RESEARCH

ANYTIME® no-bite™ LOTION MOSQUITO REPELLENT & ANYTIME® no-bite™/no-burn™ LOTION MOSQUITO REPELLENT

Summary

The efficacy of a natural formulation of mosquito repellent (Anytime Repellent) and a combination of mosquito repellent and sunscreen (Ultra 2in1Combination Mosquito Repellent & Sunscreen) was assessed by conducting repellency studies using adult mosquitoes reared from larvae sourced from a site in St. Andrew, as well as larvae from a susceptible strain of mosquitoes,  maintained under laboratory conditions. Untreated hands (control) and hands treated with the sample repellants were exposed to the adult mosquitoes and the number of female mosquitoes seeking a blood meal was recorded. The mean number of mosquitoes seeking a blood meal ranged from 10.5 ± 0.76 to 11.3 ± 0.55 for the control compared to 0.50 ± 0.34 and 0.83 ± 0.31 for Anytime Repellent and 0.17 ± 0.17 and 0.50 ± 0.22 for the Ultra 2in1 Combination. The mean level of repellency was 93.2 ± 2.3% and 94.2 ± 3.9% for Anytime Repellent and 95.8 ± 1.90% and 98.6 ± 1.4% for Ultra 2in1 Combination. The test concluded that both insect repellants were effective against the randomly selected population of mosquitoes.

Preamble

 This study was conducted at the request of Mr. Ian Gilbert to determine the repellent effect of a natural formulation of mosquito repellent (Anytime Repellent) and a combination of mosquito repellent and sunscreen (Ultra 2in1Combination) against a selected population of mosquitoes in Jamaica.

Project Goal and Objectives

Goal: To determine the relative efficacy of Anytime Repellent and Ultra 2in1 Combination against mosquitoes in Jamaica.

Objectives:

  1. Assess the level of blood seeking activity of mosquitoes on human skin treated with Anytime Repellent and Ultra 2in1 Combination.
  2. Calculate the level of repellency of Anytime Repellent and Ultra 2in1 Combination against the selected mosquito population.

Methodology

The study was conducted using mosquito larvae sourced from a site in St. Andrew, as well as larvae from a susceptible strain of mosquitoes,  maintained under laboratory conditions. Plastic containers (~1 Litre) containing at least 40 mosquito larvae were fed and placed in specially designed (~30,000 cm3 ) testing chambers to allow for pupation and the emergence of adult mosquitoes. The chambers consisted of a PVC frame covered with a 1.2 mm white nylon mesh. At one end was a specially designed opening to allow a human hand to be placed in the chamber while preventing the escape of the mosquitoes.

The chambers were checked several times each day and as soon as a minimum of 20 female adult mosquitoes were observed, the container with the water containing the immature stages was removed in preparation for the assessment. Tests were done using human volunteers. The volunteers’ forearms were washed with water and unscented soap and then dried. One hand was randomly selected and treated by applying the Anytime Repellent or the Ultra 2in1 Combination Mosquito Repellent & Sunscreen as directed on the label.

The volunteer was asked to place the untreated (control) hand in a test cage containing the adult mosquitoes for a two-minute period and the number of mosquitoes seeking to obtain a blood meal recorded. After two minutes the untreated hand was replaced with the treated hand and a similar assessment done. The untreated hand was again exposed to the adult mosquitoes after the treated hand was removed to assess the mosquitoes continued interest in seeking a blood meal. The assessment was repeated at 10-minute intervals for a total of one hour. Six replicates were done for each treatment.

The percent repellency was determined by calculating the difference between the number of mosquitoes seeking a blood meal from the control hands and the treated hands. The data was collated and analyzed in SPSS Statistics Version 19.

Results and Conclusion

Data on the number of mosquitoes seeking a blood meal from treated and untreated hands are presented in Table 1. The mean number of mosquitoes seeking a blood meal ranged from 10.5 ± 0.76 to 11.3 ± 0.55 for the control compared with 0.17 ± 0.17 to 0.83 ± 0.31 for the treated 4 hands. The mean level of repellency was 93.2 ± 2.3% and 94.2 ± 3.9% for Anytime Repellent and 95.8 ± 1.90% and 98.6 ± 1.4% for Ultra 2in1 Combination (Table 2). It should be noted that none of the mosquitoes landing on the treated hands sought to take a blood meal. Based on the data it was concluded that both products were effective in repelling both populations of mosquitoes.

