American-Eurasian Journal of Scientific Research 11 (5): 320-331, 2016
ISSN 1818-6785
© IDOSI Publications, 2016
DOI: 10.5829/idosi.aejsr.2016.11.5.10418
Corresponding Author: Shimelis Mengistu, Haramaya University, College of Veterinary Medicine,
P.O.Box: 138 Dire Dawa, Ethiopia.
320
Major Honey Bee Health Problem with Particular Emphasis to
Anti-Varroa Investigation of Propolis in Toke-Kutaye District, Ethiopia
Shimelis Mengistu, Yared Kebede and Desalegn Begna 1 12
Haramaya University, College of Veterinary Medicine, P.O.Box: 138 Dire Dawa, Ethiopia 1
Holleta Bee Research Center, P.O.Box 22, Holeta, Ethiopia 2
Abstract: A cross sectional study was carried out from November 2014 to March 2015 for problems associated
with Major Honey Bee Health problem in Toke-kutaye, District, West showa Oromia Regional State, Ethiopia
with Particular Emphasis to Anti-varroa Investigation of Propolis. Purposive sampling was used in each kebele
and a total of 40 beekeepers out off 5 kebeles were selected and Sampling was split proportionally and varies
with the number of hives available between each beekeeper apiary site. Direct observation, inspection and
laboratory examination was the main data collection techniques. Executing petr-dish bio assay set up were the
main data collection techniques used to gather the information of various concentrations of propolis extracted
in 55% ethanol. Laboratory examination results revealed that all diagnosed kebeles had varroa mite with
infestation rate ranging from 80% to 92.3% and the presence of Nosema 39% and amoeba 62.3% in honey bees
in 82 colonies tested out of 5 studied kebeles. All inspected hive had pests like wax moth, 82(61%) and spider
(Lactrodectus mactan), 82 (59%). Antivarroa investigation of propolis as through bio assay revealed that the
length of narcosis and rate of mortality of varroa dependence on the concentration of propolis used, the
duration of contact time and the origins of propolis. Thus, treatment with a 20%, propolis solution in
55%ethanol resulted in 100% mortality rate at a contact time of 5s regardless of the origins of the propolis.
However, treatment with 5% of propolis (Eastern hararge) narcosis lasted from 37.8 ± 4.41% to 30 ± 5% at 5s
and 10s contact times respectively and 100% mortal at 20s contact time. Further research in this field needs to
be encouraged, it is therefore very important that the existing problems are well managed to maintain bee health
and that the risks and consequences of pests and diseases are well understood and appropriate plans in place
to deal with any such honey bee health problem. This will help to sustain the health of honey bee already
established in the bee keeper’s apiary in the district.
Key words: Antivarroa Honey Bee Narcosis Propolis Varroa mite
INTRODUCTION Large and diverse botanical resources combined
Beekeeping is an important component of agriculture conducive for beekeeping business [4,5] and the
and rural development program in many countries and country is the leading honey producer in Africa and
useful small-scale efforts have been made to encourage is one of the ten largest honey producing countries
beekeeping interventions throughout the world [1]. It in the world [6, 7]. The most important species for bee
plays a role in providing nutritional, economic and keeping in East Africa and the wider sub Saharan Africa
ecological security. The business almost requires no land, is the Apis mellifera scutellata (the African Honey Bee)
capital and does not take much part of the farmers’ time [8].
and does not compete with other components of farming Able to survive in a wide range of climates and
systems for resource. Directly, it contributes in the values environments, honey bees are among the most successful
of the outputs produced, including honey, bee wax, queen organisms on the planet [9]. And now inhabit most areas
and bee colonies and other products such as pollen, royal of the world occupied by humans. Accompanying the
jelly, bee venom and propolis in cosmetics and medicine spread of bees globally has been a host of pathogens,
[2,3]. parasites, pests and viruses. Of these organisms, such as
with suitable climatic conditions make Ethiopia
Am-Euras. J. Sci. Res., 11 (5): 320-331, 2016
321
Nosema apis only just beginning to be understood, while Therefore the main objective of the study was to
Varroa destructor despite being well known, remain at verify major honeybee health problems of beekeeping in
forefront of bee research [10]. Toke-kutaye woreda with major emphasis to determine
The varroa mite is a parasite with the most antivarroa actions propolis collected from different
pronounced economic impact on beekeeping industry geographic origins of the country.
