Honey bees that surviving Varroa destructor infestation in France

Honey bees that surviving
Varroa destructor infestation
in France
Yves Le Conte
INRA, UMR 406 Abeilles et Environnement,
Laboratoire Biologie et Protection de l'abeille,
AVIGNON, France
Historic :Apparition of Varroa in France in 1982
Untreated colonies die after 2 or 3 years of infestation !!!
1994
Come back of feral colonies
Untreated colonies
more or less abandoned
seem to survive !
1998
=>
Characterization of the survival of those colonies
to Varroa destructor
Aims :
- validate the survival phenomenon of the bees
- test different hypothesis to explain it
I-Validation of the survival of the honey bee colonies
1.
Investigation into beekeeping journals
2.
Collection and set up different apiaries with candidate colonies
3.
Criteria studied:
•
Survival of the colonies
•
Swarming
•
Honey production
•
In Avignon, population dynamics of the varroa mite
I- Validation of the survival of the honey bee colonies
Investigation to the beekeepers => set up of a different apiaries with 70
candidate colonies :
- Four apiaries in Avignon
Two in la Sarthe,
One in Orne,
I- Validation of the survival of the honey bee colonies
Queen paint marked
2 monthly visits of the colonies from
early spring to early winter
Check for diseases
No other manipulations
I- Validation of the survival of the honey bee colonies
Survival of the colonies from ‘la Sarthe’: 7.9 years
N°Colonie
1995
1996
1997
1998
1999
2000
2001
2002
2003
2004
2005
2006
2007
2008
2009
TAB
TAM
TAG
ES1
ESCH
ES2
ES3
ES4
ES5
ESSTA
757
535
222
B
F35
7.63 ± 0.3 years
(maxi: 15).
S44
601
G1
V1
27
F
URSS25
10BC3C
URSS96
264
124
692
X
174
248
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
Surviving colonies
I- Validation of the survival of the honey bee colonies
Mortality
16,8
18
16
14
14
12
%
12,5
12,5
10
9,7
10
8
6
4
2
0
98-99
99-00
00-01
01-02
02-03
03-04
Year
No evidence for mortality due to varroa infestation
N=72
Swarming
100
Swarming (%)
52
**
80
61
60
59
67
40
69
*
74
58
34
20
55
33
41
45
0
2000
2001
2002
2003
2004
2005
Year
VSB
Control
Honey production
Honey production (Kg)
40
35
71
30
58
85
25
76
20
15
40
72
66
82
65
48
61
57
37
10
49
5
0
1999
2000
2001
2002
2003
2004
2005
Year
VSB
Control
Cost due to varroa infestation
N=72
I- Validation of the survival of the honey bee colonies
No particular deseases observed inside de collected colonies
I- Validation of the survival of the honey bee colonies
Comparaison of varroa population dynamics :
- Surviving colonies from Avignon
- ‘Non surviving’ colonies from Canada
- Daugther queen of surviving colonies
Counting varroa fall twice a week
Same study made in Canada
by Pr. Gard Otis
Gérard de Vaublanc (INRA,
Avignon)
Varroas infestation of the colonies
Sensitive colonies (per week)
1800
EBA 15
Number of varroa
1600
EBA 23
1400
EBA 03
1200
EBA 21
1000
800
EBA 14
600
EBA 37
400
EBA 38
200
0
S13 S15 S17 S19 S21 S23 S25 S27 S29 S31 S33 S35 S37 S39
EBA 18
week
Differences of varroa
infestation
Surviving colonies (per week)
1800
Number of varroa
1600
EBA 24
1400
EBA 51
1200
1000
EBA 77
800
EBA 44
600
EBA 76
400
EBA 45
200
EBA 259
0
S13
S15
S17
S19
S21
S23
S25
S27
week
S29
S31
S33
S35
S37
S39
De Vaublanc, G., Otis, G.W., Le Conte, Y.,
Crauser, D., Kelly, P. 2003 – Am. Bee J. 143
(4): 319.
Varroa fall in surviving colonies and canadian colonies (per month)
4000
3500
Varroa number
3000
2500
Canadian
2000
Surviving daughter
1500
Mother surviving
colonies
1000
500
0
April
May
June
July
August
September
October
November
• Honey Bee Colonies surviving to Varroa
Different Hypothesis :
Beekeeping
methods
Environment
The honey
bee
Co-evolution
Virus
The Varroa
Hypothesis tested:
Virulence of the varroa:
Need tolls :
Genetic markers to differentiate
populations
Complet sequence of mtDNA,
17 microsatellites nuDNA
First resultat: no variability, it’s a clonal population!