Table 1: Number of female mosquitoes from selected mosquito populations seeking a blood meal from human hands treated with Anytime Repellent or the Ultra 2in1 Combination Mosquito Repellent & Sunscreen. 

    Table 1 Mosquitos Seeking Blood Meal

Table 2: Calculated repellency of Anytime Repellent and Ultra 2in1 Combination Mosquito Repellent & Sunscreen against a Jamaican population of mosquitoes.

    Table 2

 

 

ANYTIME® no-bite™ LOTION MOSQUITO REPELLENT ANYTIME® no-bite™/no-burn™ LOTION MOSQUITO REPELLENT, STILL TAILS™, & ZONE DEFENSE

Abstract

All Bradshaw’s 4 Ring Protection ANYTIME™ formulations performed as well as Off! 7% DEET at 0, 1, 2, 4, and 6 hours post- treatment. Y&G Spray yielded the greatest repellency ranging from 94-100% throughout the five time intervals; however, there was no significant difference (p >0.05) in biting activity among the tested repellents.

Objective

Quantify and compare repellency of four Bradshaw’s 4 Ring Protection ANYTIME™ formulations to a DEET standard, and a non-treated control against yellow fever mosquitoes, Aedes aegypti.

 Materials & Methods

  1. This study was performed on March 10, 2017 in a temperature-controlled laboratory following techniques developed by Klun et al (2008).
  2. Bioassays were conducted in nine, six-chambered Plexiglasss K&D modules interconnected with hoses to a water bath supplied with a temperantre-controlled inversion circulator (Fig. l).
  3. Five colony-reared female Aedes aegypti were aspirated into each chamber (Fig 2).
  4. Nine replications of the following treatments were randomly assigned to the chambers (Fig. 3):
    1. Bradshaw’s 4 Ring Protection ANYTIME™ formulations:
      1. STILL TAILS™
      2. ANYTIME™ No-Bite
  • ZONE DEFENSE
  1. ANYTIME™ No-Burn
  1. Positive control - 7% DEET standard (Off! Family Care Unscented)
  2. Negative control - Non-treated
  1. Repellents were applied to nine ca. 2”x2’’x9” strips of organdy cloth and spread with a small paint brush over 12 cm2 ink-pen-demarcated rectangles drawn on the strips with a flat plastic template. A micropipettor set at 27.6 µl was used to apply liquid repellents, while creams were applied at 0.06 g (Fig. 4 a-d).
  2. Treated clothes were taped to flat plastic templates with openings aligning with lower Plexiglasss bases

(Fig. 5).

  1. The lower Plexiglasss bases contained shallow surface wells that were filled with a blood substitute (CDTA

and ATP) and covered with moistened collagen membranes (Fig (6) .

  1. The blood substitute was heated to 38 °C with water pumped through hose lines attached to the lower bases and the water bath (Fig. 6).
  2. The plastic templates were fitted between the K&D modules and lower Plexiglasss bases (Fig. 1).
  3. Mosquitoes were exposed to the treated surfaces by opening the K&D module sliding doors for 90-second

biting counts at five post-treatment time intervals (0, 1, 2, 4 & 6 hrs) (Fig. 1).

  1. Fresh mosquitoes were aspirated into the chambers for each time
  2. Ambient temperature and humidity were continuously recorded throughout the study with a HOBO data logger.
  3. The dependent variable was biting count mean. Treatment and module means were independent
  4. Biting counts as well as log and square root transformed biting counts by treatment were examined for normality with SAS PC Proc
  5. The variance in biting count means by treatment and time intervals was statistically analyzed with SAS Proc ANOVA and statistically tested for significant differences with Tukey’s Studentized Range test at p<0.05 and charted with 95% Confidence Intervals (C.I.).
  6. Average % repellency was calculated with the following formula and charted by treatment and time interval: (Control Biting Count - Treatment Avg. Biting Count) ÷ Control Avg. Biting Count x 100.

