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J. Plant Prot. and Path., Mansoura Univ., Vol.6 (1): 235 - 243, 2015
EVALUATION OF SOME ESSENTIAL OILS FOR
CONTROLLING VARROA MITES AND THEIR EFFECTS ON
BROOD REARING ACTIVITY IN HONEY BEE COLONIES.
El-Hady, A. M.
1
; E. E. Nowar
2
and M. F. EL-Sheikh
3
1-Department of Bee Research, plants Protection Institute, ARC,
EL- Doki -Giza. Egypt
2-
Department of plant Protection, Faculty of Moshtohor, Benha Univ.
3-Department of plant Protection, Economic Entomology,
Faculty of Tanta, Tanta Univ.
ABSTRACT
This study was carried out to assess the efficacy of controlling essential oils
i.eThymol, Cinnamon and Anise oils with different concentrations compared to 65%
Formic acid in controlling Varroa destructor mites infesting colonies of Apismellifera
during the winter and summer of 2013 season. Randomized complete block design
was applied at this study. The apiary was divided into 7 groups (3 colonies / group).
Three application of the oils. Average of Varroa infestation level before treatment
ranged between 21–29 in winter and from 37-47 in summer. The treatments were
significantly effective when applied in summer than the winter season and when used
at 30% concentration than those obtained at 15% concentration compared to the
control treatment. Average efficacies percentage ranged between 80.95% (Cinnamon
oil 30%) to 86.99 (Thymol 30%) in winter and from 67.46 %(Thymol 30%) to 82.16%
(Cinnamon 30%). Thymol oil was the best in winter, however, Cinnamon was the best
in summer. Anise oil 30% was intermediate 80.00 and 77.11% in winter and summer
treatments respectively. In concern with the effect of the tested oils on brood rearing
activity, Anise and Cinnamon oils at 30% concentration were the best (74.54 and
74.35 %. respectively) during January – February. However, in significant differences
were found between the tested oils at 30% concentration during August–September.
Thymol was the best at 15 % concentration ( 55.38%).However, Cinnamon oil 30%
was the best (64.54%) followed by Thymol(56.52%) .
Keywords :Varroa destructor ,Apismellifera ,essential oils.
INTRODUCTION
Varroa destructor mite has become a serious pest of Apis mellifera
all over the world ( Baker and Peng,1995; Rashid et al.,2014).It cause serious
losses in apiculture of A. mellifera.It feeds on haemolymph of larvae, pupa
and adult bees during all life ( Anderson and Trueman, 2000). It also,
decrease brood, colony ability to pollinate plants ( De Jong at al.,1984).V.
destructor associated with the virus diseases, which six bee viruses were
detected in bee infested by varroa( Shimanukiet al., 1994; and Correa-
Marqueset al., 2003). Using of synthetics varroacides may due to increase of
varroa resistance to these products ( Thomsonetal., 2002). Recently,
beekeepers have used some essential oils as well as volatile compounds to
control varroa mites ( El-Shemy,1997 ; Allamet al.,2004;Sammataro et al.,
2004 and Ruffinengeet al., 2007). Studies have shown that higher
concentrations of formic acid can kill mites under capping (Calderon et al.
2000, Amrineet al. 2006).The present study aim to evaluate three essential
oils i. e.Thymol, Cinnamon and Anise with two concentrations(15 and 30%) in
El-Hady, A. M. et al.
236
controlling applications compared to formic acid to controlling varroa mites in
both capped brood and on adult bees in addition, the effects of these
treatments on brood rearing.
MATERIAS AND METHODS
Present study was confirmed in an apiary selected at Ayash El-
MehallaElKobra District, Gharbia Governorate in 2013 season. All colonies
had brood nests composed of one standard deep and one shallow hive body.
Each brood box contained 5 frames in winter season and 8 frames in summer
season, had approximately equal numbers of bees, and had a laying queen.
Evaluated materials
Essential oils in two concentrations (15 and 30%) were used in this
study to management varroa mites comparing to Formic acid 65% given in
Table (1). Colonies were divided into 7 groups, three colonies for each
concentration as well as 3 colonies as a control (without treatment) .Three
applications of each treatment were applied from 16 January to – February
and from 15 August to – September , 2013.A completely randomized design
was applied in this treatment.