Does not support hypothesis of less virulent varroa populations
Navajas, M., Le Conte, Y., Solignac, M., Cros-Arteil, S. et Cornuet, J-M. 2002.
Mol. Biol. Evol. 19: 2313-2317.
Solignac, M., Vautrin, D., Pizzo, A., Navajas M., Le Conte, Y., Cornuet, JM. 2003
Molecular Ecology Notes, 3 (4): 556-559.
Hypothesis tested:
The virus
9 different virus search in different honey bee populations
Immuno-diffusion and elisa
-
Cloudy Wing Virus : CWV
-
Deformed Wing Virus : DWV
-
Black Queen Cell Virus : BQCV
-
Acute Paralysis Virus : APV
-
Bee Virus Y : BVY and bee virus X : BVX
-
Sacbrood Bee Virus : SBV
-
Chronic Paralysis Virus : CPV
-
Kashmir Bee Virus : KBV
Isabelle Mazet (INRA Avignon)
Brenda Ball (IACR- Plant and Invertebrate
Ecology Division, Rothamsted)
and Magali Ribière (AFSSA, Sophia Antipolis)
Hypothesis tested:
The virus
Results:
Significative differences beetwen surviving and control colonies
Control colonies have more APV et CPV
Almost all the colonies have DWV
Hypothesis tested:
The virus
Injections of virus:
RS 15 AFSSA
control
Survival %
100
100
80
80
60
60
40
40
20
20
0
0
0
3
6
9
days after inoculation
TNI
ABPV d10-6
CBPV d10-6
CBPV d10-8
12
ABPV d10-8
RR 51
surviving colony
Survival (%)
15
0
3
6
9
day after inoculation
TNI
ABPV d10-6
CBPV d10-6
CBPV d10-8
No significant differences between surviving and
control colonies for APV
Nor for CPV
12
ABPV d10-8
15
Hypothesis tested: Resistance of the honey bee :
Varroa Hygienic
Capping
duration
Behavior (SMR)
Regulation of the
environment
(T°C et HR%)
swarming
Grooming
behavior
Resistance
mecanisms
individual response
Reduction of
Varroa
fertility
populationnal response
honeybee - Varroa
Interaction
Regulation of Varroa population
development
Rosenkranz P. (1999) Apidologie 30, 159-172.
Resistance of the bees to the mite:
Hypothesis tested : Grooming behavior
Ability of the bees to recognize
and destroy the varroa mite
Behavioral test =>
Differences in detection of the mite beetween the differents type of colonies
• Solid Phase MicroExtraction (SPME)
• Solid Injection (IS)
Seringue SPME
• Capillary insert in the injecteur
of the GC
Fibre
Tube with 50 Varroa
30°C
Identified Substances with SPME and IS
• Micro-Extraction
• Injection Solide
– Cuticular
hydrocarbons
– 3 Acids (palmitic, oleic,
stearic)
– 1 alcool
– Cuticular
hydrocarbons
– 8 acids
– 3 esters (palmitate,
oléate, stéarate
d’éthyle)
Martin C., Provost E., Roux M., Bruchou C., Crauser D., Clement J.L., Le Conte Y.
Physiological Entomology. 2001. 26: 4, 362-370.
Behavioral effects on the bees
Three of the compounds trigger a behavioral response
Behavioral effects on the bees
Behavioral contacts between surviving and control bees.
45
% perception of the tag
40
35
30
25
colonies S
colonies R
20
15
10
5
0
T
S
0,1
1
10
100
Ethyl oleate dose
Surviving bees are responding more than control bees
EAG gave the same results
Confirm the better capacity of the surviving bees to recognize the mite
Resistance of the bees to the mite:
Hypothesis tested : Grooming behavior
Better ability of the resistant bees to recognize
and destroy the varroa mite
Behavioral test and electro-antennography =>
Differences in detection of the mite beetween the differents type of colonies
MARTIN C., SALVY, M., PROVOST E., BAGNÈRES A.G., ROUX M., CRAUSER D.,
CLÉMENT J.L., LE CONTE Y., 2002 –Physiological Entomology. 27: 175-188.
Resistance of the bees to the mite:
Hypothesis tested : Hygienic behavior
Method:
Hygienic test: frozen brood
Results:
Our surviving bees show that behavior,
but no more compared to sensitive one.
