A Comparative Study on the Efficacy of N-N-diethyl-m-toluamide, Picaridin, and Citronella as Repellents Against Aedes Aegypti

January to August 2012 –Dengue Fever:7321 cases –Dengue Hemorrhagic Fever:4181 cases January to August 2013 –Dengue Fever:2494 cases –Dengue Hemorrhagic Fever:1552 cases

Recent large clinical trial conducted in Thailand yielded only 30% effectivity of the dengue vaccine, 40% short of it’s expected outcome.

Celgosivir (6-O-butanoyl castanospermine), which misfolds the dengue NS1 protein is still in the initial stages of clinical trials.

Most Feasible Prevention

Insect Repellents DEET (N, N diethyl-meta-toluamide) –GOLD STANDARD –Blocks insect olfactory receptors –Toxic effects: impaired cognitive function Insomnia mood distrubances, severe epidermal reactions

Insect Repellents Non-DEET containing –Picaridin –Citronella

PICARIDIN ( 2-(2-hydroxyethyl)-1-piperidinecarboxylic acid 1-methylpropyl ester) used in Europe, Australia, Latin America and Asia as an alternative to DEET Unknown exact mechanism of action but repellency is comparable to that of DEET included in the list of effective repellents by the CDC

CITRONELLA plant-based insect repellent product from the steam distillation of freshly cut or partially dried grasses unkown exact mechanism of action but acts mainly as a feeding depressant US Environmental Protection Agency classifies citronella’s dermal toxicity as Category III or only slight toxicity if improperly used

General Objective –To compare the efficacy of different variants of insect repellents (DEET containing versus non-DEET containing versus plant-based) against mosquito landing and bites

Specific Objectives 1.To determine the LANDING frequency (mosquitoes landed on the subject, but not necessarily bit) for each variant of insect repellent 2.To compare the MEAN LANDING frequency of the insect repellents 3.To determine the BITING frequency for each variant of insect repellent

Specific Objectives 4. To compare the MEAN BITING frequency for the insect repellents 5. To determine which of the insect repellent variant provides the best protection against mosquito bites 6. To observe adverse events/reaction, if there is any, upon application/usage of the insect repellents

Materials and Methods STUDY DESIGN –Double blind randomized controlled trial Proposal reviewed and approved by IRB SETTING

STUDY POPULATION –any male or female –aged years old –without pre-existing co-morbidities and allergies

Aedes Aegypti Mosquitoes

SAMPLE SIZE –9 (WHO and RITM protocol)

Repellents Tested A: N-N-diethyl-meta-toluamide (DEET) 25% B: Picaridin 10% C: Citronella

Informed consents were secured. Subjects were randomly allocated to three groups.

Eight trials Test mosquitoes replaced for each trial Each trial ran for 5 minutes Repellent was not reapplied prior to the next trial

Adverse Reactions were recorded –Rashes –Pain –Discomfort –Difficulty of Breathing

Events LANDING FREQUENCY: number of mosquitoes that LANDED on the test limb instantaneously BITING FREQUENCY: number of mosquitoes that landed on the test limb and are able to BITE the test limb, may be evidenced by the appearance of a rash/papule on the test limb

Statistical Analysis One-Way Analysis of Variance –To prove that at least two of the means are significantly different Tukey’s Test –To compare specific pairwise differences

Results

Landing Frequencies TRIA L MEANTOTA L DEET A B C PICARIDIN A B C CITRONELLA A B C

Hourly Landing Frequency

Biting Frequencies TRIA L MEANTOTA L DEET A B C PICARIDIN A B C CITRONELLA A B C

Hourly Biting Frequency

One Way ANOVA (Landing) Critical Value (F): 5.14 Computed Test Statistic: Decision: Reject Null Hypothesis SOURCEDegrees of Freedom Sum of Squares Mean Squares F Treatment Error Total

One Way ANOVA (Biting) Critical Value (F): 5.14 Computed Test Statistic: Decision: Do not Reject Null Hypothesis SOURCEDegrees of Freedom Sum of Squares Mean Squares F Treatment Error Total

Tukey’s Test (Landing) D min: SET-UPABC A B C

NO allergic reactions noted

Discussion DEET, the gold standard, provided the best protection against mosquito bites based on landing and biting frequency Picaridin and citronella subjects acquired more landing and bites

-Consistent with studies done by Fradin and Beeyer -DEET is still the most effective mosquito repellent in the market.

Discussion INCREASE in the number of landing and bites as the trials progressed (waning of repellency)

Discussion One Way ANOVA for LANDING frequency: mean landing frequencies are SIGNIFICANTLY DIFFERENT from one another. One Way ANOVA for BITING frequency: mean biting frequencies are NOT SIGNIFICANTLY DIFFERENT from one another.

Discussion Less number of bites compared to landing in general Raw data: DEET subjects had the LEAST NUMBER of bites (best protection among the three)

Discussion Tukey’s test: In general, mean landing frequencies are SIGNIFICANTLY DIFFERENT

Discussion Consolidating raw data and statistical analysis –DEET is SUPERIOR to Picaridin and Citronella in repellency, based on LANDING and BITING frequency

Conclusion DEET is SUPERIOR to Picardin and Citronella based on LANDING and BITING frequency raw data One Way Analysis of Variance: MEAN LANDING frequencies are SIGNIFICANTLY DIFFERENT, MEAN BITING frequencies are NOT SIGNIFICANTLY DIFFERENT

Tukey’s Test: In general, MEAN LANDING frequencies are SIGNIFICANTLY DIFFERENT DEET provides the BEST PROTECTION against Aedes Aegypti

Limitations and Recommendations Minimum required sample size employed, larger sample size needed for a more conclusive study Only the landing and biting frequency were measured. Toxicity is another important parameter Future studies: recommendations on the safe levels of insect repellents