DOI: 10.5061/dryad.sxksn03fk
title: Enhanced insecticidal activity of isoparaffin by ozone as an adjuvant

description: Recently, concerns have been raised regarding control methods for
 domesticated sanitary pests. For many years, we have relied on chemical
 pesticides for control, but their negative effects on the human body and
 increased insecticide resistance have been reported, highlighting the
 necessity to develop new control technologies. Therefore, we focused on
 insect respiratory systems, for which it is considered difficult to
 develop insecticide resistance. In this study, the synergistic effects of
 isoparaffin and ozone were tested. The insect mortality rates (%) were not
 significantly different, reaching almost 100 percent when sprayed with SS
 (spray substance) and SS+O3 (SS containing Ozone). However, death times
 (minutes) were noticeably different. In cockroach P. fuliginosa, the death
 times were 10.5 ± 2.6 min. and 1.6 ± 0.5 min. (mean ± SD) for SS and
 SS+O3, respectively. The Asian tiger mosquito A. albopictus exhibited a
 death time of 13.5 ± 20.7 min from SS and 1.6 ± 1.4 min from SS+O3.
 Insects sprayed with SS+O3 died significantly faster, at speeds up to 6
 times those observed in SS treatment alone. SEM observation indicated
 death was caused when the spiracles of the insects were covered with
 isoparaffin degraded by ozone, resulting in suffocation. From these
 results, we conclude the combination of isoparaffin and ozone offers a
 promising new insecticide against pest insects. 
description: # Enhanced insecticidal activity of isoparaffin by ozone as an adjuvant
 Dataset DOI: [10.5061/dryad.sxksn03fk](10.5061/dryad.sxksn03fk) ##
 Description of the data and file structure These data files were used in
 *Enhanced insecticidal activity of isoparaffin* *by ozone as an adjuvant*.
 This file contains raw data on mortality, time to death, and spray volume
 organized in a CSV file. The datasets are included in a Zip file. The
 files display the raw data for the figures. "n/a" indicates
 "not applicable," and "null" signifies "no
 data" in the datasets. The file name and section of each file
 correspond to the figure and figure panel of the manuscript.
 Abbreviations: LPG (Liquefied Petroleum Gas), SS (Spray Substance),
 SS+Ozone (Spray Substance + Ozone).​ ### Files and variables #### File:
 Mortality\_rate\_Rawdata\_Fig2.csv **Description:** This file is Mortality
 rate (%) corresponding to Fig. 2 in the manuscript. Abbreviations: LPG
 (Liquefied Petroleum Gas), SS (Spray Substance), SS+Ozone (Spray Substance
 + Ozone). The files display the raw data for the figures. For Oriental
 termite *Coptotermes formosanus*, Brown marmorated stinkbug *Halyomorpha
 halys*, and Asian tiger mosquito *Aedes albopictus* were sprayed in a
 1.4-liter polyethylene container through a transparent cylindrical tube 45
 cm high (inner diameter 20 cm) for about 3 seconds. Smoky brown cockroach
 *Periplaneta fuliginosa* was sprayed from 20 cm high for 3
 seconds. "n/a" indicates "not applicable," and
 "null" signifies "no data" in the datasets. #####
 Variables * Mortality_rate: Mortality rate (%) was calculated as the
 number of deaths/total number x 100 when LPG (Liquefied Petroleum Gas), SS
 (Spray Substance), and SS+Ozone (Spray Substance + Ozone) were sprayed on
 the insects. For Oriental termite *Coptotermes formosanus*, Brown
 marmorated stinkbug *Halyomorpha halys*, and Asian tiger mosquito *Aedes
 albopictus* were sprayed in a 1.4-liter polyethylene container through a
 transparent cylindrical tube 45 cm high (inner diameter 20 cm) for about 3