Resistance of the bees to the mite:
Hypothesis tested : Varroa Hygienic behavior
Method:
Varroa Hygienic Sensitive bees
(Harbo JR, Harris JW. 2005. J. APIC. RES. 44:21-3)
Results:
Our surviving bees show that behavior
Resistance of the bees to the mite:
Hypothesis tested : Infertility of the varroa mite and honey bee survival.
Last week:
From 2000 to 2005,
50% for surviving colonies.
very variable results depending of the
season:
From 70% to 19% for surviving
colonies.
From 49% to 15% for canadian
colonies.
Gérard de Vaublanc (INRA, Avignon)
20 % for control colonies.
Barbara Locke
Honey bee Varroa tolerance :
Complexe phenomenon, involve many genes associated with :
metabolism, behavior, chemical communication,
reproduction…
Pangenomic approach
method for the identification of candidat genes
Using gene expression as a tool :
• to study molecular basis of host/parasite interactions and
functional genomics
• for honey bees selection against Varroa destructor (Varroa
tolerant bees could have a specific gene expression pattern that we
could use for selecting bees against the mite)
Maria NAVAJAS, INRA Montpellier, France
Yves LE CONTE, INRA Avignon, France
Gene ROBINSON, University of Illinois, USA
Charlie WHITFIELD, University of Illinois, USA
Jay EVANS, USDA, Beltsville, USA
Use gene expression to investigate
response of immature honey bees to Varroa
destructor
Effects of the parasitism and of the honey bee genotype
DNA chips
EST Chips
≈ 7000 cDNAs - ≈ 5000 genes ≈ 50% annoted
Oligo Chips
Univ. Illinois – USDA - NSF
(A)
Atg18, pUf68
Rab7
Immune system
Inflammatory
response
Deformed
Effects of the parasitism:
Wing Virus
Pcmt , Nedd8
Cellular and molecular
31 genes were moving
damages
baz , dgl1,
sgl
Embryonic
ple , Atg18,
Dlic2
Brain
development
Deformed
adult
Cognitive
impairment
Hypothetical pathways and models of honey bee responses to Varroa-parasitism. Arrows
and dashes indicate positive and negative regulation, respectively; dotted lines are possible links
between two factors.
NAVAJAS M., MIGEON A., ALAUX C., MARTIN-MAGNIETTE ML., ROBINSON GE., EVANS JD., CROS-ARTEIL S., CRAUSER D., LE CONTE Y.
2008. BMC Genomics 2008, 9:301doi:10.1186/1471-2164-9-301.
(B)
Candidate genes
for afor
behavioural
resistance
Candidate
genes
a behavioural
resistance
Futsch , scrt , otk ,
Bchs
Mhcl , gro , fng , Bchs
Nervous system
Dhc64c
Dhc64c
poe
α, para
poe, GluCl
GluClα,
para
Responsiveness
to stimuli
Mushroom
bodies
smi21F
su(w a), poe
poe ,
smi21F , su(w
para
rogdi
para, rogdi
Effects of the genotype:
Dscam , fwd
Olfaction
99 genes were moving
Others genes
Others genes
Dscam , otk
Immune system
Ahcy13, para
Ahcy13, para
ResistanceResistance
to toxins to toxins
Hypothetical pathways and models of honey bee response to the bee tolerant genotype.
Gene names are up-regulated.
NAVAJAS M., MIGEON A., ALAUX C., MARTIN-MAGNIETTE ML., ROBINSON GE., EVANS JD., CROS-ARTEIL S., CRAUSER D., LE CONTE Y.
2008. BMC Genomics 2008, 9:301doi:10.1186/1471-2164-9-301.
2006 :
Year of the sequencing of the honey bee genome
=> New ADN cheap including the hole genome
conception: Université d’Illinois + USDA + NSF
13440 oligos
Second approach: adult worker responses
USDA Baton Rouge, Louisiane
J. Harris and J. Harbo
Gene expression on the Varroa hygienic trait
Results:
• olfaction involved
Prospects:
Make replicates on other selected bees
Find the genes involved
Use them as toll in honeybee selection
Conclusion
• Some honey bee strains can survive to the Varroa mite in
France
• It is a good start to develop IVM.
• Olfaction clearly involved in varroa tolerance
• Varroa hygienic behavior seems to be a good trait to select to
get resistant bees (behavioral and molecular results on
olfaction).
• Genome sequencing and gene expression techniques should
bring new tools for honey bee selection in the future
As for the case of Acarapis woodi in the past, we can hope that most of
the bees will become resistant to Varroa destructor