Table.1. Essential oils and their concentrations.
Serial No. Name of acaricides Concentrations %
1 Thymol 15%& 30%
2 Anise oil 15%& 30%
3 Cinnamon oil 15%& 30%
4 Formic acid 65%
Application methods:
1- The mixture of Essential oil Thymol or Anise and Cinnamon was
prepared in two concentrations, 15 and 30% by mixing with olive oil
(15ml: 85ml and 30ml:70 ml, respectively) . Talcum powder was added to
each mixture at rate 1 oil: 2 powder) to make solid mix (3x3x1 cm)
weighted 30 gm. .Each mixture was placed on the top of combs, changed
every 10 days.
2- 120 ml of 65% Formic acid was kept in a corked plastic bottle with cotton
strip to regulate 4 mL evaporation / daily of Formic acid. The bottle was
adjacent to an empty space above the brood.
Assessment of bee infection
For bees, pre-treatment count of mites was taken before application.
About 100 adult bees/ colony were used to calculate varroa infestation using
alcohol wash technique (De Jong et al., 1982), Post-treatment Assessment
count of mites was taken after the end of each treatment application.
For brood, the mite infestation was evaluated by opening cells of sealing
brood before treatment and mite infestation was assessed in inch
2
, three
replicates / colony (Burgett and Burikam, 1985).
Efficacy of oils.
Effectiveness % of the tested treatments were determined according
to the formulae adopted by Liorente J-Martines(1989) as follow:
J. Plant Prot. and Path., Mansoura Univ., Vol.6 (1), January, 2015
237
Effectiveness in controlling Varroa mites %=
Final of fallen mites in treatment−Final of fallen mites in control
............×100
Final of fallen mites in treatment
Whoever, on brood rearing activity, an area of 5x5 cm in the middle
of a worker comb (sealed brood) was selected after treatment to determine
the effect of oils on brood rearing activity.
Effectiveness in brood rearing activity %=
Final of increasing in treatment −Final of increasing in control
....×100
Final of increasing in treatment
Statistical analysis.
Randomized complete block design were applied to this experiment
Mean mite mortality (fallen mites) and brood rearing activity were compared
by analysis of variance technique to test the significance of data using the
least significance test (LSD) at 5% probability level (Montgomery,
2001).Duncan analysis was used to compare between mean of fallen mites.
RESULTS AND DISCCUSION
Efficacy of some essential oils at different concentrations as well as
Formic acid 65%was evaluated for controlling of varroa mites during the
winter and summer seasons of 2013. Each treatment was applied three times
from the first of January to the half of February and from the half of August to
the half of September. Pre-treatment count of fallen mites was taken before
treatment. The pre-count of the beginning of fallen mites was higher in
summer than the winter season. It ranged from 21 to 29 infestation in winter
and from 39 to 47 in summer.
Data in Table (2) revealed that from January to February, the fallen
mites increased and the first application was the highest one. Thymol 30%
showed the highest fallen mites (225 mites) at 1
st
application, followed by 94
mites at the 2
nd
application then 50 mites at the 3
rd
application.. In case of
Cinnamon 30% recorded the second rank, it showed 103, 102 and 47 fallen
mites, respectively. Anise oil 30% came in the 3
rd
rank, the fallen mites were
155 mites at the 1
st
application, 48 mites at the 2
nd
application then 38 mites
at the 3
rd
application. Consequently, it could be ranking the essential oils
according to the total fallen mites in descending order as follow, Thymol 30%
(369 mites), Cinnamon oil 30% (252 mites) then Anise oil 30% ( 241 mites).
In concentration 15 5 of the essential oils, Anise oil came in the first rank (205
mites) followed by thymol (175 mites) then cinnamon oil (142 mites). Formic
acid 65% showed the highest total fallen mites (573 mites) compared to the
tested essential oils.
El-Hady, A. M. et al.
238
Table.2. Effectiveness of essential oils for controlling of Varroa mites
during January – February, 2013.
