Follow-up training MediLabSecure

Training follow-up & evaluation External Quality Assessment and MosPictoQuizz WP4 Medical entomology - December, 2017

REVIEW Novembre 2017

Reminder of the capacity building activities

Training needs assessement • Questionnaire • Tailored trainings

2014

2015

Training courses • Capacity building in mosquito vector of arboviruses • In Serbia, Turkey, Tunisia • For the 19 participating countries

2016

Training follow-up & evaluation • EQA (External Quality Assessment) • MosPictoQuizz

2017

2018

EQA (External Quality Assessement) on mosquito identification

To evaluate the laboratory capacity to correctly identify vectors

Focuses on the mosquitoes (adult and larva) For all the medical entomology laboratories of the MediLabSecure network.

Participant laboratories 19 laboratories from the medical entomology network (WP4) Kosovo / Institute of Vet. medecine Fac of Univ. of Prishtina, Fac of Agri and Vet.

Bosnia &Herz. / Lab. Diagnostic Veterinary Institute of the Rep. of Srpska

Serbia / Lab for med and vet entomology Fac of Agriculture, Univ. Novi Sad

Montenegro / Lab for applied zoology Biotechnical faculty

Moldova / Lab of systematics and molecular phylogeny Institute of zoology Ukraine / Lab of Especially Dangerous Infections Epizootology State Body "Ukrainian I.I. Mechnikov Research AntiPlague Institute of Ministry of Health of Ukraine" Turkey / HU-ESRL-VERG (Vector ecology research group) Hacettepe Univ.

FYROM / Lab for virology and molecular diag Institue of Public Health

Georgia / The R. Lugar Center for Public Health Research National Center for Disease Control and Public Health

Albania / Entomology lab Institute of Public Health

Armenia/ Reference lab center National Center for Disease Control

Morocco / Medical entomology lab Institut National d'Hygiène Morocco / Lab. des Maladies Vectorielles Institut Pasteur

Lebanon / Lab of immunology National Center for Disease Control and Public Health

Algeria / Eco-epidémiologie parasitaire et génétique des populations Institut Pasteur

Jordan / Parasitic and Zoonotic Diseases Department Ministry of Health

Palestine / Vector Control Unit Ministry of Health Tunisia/ Medical entomology lab Institut Pasteur

Libya / Lab of parasitology and vector born diseases National Center for Disease Control

Materials and methods A mosquito box composed of: - 7 or 8 mounted adult mosquitoes - 4 mosquito larvae in alcohol All the specimens were fresh, collected the previous few months.

All available resources were accepted to support identification (dichotomical keys, interactive key for mosquito species, molecular methods‌).

Constraints Despite the precautions regarding the provided materials, some specimens could have been damaged during the transportation. This option has been considered and it has been admitted that the challenging determination of poor preserved specimen relies within the area of the medical entomology expertise. Participants have been invited to take this challenge and to manage this possibility the best they could.

9/143 specimens were considered as damaged by the participating labs.

Damaged specimens were not taken into account in the results.

Methods and resources used by participants Resources used for identification 8

100 % morphological identification

7 6 5 4 3

2 1 0 MosKeyTool only

Dicho key + MosKeyTool

Dicho key only

Dicho key + Books (Becker, Harbach)

All labs performed the exercise by

morphological identification, using interactive identification key and dichotomical key.

Dichoto key + MosKeyTool + Books (Becker, Harbach)

Results General results of the mosquito identification from the participant laboratories (rate of correct answers)

100%

87%

90% 80%

80%

77%

% right answers

70% 60%

66% 62%

65% Genus

50%

Species 40% 30% 20% 10%

0% Adults

Larvae

All (adults+larvae)

Results per lab (adults + larvae) General results of the mosquito identification (adults + larvae) from the participant laboratories 100% 90%

% right answers

80% 70% 60% 50%

40% 30% 20% 10% 0%

Lab1 100%

Lab2 100%

Lab3 100%

Lab7 100%

Lab4 92%

Lab5 92%

Lab6 92%

Lab8 100%

Lab9 83%

Species 100%

100%

100%

92%

92%

92%

92%

83%

75%

Genus

Total rate of correct answers for all labs:

Lab10 Lab11 Lab12 Lab13 Lab14 Lab15 Lab16 Lab17 Lab18 Lab19 92% 75% 83% 73% 82% 64% 67% 45% 44% 42% 64%

63%

58%

50%

45%

45%

33%

18%

Genus identification: 80% Species identification: 65%

11%

8%

Results per lab (adults) General results of the mosquito identification (adults) from the participant laboratories 100% 90%

% right answers

80% 70% 60% 50% 40% 30% 20% 10% 0%

Lab1 100%

Lab2 100%

Lab7 100%

Lab3 100%

Lab8 100%

Lab6 100%

Lab4 88%

Lab5 88%

Lab9 75%

Species 100%

100%

100%

100%

100%

100%

88%

88%

75%

Genus

Total rate of correct answers for all labs:

Lab12 Lab10 Lab11 Lab16 Lab13 Lab14 Lab15 Lab17 Lab18 Lab19 88% 86% 50% 75% 57% 71% 43% 43% 40% 38% 63%

57%

50%

50%

43%

43%

43%

29%

Genus identification: 77% Species identification: 66%

20%

0%

Results per lab (larvae) General results of the mosquito identification (larvae) from the participant laboratories 100% 90%

% right answers

80%

70% 60% 50% 40% 30% 20% 10% 0%

Lab1 100%

Lab2 100%

Lab4 100%

Lab5 100%

Lab3 100%

Lab7 100%

Lab9 Lab10 Lab13 Lab11 100% 100% 100% 100%

Lab6 75%

Lab8 Lab14 Lab15 Lab12 Lab18 Lab19 Lab16 Lab17 100% 100% 100% 75% 50% 50% 50% 50%

Species 100%

100%

100%

100%

100%

75%

75%

75%

50%

Genus

75%

75%

Total rate of correct answers for all labs:

75%

50%

50%

50%

50%

25%

Genus identification: 87% Species identification: 62%

0%

0%

List of species to identify Adult species

Larva species

List of adult species to identify for the EQA Species (adult) Aedes albopictus Aedes vittatus Culiseta longiareolata Orthopodomyia pulcripalpis Aedes vexans Culex pipiens Culex hortensis Anopheles maculipennis s.l. ≠Culicidae Aedes caspius Culiseta annulata Anopheles claviger s.l.

Total

Nb specimens to identify 19 19 17 19 15 18 11 6 4 3 2 1 134

Nb of adult specimens to identify per lab: 7

or 8

List of larva species to identify for the EQA Species

Nb specimens to identify

Aedes vittatus Aedes detritus Culiseta longiareolata Orthopodomyia pulcripalpis Total

Nb of larva specimens to identify per lab: 4

19 19 19 19 76

Results per species (adults) General results per species of the adult mosquito identification from the participant laboratories 100% 100%

95%

90%

82%

80%

74%

71%

71%

74% 68%

67%

70%

% right answers

73% 63%

67%

56%

60%

45%

50%

Genus Species

40% 30% 20% 10% 0% Aedes albopictus

Culiseta longiareolata

Aedes vittatus

Aedes vexans

Total rate of correct answers for all labs:

Orthopodomyia pulcripalpis

Culex pipiens

Culex hortensis

Genus identification: 77% Species identification: 66%

Results per species (larvae) General results per species of the larva mosquito identification from the participant laboratories 95%

100%

90%

84%

84%

74%

80%

% right answers

84%

84%

70% 60%

47%

Genus

50%

37%

40% 30% 20% 10% 0%

Orthopodomyia pulcripalpis

Culiseta longiareolata

Total rate of correct answers for all labs:

Aedes detritus

Aedes vittatus

Genus identification: 87% Species identification: 62%

Species

Results Are the EQA results linked to the presence/absence in the country ?

% of species right answers 100% 90% 80% 70%

68%

57%

60% 50% 40% 30% 20% 10% 0%

Species present in the country

Species absent in the country

EQA feedback For your lab, this testing has been...

Success rate EQA vs estimated difficulty level 120% Lab1

Difficult 23%

Easy 59%

Lab2

100%

Lab3

% right answers

Very difficult 6% Very easy 12%

Lab4

80%

Lab5 Lab6

60%

Lab7 Lab9

40%

Lab10

Lab11 20%

Lab12 Lab13 Lab14

0% Very easy

Easy

Difficult

Very difficult *5 non-respondent labs

EQA feedback Number of person per lab who performed the EQA

8 7 6 5 4 3 2 1 0 1

2

3 or more

Don't know

EQA comments… … about the damaged materiels Some samples were broken/damaged

Some insects have been damaged, which could lead to errors in identification

… about the EQA benefits

Thank so much for this opportunity to evaluate our ability to identify mosquito

… about the difficulty to identify adult or larva

For our lab in general it will be difficult

We had a bit difficulties during the larvae identification!

This is a good exercise for our laboratory. We found difficulties in identifying adults.

Thank you

Overview of the species to identify

Aedes albopictus An invasive species Mainly black with white stripes on legs and dorsal abdomen. silvery median white line on the scutum

Aedes vittatus Scutum with 6 pale spots, tibiae with median pale ring are diagnostic characters.

Aedes vittatus The larva is recognisable by the implantation of the seta 1-S between the last and the penultimate tooth of the siphon comb.

7/17 labs confused this species with other Aedes sp.: Aedes aegypti (3), Aedes vexans (2), Aedes albopictus (1), Aedes cinereus or rossicus (1)

Aedes vexans Tarsi with pale basal rings and tergum with bilobed basal bands.

Culiseta longiareolata Easy to identify due to the presence of longitudinal lines on the scutum and rows of pale patches on the femur.

3/17 labs confused this species with Orthopodomyia pulcripalpis

Culex pipiens It's impossible to differentiate Culex pipiens s.s. from Culex quinquefasciatus. Difficult to differentiate from Culex torrentium, male genitalia examination is necessary.

