed on day 5 for R. dominica exposed with and without wheat to 0.21 or 0.42 g/m3 of ozone.

(A) Without wheat and 0.21 g/m3 of ozone;

(B) Without wheat and 0.42 g/m3 of ozone;

(C) With wheat and 0.21 g/m3 of ozone;

(D) With wheat and 0.42 g/m3 of ozone.


What is the main message of this slide?Lethal times (LT99) of phosphine susceptible and resistant strains of four insect species

after exposed to 0.42 g/m3 of ozone in the presence or absence of wheat

Species Strains Lethal time (95% CL) (h) Ratio (95% CL)LT99 with wheat LT99 without wheat

T. castaneum

LAB 13.64 (8.64-92.79) 10.65 (8.31-19.90) 1.26 (0.58-2.73)

CF 15.91 (11.79-24.91) 14.35 (11.37-20.30) 1.11 (0.71-1.73)

PD 25.81 (16.19-64.99) 29.89 (21.74-47.59) 1.16 (0.57-2.36)

O. surinamensis

LAB 7.07 (4.24-25.46) 4.33 (3.49-5.98) 1.63 (0.82-3.25)

AB2 11.18 (9.33-14.16) 9.05 (7.64-11.25) 1.24 (0.94-1.63)

S. zeamais

LAB 5.56 (4.65-7.20) 4.20 (3.56-5.29) 1.32 (1.00-1.75)

TX 3.80 (3.37-4.50) 3.14 (2.52-4.59) 1.21 (0.89-1.64)

S. oryzae

LAB 3.11 (2.76-3.65) 2.32 (2.02-2.87) 1.34 (1.08-1.65)

TX 2.00 (1.90-2.13) 2.91 (2.07-8.17) 1.45 (0.96-2.20)


What is the main message of this slide?Progeny production (% reduction) of laboratory and field strains of four insect species after exposure to 0.42 g/m3 of ozone for different time periods in the presence of wheatTime (h) T. castaneum O. surinamensis S. zeamais

S. oryzae

LAB CF PD LAB AB2 LAB TX LAB TX

0 115.8 14.2 41.0 260.6 355.8 278.4 160.4 223.0 190.8

1 70.8(38.9%)

16.4(-15.5%)

57.0(-39.0%)

242.4(7.0%)

229.4(35.5%)

215.4(22.6%)

93.2(41.9%)

102.4(54.1%)

98.2(48.5%)

2 58.0 (49.9%)

13.8(2.8%)

41.4(-1.0%)

49.4(81.0%)

149.8(57.9%)

125.6(54.9%)

56.6(64.7%)

4.4(98.0%)

1.8(99.1%)

3 45.0(61.1%)

46.4(-226.8%)

35.8(12.7%)

69.4(73.4%)

92.0(74.1%)

103.0(63.0%)

4.0(97.5%)

1.4(99.4%)

0.6(99.7%)

4(5) 65.2(43.7%)

54.4(-283.1%)

66.0(-61.0%)

2.4(99.1%)

3.8(98.9%)

4.8(98.3%)

3.8(97.6%)

2.0(99.1%)

1.2(99.4%)

6 5.6(95.2%)

10.6(25.4%)

60.6(-47.8%)

2.4(99.1%)

1.8(99.5%)

0(100%)

1.2(99.3%)

1.4(99.4%)

4.2(97.8%)

8 8.4(92.7%)

29.2(-105.6%)

46.4(-13.2%)

2.6(99.0%)

0.4(99.9%)

1.0(99.6%)

0.2(99.9%)

3.8(98.3%)

0.4(99.8%)

10 3.6(96.9%)

19.8(-39.4%)

40.6(1.0%)

1.2(99.5%)

0(100%)

- - - -


What is the main message of this slide?Progeny production (% reduction) of laboratory and field strains of four insect species after exposure to 0.42 g/m3 of ozone for different time periods in the absence of wheat

Time (h) T. castaneum O. surinamensis S. zeamais

S. oryzae

LAB CF PD LAB AB2 LAB TX LAB TX0 47.6 11.4 39.8 339.6 194.8 285.8 179.6 162.8 114.6

1 73.6(-54.6%)

16.2(-42.1%)

65.0(-63.3%)

282.2(16.9%)

271.6(-39.4%)

47.0(83.6%)

7.8(95.7%)

65.4(59.8%)

11.8(89.7%)

2 46.6(2.1%)

24.4(-114%)

89.2(-124.1%)

138.2(59.3%)

129.6(33.5%)

49.6(82.6%)

0.8(99.6%)

0(100%)

0(100%)

3 46.2(2.9%)

69.6(-510.5%)