 seconds. Smoky brown cockroach *Periplaneta fuliginosa* was sprayed from
 20 cm high for 3 seconds. * Number_of_Deaths: This number was counted when
 LPG (Liquefied Petroleum Gas), SS (Spray Substance), and SS+Ozone (Spray
 Substance + Ozone) were sprayed on the insects. For Oriental termite
 *Coptotermes formosanus*, Brown marmorated stinkbug *Halyomorpha halys*,
 and Asian tiger mosquito *Aedes albopictus* were sprayed in a 1.4-liter
 polyethylene container through a transparent cylindrical tube 45 cm high
 (inner diameter 20 cm) for about 3 seconds. Smoky brown cockroach
 *Periplaneta fuliginosa* was sprayed from 20 cm high for 3 seconds. *
 Survival_count: This number represents the number of insects that survived
 being sprayed with LPG (Liquefied Petroleum Gas), SS (Spray Substance), or
 SS+Ozone (Spray Substance + Ozone). For Oriental termite *Coptotermes
 formosanus*, Brown marmorated stinkbug *Halyomorpha halys*, and Asian
 tiger mosquito *Aedes albopictus* were sprayed in a 1.4-liter polyethylene
 container through a transparent cylindrical tube 45 cm high (inner
 diameter 20 cm) for about 3 seconds. Smoky brown cockroach *Periplaneta
 fuliginosa* was sprayed from 20 cm high for 3 seconds. * Total: The total
 number of insects used in this experiment is shown. #### File:
 Mortality\_rate\_Rawdata\_FigS1\_BandC.csv **Description:** This file is
 Mortality rate (%) corresponding to Fig. S1 B and C in the
 manuscript. Abbreviations: LPG (Liquefied Petroleum Gas), SS (Spray
 Substance), SS+Ozone (Spray Substance + Ozone). The files display the raw
 data for the figures. The method involved spraying insects from a height
 of 20 cm for 3 seconds. "n/a" indicates "not
 applicable," and "null" signifies "no data" in
 the datasets. ##### Variables * Mortality_rate: Mortality rate (%) was
 calculated as the number of deaths/total number x 100 when LPG (Liquefied
 Petroleum Gas), SS (Spray Substance), and SS+Ozone (Spray Substance +
 Ozone) were sprayed on the insects. The method involved spraying insects
 from a height of 20 cm for 3 seconds. * Number_of_Deaths: This number was
 counted when LPG (Liquefied Petroleum Gas), SS (Spray Substance), and
 SS+Ozone (Spray Substance + Ozone) were sprayed on the insects. The method
 involved spraying insects from a height of 20 cm for 3 seconds. *
 Survival_count: This number represents the number of insects that survived
 being sprayed with LPG (Liquefied Petroleum Gas), SS (Spray Substance), or
 SS+Ozone (Spray Substance + Ozone). The method involved spraying insects
 from a height of 20 cm for 3 seconds. * Total: The total number of insects
 used in this experiment is shown. #### File:
 Death\_time\_Rawdata\_Fig3.csv **Description:** This file is Death time
 [min] corresponding to Fig. 3 in the manuscript. Abbreviations: SS (Spray
 Substance), SS+Ozone (Spray Substance + Ozone).​ The files display the raw
 data for the figures. For Oriental termite *Coptotermes formosanus*, Brown
 marmorated stinkbug *Halyomorpha halys*, and Asian tiger mosquito *Aedes
 albopictus* were sprayed in a 1.4-liter polyethylene container through a
 transparent cylindrical tube 45 cm high (inner diameter 20 cm) for about 3
 seconds. Smoky brown cockroach *Periplaneta fuliginosa* was sprayed from
 20 cm high for 3 seconds. "n/a" indicates "not
 applicable," and "null" signifies "no data" in
 the datasets. ##### Variables * Death_time_[min]:  We measured the number
 of minutes after the insects died after being sprayed with the various
 methods written in the Description. * Standard_Deviation: Calculated from
 insect Death speed [min]. This number was calculated using Excel. *
