Facilitating Dengue Vector Surveillance for Disease Control: A pilot study

Gulaiim Almatkyzy, M.S., John P. Grieco, Ph.D., Theeraphap Chareonviriyaphap, Ph.D., Sunny Shah, Ph.D., Nicole L. Achee, Ph.D.

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What would you say is the deadliest animal on Earth?

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Global Burden of Dengue

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Endemic in >100 countries; 390 million infections, 96 million clinical cases annually

Sources: 1. World Health Organization. (2017). Dengue and Severe Dengue. Retrieved from http://www.who.int/mediacentre/factsheets/fs117/en/2. HealthMap. (2018). Dengue Alerts for Past three months. Retrieved from http://www.healthmap.org/dengue/en/

Background: Dengue Virus

• Mosquito-borne viral infection

• Confirmed serotypes: DENV1-4 (DENV-5 is emerging)

• Range of disease -asymptomatic to flu-like symptoms and severe, fatal disease

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Female Ae. aegypti mosquito in the process of acquiring a blood meal from her human host . Source: Centers for Disease Control and Prevention

Source: Mustafa, M. S., Rasotgi, V., Jain, S., & Gupta, V. (2015). Discovery of fifth serotype of dengue virus (DENV-5): A new public health dilemma in dengue control. Medical Journal Armed Forces India, 71(1), 67-70.

Dengue Vector Control Strategies

6Source: Achee ,N.L., Gould, F, Perkins, T.A., Reiner, R.C. Jr, Morrison, A.C., Ritchie, S.A., et al. (2015) A Critical Assessment of Vector Control for Dengue Prevention. PLoS NeglTrop Dis 9(5): e0003655. https://doi.org/10.1371/journal.pntd.0003655

Diagnostic Limitations

• PCR

– Expensive

– Time consuming

– Specialized laboratory

• ELISA

– Cross-reactivity

– Seroprevelance only

• PRNT

– Expensive

– Time consuming

– Specialized laboratory

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Primary dengue infection with timing of diagnostic tests . Source: Tropical Regional Services, Queensland Health

Source: CDC. (2017). Laboratory Guidance and Diagnostic Testing. Retrieved from https://www.cdc.gov/dengue/clinicallab/laboratory.html

VecTOR dipstick

• Immunochromatographic wicking dipstick assay

• Works by antigen-antibody sandwich principle

• Developed by VecTOR Test Systems

8Source: Wanja, et al., (2014). Immuno-chromatographic wicking assay for the rapid detection of dengue viral antigens in mosquitoes (Diptera: culicidae). Journal of Medical Entomology, 51(1), 220-225.

Testing VecTOR dipstick assay. Source: Personal Archive

Study Purpose

• Goal

Improve dengue vector surveillance system at local level

• Objective

Evaluate a rapid diagnostic tool to detect DENV in mosquitoes

• Aims

– Quantify the specificity and sensitivity of the VecTOR dipstick assay to detect DENV in field-collected mosquitoes and control samples

– Assess the acceptability of the VecTOR distick assay among laboratory personnel via questionnaire

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Study Sites

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Nakhon Pathom Province,Sam Phran District

Bangkok Metropolis,Sai Mai district

Source: Maps of World. (2016). Thailand. Retrieved from https://www.mapsofworld.com/thailand/provinces/nakhon-pathom-map.html

Field Activities

• Household selection

• Indoor collection(A)

– Purposive sampling

– Prokopack aspirator

• Outdoor collection (B)

– Simple Random Sampling

– BG-Sentinel traps

• Mosquito storage

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Mosquito collection. Source: Personal Archive

Laboratory Test Work Flow

50 dipsticks

30 (pooled mosquitoes)

30 samples

4 (preliminary research)

16 (training)

8 positive control

8 negative control

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DENV Detection Summary

1306 total field-collected mosquitoes (61 households)

Site of collection Culex and Anopheles Aedes Female

Sai Mai district 57 7

Sam Phran district 947 260

KU campus 21 14

Total 1025 281

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Out of 281 pooled mosquitoes into 30 tubes

Sample Type PCR VecTOR dipstick

DENV+ (n=1) n=1 N=0

DENV- (n=29) n=29 N=29

DENV+ control samples (n=8) n=8 N=4

DENV- control samples (n=8) n=8 N=8

56.3% Cohen’s Kappa agreement between PCR and VecTOR dipstick

Questionnaire

• Distributed to laboratory personnel

• 15 closed-ended questions

• Three sections (5 questions each)

– Ease-of-use

– Practicality

– Integration

• Six-point Likert rating scale

– “0” (not applicable)

– “1” (strongly disagree)

– “5” (strongly agree)

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VecTOR dipstick assay training. Source: Personal Archive

4.2 4.2 44.6 4.6 4.8 4.4

54.4 4.47

42.8 3

3.83.2

3.8

3.2

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1.8

3.29

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2.6 2.8

3.6

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3.4

3.2

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2.2

3.2

user 1 user 2 user 3 user 4 user 5 user 6 user 7 user 8 user 9 average

Section Scores(VecTOR Dipstick)

Ease of Use Practicality Integration

Questionnaire Summary

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4.47 3.29 3.20

Summary

• Goal of dengue surveillance systems is to control spread of virus within human populations

• Dengue diagnostic tests are an essential component of disease surveillance and management systems - need for rapid, inexpensive, point-of-care tools with high sensitivity and specificity

• VecTOR dipstick assay has potential as a ‘user-friendly’ tool where capacity for PCR testing is limited:– High specificity (100%)– Fair sensitivity (44%)– Considered easy to use

• Practicality and integration were questionable

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Recommendations and Next Steps

• Train other lab personnel and community members

• Test in other regions (field-collected mosquitoes) - other flaviviruses and arboviruses to assess cross-reactivity

• Larger sample size (for sensitivity and specificity)

• Further research on practicality of the assay

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Acknowledgements• Supervisor

– Dr. Nicole L. Achee

• In-Country Supervisor

– Dr. Theeraphap Chareonviriyaphap

• Committee members:

– Dr. Sunny Shah

– Dr. John Grieco

• Special thanks:

– Dr. Wunrada Surat

– Ms. Flora Klacsmann

– Ms. Amonrat Panthawong

– Ms. Thikhumporn Sittivicharpinyo

• Laboratory assistants:

– Students from Department of Entomology, KasetsartUniversity

– Students from Department of Genetics, KasetsartUniversity

• Field assistants:

– The Department of Environment, Saimai District, Bangkok Metropolis

– The Department of Fishery Science, KasetsartUniversity

– The Head Office of Sam Pran District, Nakhon PathomProvince

– All community workers of Sam Pran District and Saimai District

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Thank You for Your Attention!

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