Pre-
treatment
count
1
st
Application
2
nd
Application
3
rd
Application
Treatment No. of
mites 2/1 – 14/1 15/1-26/1 28/1 – 10 / 2
Total
fallen
mites
Efficacy %
Thymol 15% 23 82e 59d 34d 175 72.57
Anise oil 15%
29 89de 78c 38d 205 76.58
Cinnamon oil15 %
24 61 37e 44c 142 66.19
Thymol 30% 21 225a 94b 50b 369 86.99
Anise oil 30%
25 155b 48de 38d 241 80.00
Cinnamon oil 30 %
24 103cd 102b 47bc 252 80.95
Formic acid 65%
22 108c 223a 242a 573 91.62
Control 21 16f 15f 17e 48
L.S.D at 5% 14.75 12.87 5.41
Data in Table (3) revealed that the total fallen mites as a result to use
the essential oils from August to September were more than those in from
January to February, 2013. The total fallen mites due to the use of essential
oils at 30% concentration were more than those at 15% concentration. On
contrast of winter season, Cinnamon oil 30% showed the highest total fallen
mites (684 mites) followed by Anise oil% (533 mites), It could be noticed that
Thymol 15 % concentration was more effective (375mites) than the
concentration 30% (447 mites). Also, Formic acid 655 gave the highest fallen
mites (976 mites) compared to the tested essential oils and it was more
effective in summer season than the winter season.
Table.3. Effect of different essential oils for controlling the management
of Varroa mites during August – September, 2013.
Pre-
treatment
count
1st
Application
2nd
Application
3
rd
Application
Treatment
No. of mites
17/8 – 26/8 27/8-6/9 7/9 – 15/9
Total
fallen
mites
Efficacy %
Thymol 15% 43 291a 107d 49d 375 67.46
Anise oil 15%
37 112c 110d 48d 270 54.81
Cinnamon
oil15% 39 147b 106d 35d 288 57.63
Thymol 30% 47 157b 157c 61cd 447 72.70
Anise oil 30%
41 293a 151c 89c 533 77.11
Cinnamon Oil
30% 45 314a 257b 113b 684 82.16
Formic acid
65% 42 288a 320a 368a 976 87.50
Control 38 28e 33e 61c 122
L.S.D at 5% 29.53 30.28 37.36
The effectiveness of different essential oils for controlling varroa
mites ranged between 66.19 to 86.99 % in winter (January – February) and
54.81 to 82.16% in summer (August to September). In the period from
January – February, Thymol oil 30% was the best (86.99% followed by
Cinnamon oil 30% (80.95%) then Anise oil 30% ( 80.00%). However, the
same oils at 15% were less effective. Floris, et al.(2004) stated that both
Thymol formulations reduced significantly the levels of mite infestations of
J. Plant Prot. and Path., Mansoura Univ., Vol.6 (1), January, 2015
239
adult bees and sealed brood and its efficacy, expressed as percentage of
mortality, was lower for both products (Apilife VAR 74.8 ± 13.1 and 81.3 ±
15.5, Apiguard 90.4 ± 8.3 and 95.5 ± 8.7 for sealed brood and adult bees,
respectively).
On the other hand, treatments from August to September showed
that Cinnamon oil 30% was the best and occupied the first rank (82.16 %
efficacy) in summer followed by Anise oil 30% (77.11%) then Thymol 30%
(67.46%). Similar results with different oils were suggested by Whittington et
al (2000) examined the botanical oils neem, thymol, and canola control of
parasitic mites (Varroa jacobsoni and Acarapiswoodi) in honey bee (Apis
mellifera) colonies in British Columbia, Canada. Neem oil spray (5% solution)
killed 90±6% of varroa mites, 3 times more than that died in the untreated
group. Thymol-oil spray (4.8g Thymol/litre in 20% canola oil solution), thymol
in vermiculite (3.6g/vermiculite block), and canola oil spray (20% solution)
killed 79±8%, 68±6%, and 65±6% of the varroa mites, respectively. Colonies
treated with the Thymol-oil spray had a significantly lower tracheal mite
infestation (1.3±7.5%) at the end of the treatment period than the untreated
group (23.3±6.0%). All other treatments showed a slight but not statistically
significant decrease in tracheal mite infestation levels. Also, Akyol and
Yeninar(2008) reported that average efficacies of Thymovar and Bee Vital
were found to be 96.91 and 88.66%, respectively. El-Zemityet al.(2006) who
found that clove oil gave good results in controlling varroa mites. Fouly and
Al-Dehhairi (2009) stated that clove showed 62% efficacy in controlling varroa
mites. Hussein et al.(2001) tested the 6 plants oils to control varroa mites and
rated clove oil, the pest of among the other oils.