Fig. 1. Upper K&D modules stacked on lower bases and treated cloth-covered templates. Doors opened to expose mosquitoes to treatment surfaces.

Fig. 1. Upper K&D modules stacked on lower bases and treated cloth-covered templates. Doors opened to expose mosquitoes to treatment surfaces.


Fig. 2. A K&D module equipped with a sliding door beneath each of six chambers stocked with 5 female mosquitoes.
Fig. 2. A K&D module equipped with a sliding door beneath each of six chambers stocked with 5 female mosquitoes.

Fig . 3. Repellent treatments (STILL TAILS™, ZONE DEFENSE, no-bite, no-bite/no-burn, and DEET 7%) (left to right).

Fig . 3. Repellent treatments (STILL TAILS™, ZONE DEFENSE, no-bite, no-bite/no-burn, and DEET 7%) (left to right).

             test a                      test b

          test d      test c

Fig. 4. Process for applying repellents to cloth strips, a. ink-demarcated cloth strips taped to dish pan,

  1. liquid repellents applied with micropipettor and paint brush, c. cream repellents weighed and applied with paint brush, d. cloth strips with treatments applied.

Fig. 5. Treated clothes attached to plastic templates placed on top of lower Plexiglasss bases.


Fig. 5. Treated clothes attached to plastic templates placed on top of lower Plexiglasss bases.

 

Fig. 6. Lower bases containing blood substitute filled surface wells covered with collagen and interconnected to temperature controlled water bath.

Fig. 6. Lower bases containing blood substitute filled surface wells covered with collagen and interconnected to temperature controlled water bath.

Results 

Environmental Data:

The bioassays were started at 8:15 a.m. and ended at 2:30 p.m. (Fig. 7). The range in ambient temperature and humidity was 81-82 °F and 40-46% R.H., respectively. Temperature and humidity were relatively stable throughout the post-treatment intervals.

graph1

Fig. 7. Ambient temperature and humidity data during bioassays.

Normality Tests:

Biting counts by treatment were found to follow closer to a normal distribution compared to log or square root transformed data. Consequently, all statistical analysis was conducted on non-transformed data.

Biting Count and Repellency:

There was no significant difference (p>0.05) in average biting counts among the nine K&D modules. Biting counts in the controls were significantly (p<0.05) greater than all of the repellent treatments at 0, 1, 2, and 6 hours post-treatment (Fig. 8). At 4-hours, there was no significance difference (p>0.05) in biting activity among the control, DEET and no-burn/no-bite; however, significantly (p<0.05) fewer bites occurred for Y&G Spray and Hooves compared to the control. There was no significant difference (p>0.05) in bites among the five repellents at 4 hours.

graph2


Fig. 8. Aedes aegypti biting count averages and 95% C.I. at 5 time intervals for Control, DEET, 
no-bite/no-burn, STILL TAILS™, and ZONE DEFENSE. Means within time intervals followed by different letters were significantly different at p<0.05.

When biting count data were converted to percent repellency, all of the Bradshaw’s 4 Ring Protection ™ formulations appeared to perform as well, if not better, than the 7% DEET (Fig. 9). The ZONE DEFENSE yielded the highest   level of repellency ranging from 94-100% throughout 6-hour. Hooves ranged from 81-100%. None of the other formulations including the DEET 7% standard produced >95% repellency. DEET and no-bite/no-burn yielded the lowest repellency at 53% at 4 hours.

graph3

Fig. 9. Percent repellency of Bradshaw’s 4 Ring Protection™ formulations compared to 7% DEET

against Aedes aegypti at 0-6 hours post-treatment.

Literature Cited 

Klun, J.A., M.A. Kramer, A. Zhang, S. Wang, and M. Debboun. 2008. A quantitative in vitro assay for mosquito deterrent activity without human blood cells. J Am. Mosq. Contr. A