Confirmation with molecular techniques is recommended.

The apex of all femora with narrow spot of yellowish scales.

Tarsi are uniformly dark brown.

Culex hortensis This species has numerous white scales on the thoracic pleurae. And apical pale bands that extends forward in a median triangle on the abdomen.

Aedes detritus Impossible to differentiate from Ae. coluzzi with only morphological criteria. Isoenzymes are diagnostic and a multiplex PCR will be useful for identification. Differentiable also by bioecological specificities.

7/17 labs confused this species with other Aedes sp.: Aedes vexans (3), Aedes vittatus (2), Aedes caspius (1), Aedes rossicus (1)

Orthopodomyia pulcripalpis Distinctive pattern on the scutum and white scale patch on the basal part of the wing allows to identify this species very easily.

5/19 labs confused this species with Aedes aegypti

Orthopodomyia pulcripalpis Abdominal plates on segments VI, VII, and VIII are diagnostic for the genus Orthopodomyia.

MosPictoQuizz

To maintain the mosquitoes species determination among the routine activities A bimonthly online picture activity to identify adult mosquitoes

Materials and methods A online picture quiz open to all, started in January 2017.

Every 2 months, a set of pictures of an unidentified mosquito are proposed for identification.

The results are analyzed and reported to participants the following month, accompanied by a mosquito factsheet focusing on the characteristics of the given species, its geographical distribution, the species not to be confused with, etc.

Mosquito factsheet

MosPictoQuizz#1 results

98%

Aedes albopictus

gave the right answers for the genus identification

91% gave the right answers for the species identification

46 participants

19 countries

Aedes albopictus is currently the most invasive mosquito in the world.

The contrast between dark and silver scales, and the median line on the scutum are indicative. Note the scutellum in three parts, covered of silvery scales.

MosPictoQuizz#2 results

97%

Anopheles sacharovi

gave the right answers for the genus identification

80% gave the right answers for the species complex identification Anopheles sacharovi is an important malaria vector of the Maculipennis complex. The wing pattern of Anopheles maculipennis s.l. -absence of a patch of clear scales on the wing fringe- is here typical.

33 participants

14 countries

MosPictoQuizz#3 results

90%

Culiseta annulata

gave the right answers for the genus identification

90% gave the right answers for the species identification

29 participants

17 countries

Culiseta annulata is a widespread and common mosquito in Western Europe and parts of the Middle East.

Diagnostic characters were: well marked clear band on the abdominal terga, spots of dark scales on the wings, etc.

MosPictoQuizz#4 results

100%

Aedes aegypti

gave the right answers for the genus identification

100% gave the right answers for the species identification

26 participants

15 countries

Aedes aegypti is an invasive mosquito recently reestablished in Madeira and around the Black Sea. Diagnostic characters are mainly on the scutum with the famous "lyre-shaped" with the two fine central longitudinal bands (the strings of the lyre!).

MosPictoQuizz#5 results

100%

Culex pipiens

gave the right answers for the genus identification

90% gave the right answers for the species identification

20 participants

13 countries

Culex pipiens s.s., as well as its tropical vicarient Culex quinquefasciatus, are members of the Culex pipiens complex.

Morphology of females does not permit to distinguish the two species.

MosPictoQuizz#6 results

100%

Anopheles claviger

gave the right answers for the genus identification

57% gave the right answers for the species identification Anopheles claviger s.s., as well as its sibling species Anopheles petragnani, are members of the Claviger Complex. An. claviger s.s. occurs across most of Europe, up to the Middle East and North Africa, whereas An. petragnani is so far restricted to the western Mediterranean region.

35 participants

13 countries

MosPictoQuizz participation Nb of participant /

Nb of country /

50 46 45 40 35 35

33 29

30

26 25 20

19

20

17 15

14

15

13

13

10

5 0 #1

#2

#3

#4

#5

#6

MosPictoQuizz feedback MosPictoQuizz#1

MosPictoQuizz#3

Summary of results

~50% of the labs (9/19) were able to identify more than 75% of the specimens, which means between 0 and 3 mistakes.

19 participant labs to the EQA

12 different species

210 specimens to identify

Good results (87%)

#6

on genus identification of larvae.

MosPictoQuizz

All the entomological labs are able to identify

invasive species (Aedes albopictus and Aedes aegypti).

Recommendations Mosquito identification is an important step in entomological surveillance.

A regular practice, on top of routine activities, is highly recommended.

Even though we usually feel more comfortable with larva and/or adult identification, do not neglect any of those two.

Practice equally larva and adult identification.

The profession of medical entomologist have changed considerably these last decades.

Fieldwork remains fundamental as the transmission of infectious agents is above all a question of ecology.

This MediLabSecure activities has been organized by the French National Research Institute For Sustainable Development (IRD).

This medical and veterinary entomology group is coordinated by Dr. Vincent Robert and Marie Picard at MIVEGEC unit, IRD.

The MediLabSecure project is supported by the European Commission (DEVCO: IFS/201010/23/_194)