 F-test: To calculate the comparison of variances from insect Death speed
 [min],  *F*-test was calculated using Excel.  * P_value: To conduct a
 statistical analysis, a two-tailed unpaired Student’s or Welch’s *t*-test
 was performed to calculate the P value in this study after comparing
 variances by *F*-test (p &lt; 0.05). * Significant: To conduct a
 statistical analysis, two-tailed unpaired Student’s or Welch’s *t*-test
 (*, p &lt; 0.05; **, p &lt; 0.01; ***, p &lt; 0.005; ****, p &lt; 0.0001;
 ns, no significant) were employed in this study after comparing variances
 by *F*-test (p &lt; 0.05). * Insect_number: The number of insects used in
 this experiment is shown. #### File:
 Death\_time\_Rawdata\_FigS1\_D\_and\_E.csv **Description:** This file is
 Death time corresponding to Fig. S1 D and E in the
 manuscript. Abbreviations: LPG (Liquefied Petroleum Gas), SS (Spray
 Substance), SS+Ozone (Spray Substance + Ozone).​ The files display the raw
 data for the figures. The method involved spraying insects from a height
 of 20 cm for 3 seconds. "n/a" indicates "not
 applicable," and "null" signifies "no data" in
 the datasets. In the raw data of *Coptotermes formosanus* represents
 "not_analyzed" in F-test. The result of *C. formosanus* was
 analyzed by Mann-Whitney *U*-test. ##### Variables * Death_time_[min]: We
 measured the number of minutes after the insects died after being sprayed
 with the various methods written in the Description. * Standard_Deviation:
 Calculated from insect Death speed [min]. This number was calculated using
 Excel. * F-test: To calculate the comparison of variances from Death speed
 [min] in *Halyomorpha halys*,  *F*-test was calculated using Excel.  *
 P_value: The result of *H. halys* was analyzed using two-tailed unpaired
 *t*-test with Welch’s correction, followed *F*-test (p &lt; 0.05).  The
 result of *C. formosanus* was performed by Mann-Whitney *U* test. *
 Significant: The significances were presented as follow: *, p &lt; 0.05;
 **, p &lt; 0.01; ***, p &lt; 0.005; ****, p &lt; 0.0001. *
 Insect_number: The number of insects used in this experiment is shown.
 #### File: Spray\_amount\_Rawdata\_FigS2.csv **Description:** This file is
 the spray amount corresponding to Fig. S2 in the
 manuscript. Abbreviations: LPG (Liquefied Petroleum Gas), SS (Spray
 Substance), SS+Ozone (Spray Substance + Ozone). The files display the raw
 data for the figures. Spray amount (gram) *Coptotermes formosanus*,
 *Halyomorpha halys*, and *Aedes albopictus* were tested. They were sprayed
 in a 1.4-liter polyethylene container through a transparent cylindrical
 tube 45 cm high (inner diameter 20 cm) for about 3 seconds."n/a"
 indicates "not applicable," and "null" signifies
 "no data" in the datasets. ##### Variables *
 Significant_difference: The significances were presented as follow: *, p
 &lt; 0.05; **, p &lt; 0.01; ***, p &lt; 0.005; ****, p &lt; 0.0001 and
 not_significant. * P_value: *C. formosanus* was performed by Mann-Whitney
 *U* test. The result of *H. halys* and *A. albopictus* was analyzed using
 two-tailed unpaired *t*-test with Student’s correction, followed *F*-test
 (p &lt; 0.05). * Average_of_the_spray_amount: The average amount sprayed
 on insects was calculated. Calculations were made using Excel. *
 Insect_number: The number of insects used in this experiment is shown. ##
 Code/software We used the software below. 1. Prism 10 (Statistical
 analysis software): 
 [https://www.graphpad.com/features](https://www.graphpad.com/features) 2.
 Microsoft Excel (Statistical analysis):
 [https://www.microsoft.com](https://www.microsoft.com)