[11]. It is a major pest of the honeybee, spreads very
quickly and causes serious damage to its host colonies Objectives
where it reproduces inside the capped brood cells [11,12]. Main Objectives:
Two microsporidian species infect honey bees
worldwide: Nosema apis and Nosema. ceranae. The term To identify the major honeybee health problems in
nosemosis is considered to be the infection of ventricular tokke kutaye districts.
cells of adult honey bees by Nosema. apis and reduces To determine antivarroa actions propolis collected
worker longevity by 22-44% [13]. Nosema apis are the from different geographic origins of the country.
causative agent of nosemosis in honeybees (Apis
mellifera). This disease is widespread and found in every Specific Objectives:
beekeeping country [14,15]. The world trade in honey-bee
products and beekeeping materials may play an important To identify and verify the infestation rate of varroa
role in the dispersal of infective spores of N. ceranae from mite, Nosema and amoeba
apiary to apiary over different geographical areas [16]. To verify the infestation rate of pests through hive
Despite the diversity of infectious diseases and their inspection in Tokke Kutaye District.
agents that can cause bee mortality, surveillance has been To assess type of propolis caused narcotizing and
fragmented and thus it has been difficult to gain a clear effect of different concentration (%w/v) of propolis
historical view of bee health [17]. Symptoms seen in a on varroa mite.
colony vary according to the type of organism causing
the disease. Many of these symptoms are none specific MATERIALS AND METHODES
and so the only way of confirming pathogen is through
appropriate laboratory diagnostics. Their severity largely Study Area Description: The study was conducted in
depends upon the vigor of the affected colony [12]. Even Toke Kutaye districts, West Shewa Zone of the Oromia
though varroosis once established in the apiary it Regional State and Investigation of antivarroa actions of
destroyes colonies within 2 to 5 unless treated [18]. Since propolis was conducted in Holeta bee research center
forces beekeepers around the world to treat their colonies located in Holota, Oromia Special Zone around Finfine,
with different types of acaricides which applied to Ethiopia. The Toke Kutaye woreda has 32 rural kebeles
minimize mite infestation rate and subsequently reduce or and 4 administrative towns. This woreda located at 126 km
prevent colony death. However, these acaricides to the west from the capital city Addis- Ababa and
applications led to emergency of resistant mites [19] [20] geographically located at 8° 58’N, 37° 46’E and altitude of
and undesirable residue problems [21,22]. ranging from 1380m to 3194m above sea level. The area
Therefore diagnostic surveys are conducted in receives a mean annual rainfall of 900 mm (800 to 1000 mm)
different parts of the country at different times by various and annual temperature ranging from 15 to 29°C with
investigators on honey bee health problem. Similarly, the average temperature of 22°C. West Shewa zone is
works of laboratory analysis reported two adult honeybee generally a highland whose topography gave the area a
diseases nosema and amoeba and determined their characteristic climate that is conducive for the apiculture.
distribution and the work of investigation of propolis on Toke Kutaye woreda has number of livestock on the
varroa mite. According to the damage caused by Varroa bases of species as 13 7180 cattle; 46,556 sheep;
mite, Nosema and amoeba and pests on honeybee 22,611 goats; 79,504 poultry; 26792 equines and
colonies, the recent statistics about varroa infestation, 17,269 bee hives (Toke kutaye woreda livestock and
epidemiological studies in variant geographical regions of agricultural offices, 2013).
beekeeping provides set information on major findings in The Holota town is located 25 km west of Addis
relation to the works of diagnostic survey and Ababa situated at latitude of 9°3’ N and longitude of
characterizations of major honeybee health problems in 38°30’ E. The area has annual rain fall and daily
Tokke Kutaye Districts and investigation of anti-varroa temperature, which ranges from 834 mm to 1300 mm and
action of propolis. 5°C to 28°C, respectively.