As far as the timing of application, the number of fallen mites was
higher after the 1
st
application than the 2
nd
and 3
rd
applications either in winter
or summer treatments. These results are similar with other findings and
treatments (Shoreit and Hussein,1994) who found that the maximum fallen
mites was happen after the first treatment. Abdel Rahman and Rateb (2008)
suggested that the mean number of fallen mites were higher after 24 h., than
after 48 h and after 72 which showed the lowest numbers. Also, Rashid et
al.(2014) who stated that the mean number of fallen mites were higher after
24 h of treatment.
In case of Formic acid was more effective than the tested oils, which
showed 91.62% in January to February and 87.50% in August to September.
Also, the mean number of fallen varroamites increased gradually with timing
application. The number of fallen mites was lower in the first treatment than
the second and third treatment either in winter or summer application. Similar
results were obtained by Rashid et al.(2011) who stated that formic acid
significantly killed higher number mites compared to thymol . Also, Calderon
et al. (2014) found that formic acid 86% gave effectiveness in controlling
varroa mites 96.9%, while Thymol released 94.7%.
Effect of essential oils on brood rearing activity.
Data in Table (4) showed that both of Cinnamon and Anise oils
treatments increased brood rearing activities percentage than Thymol in the
two concentrations 15 and 30% compared to formic acid and the control
El-Hady, A. M. et al.
240
treatments. No significant differences were found between Aniseoil,
Cinnamon oil at 15 % and Thymol at 30% concentrations. Thymol oil 15 %
recorded the lowest increase in brood rearing (200.99 inch
2
) and efficacy%
44.28%. No significant differences were found between the Anise and
Cinnamon at 30% concentration (439.99 and 436.65 inch
2
). All the tested oils
increased brood rearing compared to Formic acid( 297.99 inch
2
)and control
treatment (111.99 inch
2
)
Table.4. Effectiveness of essential oils on brood rearing activity during
January – February, 2013.
1st
Application
2
nd
Application
3rd
Application
Treatment 2/1 – 14/1 15/1-26/1 28/1 – 10 / 2
Brood
rearing
(Inch2)
Efficacy %
Thymol 15% 43 72.66 85.33 200.99d 44.28
Anise oil 15% 121.66 130.33 136.33 388.32b 71.16
Cinnamon oil 15 %
121 138 160 385.88b 70.97
Thymol 30% 103.33 132.66 155.33 391.32b 71.38
Anise oil 30% 128.33 144.66 167 439.99 a 74.54
Cinnamon oil 30 %
131.66 145.66 159.33 436.65a 74.35
Formic acid 65% 98.33 110 89.66 297.99c 62.41
Control 36.66 37 38.33 111.99e
L.S.D at 5% 41.28
Data in Table (5) showed contrast results during January – February,
Out of concentration 15%, Thymol 15% ranked the first in increasing brood
rearing activity(55.38%).However, Cinnamon oil 30% and Thymol 30 and15
% were the best 64.54, and 56.32 %, respectively. It could be noticed that
Formic acid showed lower efficiency in summer (40.18%) than those in
summer months (62.41%) compared the control treatment. No significant
differences were found between the tested oils at 30% concentration.
Table.5. Effectiveness of essential oils on brood rearing activity during
August – September, 2013.
1
st
Application
2
nd
Application
3
rd
Application
Treatment 2/1 – 14/1 15/1-26/1 28/1 – 10 / 2
Brood
rearing
(Inch
2
Efficacy%
Thymol 15% 148.33 181 203.33 532.66b 55.38
Anise oil 15% 81.66 100.66 117.33 299.65f 20.68
Cinnamon oil
15 % 110 116.33 132.33 358.66e 33.73
Thymol 30% 166.66 182.66 197.33 546.65b 56.52
Anise oil 30% 146.66 168 126 440.66c 46.06
Cinnamon oil
30 % 193.33 220 257 670.33a 64.54
Formic acid
65% 126 141 130.33 397.33d 40.18
Control 133.33 131 123.33 237.66d
L.S.D at 5% 28.72
J. Plant Prot. and Path., Mansoura Univ., Vol.6 (1), January, 2015
241
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