92.4(-132.2%)

29.0(91.5%)

42.6(78.1%)

15.4(94.6%)

0(100%)

0(100%)

0(100%)

4(5) 45.8(3.8%)

55.6(-387.7%)

95.6(-140.2%)

1.0(99.7%)

0.6(99.7%)

0(100%)

0.4(99.8%)

0(100%)

0(100%)

6 11.2(76.5%)

46.8(-310.5%)

90.8(-128.1%)

0.6(99.8%)

0.8(99.6%)

0.2(99.9%)

0.4(99.8%)

0(100%)

0(100%)

8 6.0(87.4%)

12.6(-10.5%)

86.0(-116.1%)

1.6(99.5%)

0.4(99.8%)

0(100%)

0.4(99.8%)

0(100%)

0(100%)

10 2.6(94.5%)

0.6(94.7%)

49.4(-24.1%)

0.4(99.9%)

0.4(99.8%)

- - 0(100%)

0(100%)

Percent reduction (mean ± SE) in adult emergence from four young stages of Rhyzopertha dominica after exposure to 0.42 g/m3 of ozone for various time periods

Stages Exposure time (h)

Percent reduction (mean ± SE) in adult emergence after ozone exposure

LAB CS FL

Eggs 24 73.3 ± 3.9 64.4 ± 3.6 47.7 ± 12.4

48 85.4 ± 3.9 85.3 ± 4.5 92.4 ± 4.8

72 99.7 ± 0.3 96.3 ± 2.0 100.0 ± 0

Young larvae 10 100.0 ± 0 100.0 ± 0 99.0 ± 1.0

24 100.0 ± 0 99.2 ± 0.5 97.1 ± 2.0

34 99.7 ± 0.3 98.4 ± 0.7 99.0 ± 1.0

Old larvae 10 53.3 ± 3.9 b 64.6 ± 6.7 b 93.3 ± 4.3 a

24 99.1 ± 0.4 98.8 ± 1.2 98.3 ± 1.0

34 99.6 ± 0.3 98.8 ± 0.5 99.2 ± 0.8

Pupae 2 12.4 ± 8.9 12.1 ± 11.6 19.6 ± 17.7

6 16.4 ± 13.8 b 35.2 ± 6.0 b 76.5 ± 7.0 a

10 32.8 ± 8.6 b 54.4 ± 6.9 b 96.6 ± 3.4 aMeans by species and strains followed by different letters was significantly different (Bonferroni, P=0.05).


What is the main message of this slide?Significant findings Both ozone and chlorine dioxide can effectively kill adults of phosphine susceptible and resistant strains

of five common insect pests after exposed to 200-500 ppm of gas for less than 24 hours. They can also effectively suppress the progeny production of Rhyzopertha dominica, and Sitophilus spp.

Young stages of Rhyzopertha dominica are susceptible to ozone exposure (200 ppm), especially for larvae.

Mortality of adults of all strains and species increased as the post-exposure time increased, indicating delayed toxicity effect.

Both gases have the ability to react with the active sites on the surface of the grain, and longer equilibrium time is expected to reach to the targeted lethal concentration.

There is no cross-resistance between ozone and phosphine or chlorine dioxide and phosphine. High temperatures promote the efficacy of chlorine dioxide against adults.

Wheat restricts the penetration of chlorine dioxide and ozone by surface binding reaction and porosity, resulting lower mortality.


What is the main message of this slide?Research Outcomes• Subramanyam, Bh., E, X.Y., Savoldelli, S., Sehgal, B. , Maier, D. E. , Ren, YL (2014) Efficacy of ozone against

stored grain insect species in wheat: Laboratory and field observations. 11th International Working Conference on Stored Product Protection.

• Subramanyam, Bh., E, X.Y. (2015) Efficacy of chlorine dioxide gas against five stored-product insect species. 10th Conference of International Organization for Biological and Integrated Control (IOBC)-Integrated Protection of Stored Products.

• Subramanyam, Bh., E, X.Y., Li, B. (2016) Efficacy of a high concentration of chlorine dioxide gas against laboratory and field strains of five stored product insect species. XXV International Congress of Entomology.

• E, X.Y., Li, B., Subramanyam, Bh. (2016) Efficacy of ozone fumigation against phosphine susceptible and resistant strains of Rhyzopertha dominica. XXV International Congress of Entomology.


What is the main message of this slide?Industry Impacts PureLine is planning to register chlorine dioxide as a fumigant for stored products in U.S.

A broader spectrum will be investigated with PureLine’s financial support including the deactivation of microbes and denaturation of mycotoxins in stored grains.

Popcorn county


What is the main message of this slide?Future Plans