Am-Euras. J. Sci. Res., 11 (5): 320-331, 2016
322
Study Population: The study populations were honey bee 70% ethanol and from brood comb capped brood cells
colonies from purposely selected 5 kebeles of the district (internally) labeled accordingly. Number and Type of hive,
and propolis from diverse geographical origins. type of sampled hive, presence of wax moth and spider
Sample Size Determination: The five districts were transported to Holeta bee research center laboratory for
selected for the study following purposive sampling infestation rate of varroa mite and about 30 adult honey
approach considering potential in honey bee production. bee from collected adult bee were diagnosed for the
A multi stage sampling technique was used in this study. detection of amoeba and nosema spore which represents
In the first and second stage, honey bee potential kebeles sampled colonies.
and 8 beekeepers from each kebeles and total of 40
registered beekeepers from livestock and agricultural Examination Procedure: In the laboratory, preserved
office of woreda have been identified purposively and in worker and brood were examined.
the third stage, sample size of 82 honeybee colonies was
set for this study using quota method. Adult Examination: In the laboratory, preserved bee
Study Design: Cross sectional study (diagnostic survey) into each of the jar containing bees up to half of the jar.
and anti-varroa investigation of propolis was carried out Then the jar was shaken for 1 minute, until the mite
from November 2014- March 2015. The honey bee separated from honey bees. The solution strained through
colonies of study area were sampled for laboratory a ladle (8-12 mesh) to remove the bees and then sieving
investigation for their potential health problems and the the solution through tea strainer. Finally, the tea strainer
colonies were inspected in connection with factors that was examined and counted for varroa mite.
potentially affect bee health and the anti-varroa actions of
propolis with different levels of ethanol extracted, anti- Brood Examination: The mites themselves should be
varroal property of propolis from different agro ecological sought for confirmation, by examining the bottom of the
zones and type, level of extracts and time of efficacy cell and the brood for attached mites. Brood examinations
identified. were done by random opening of 200 to 300 brood cells.
Sampling Methods: Livestock Development and Health and the cell was inspected for the presence of mite using
District agencies of Toke-Kataye Woreda were consulted by naked eye.
to selected sampling site. Accordingly 5 sampling sites
and 40 beekeepers having bee colonies were selected Microscopy: A laboratory method consists of the
purposely and honey bee colonies randomly sampled and individual examination of the colonies for the
examined from selected kebele. Proportional sampling simultaneous detection of Nosema spores and Amoeba
method was used to fix sample size for each beekeeper cysts using 30 bees per colony was employed
and the diagnosis and examination was conducted on The abdomens of the bees to be examined were
82 randomly drown brood and adult honey bee separated and ground/grind up in 2 ml of water. Three
represented honey bee colonies of selected apiary site. drops of the suspension were placed on a slide under a
Propolis samples used in experiments were obtained from cover-slip and examined microscopically at ×400
the Holota Bee Research Center which was collected from magnification, under bright-field or phase-contrast optics.
beehives of different areas of the country.
Study Methodology: The colonies were examined for pests used in experiments were obtained from beehives in the
according to OIE [23] methods and disease diagnosis was colonies in the apiary site of Bako, Eastern harage
carried out according to Shimanuki and Knox [24] on the and Toke kutaye woreda and extracted and investigated
colonies and broods to verify the levels of infestation rate in Holeta Bee research Center. In preparation for
with varroa mite, presence of bee lice, Amoeba and extraction, weighed and frozen propolis samples were
Nosema. homogenized using a coffee mill. Samples were frozen
About 200-300 bees and brood cape cells were before homogenization since unfrozen propolis, due to its
collected from each selected hive in a jar, using bee brush sticky nature, does not lend itself to easy homogenization.
from hive entrance (externally), killed and preserved in The homogenate powder was then suspended in 70 %
were recorded corresponding to the label and then
samples were taken and a detergent solution was poured
The brood was removed from the cell with a fine forceps
Propolis Extraction and Preparation: Propolis samples
Am-Euras. J. Sci. Res., 11 (5): 320-331, 2016
323
ethanol, in a ratio of 1:9 (w/v) for effective extraction [25] the tissue paper using micropipette. The treatment was
and extracted in a rotary evaporator at 60 degrees Celsius stopped after the allocated time by removing the mites
for 2 h. After the allocated extraction time, the suspension from the Petri dish and Control experiments for each
was cooled at room temperature for about 1 h and suction experimental group were run by treating the mites, for the
filtered. corresponding treatment times, with 55% ethanol solution
The filtrate was dried in an incubator at 40°C to and also with distilled water.
weight constancy, which was achieved in 2 weeks time. An individual was considered dead if it showed no
The dried 70% ethanol extract was dissolved in 55% leg movement and/or movement of any other body part
ethanol to make a 10% (w/v) stock solution of propolis. when gently prodded with a probe. If it showed
Even though 70% ethanol was used for extraction movement, whether it was partially paralyzed or normal,
purpose, 55% ethanol was employed as a solvent in the was counted as alive. Each treatment was repeated three
bioassay in order to reduce the effect of strong ethanol times and the mean ± S.D values were used in the
solution on the experimental organisms. The little amount presentation of results.
of precipitation observed while suspending the 70%
extract in 55% ethanol was brought into solution by Data to Be Collected: Proportion of narcotized mites just
agitation. The stock solutions were stored in a refrigerator after treatment, Time at which the treated mites narcotized,
for later use. Effect of different concentrations (%w/v) of propolis,
Different concentrations of propolis were prepared Type of propolis caused effective narcotizing
for the bioassay by diluting the stock solutions with
ethanol of the concentration used to prepare the Data Management and Analysis: Data analysis was
corresponding stock solution. conducted using SPSS version 20. All collected data was
Mite Collection: Mites were collected from infested statistics such as mean, standard error; frequency,
colonies in the apiary site of Holeta Bee research Center. percentages were used to analyses the data using SPSS
Infested combs or pieces of combs containing drone (version 20). Chi-square (÷2) was used to determine the
broods were brought to the laboratory and Varroa mite statistical association between factors. The statistically
were collected from capped cells by opening and significant associations between variables were
inspecting individual cells. In order to avoid starvation of considered if the calculated p-value was less than 0.05
the mites during the collection process collected mites with 95% confidence level.
were kept in a Petri dish on bee larvae or pupae. Collection
of mites was done from both the larval and pupal stage of RESULTS
healthy brood, since mites at the different developmental
stages of the brood did not show any significant Adult Bee Diagnosis: This study showed that varroa mite
difference in their sensitivity for pyrethroids in a infestations were found in 70 (85%) out of 82 sampled
laboratory bioassay [20]. Newly moulted adult mites, colony of studied area. From the total 82 bee colonies
identified by their pale colour, relatively smaller size and diagnosed, varroa mites were detected in 70 with overall
weak locomotion activities were excluded from the infestation rate of 85%. An average of 262 ± 22 bees per
experiment since they may have a different response as colony was examined through adult bee colonies and an
hardening of the cuticle is still in progress. Mites, which average of 7 ± 6.7 (ranging from 0-14) varroa mites were
seemed physically weak and abnormal and also those recovered. Although there is no apparent difference,
obtained from diseased brood were discarded. infestation rates were high for Melka-nega-denbi 92.3%,
Petri Dish Bioassay: In order to observe the effect of and Negafile 80% with decreasing infestation rate.
contact time of different propolis concentrations on the However, per colony recovered the average number of
mortality rate [26] of V. destructor mites, six mites per varroa mites was high for Melka-nega-denbi and Negafile
experiment were placed on a 3 cm x 3 cm single layer tissue and low for Lenca (Table 1). The mite infestation cannot
paper in a Petri dish. The mites were treated by applying be established whether due to poor hive management or
250 micro liters of the concentrations to be used in the climatic factors or related to the geographical location of
bioassay: 5%, 7.5%, 10%, 15%, 20% (w/v) of propolis on the apiaries.
entered into Microsoft Excel 2007 and descriptive
Himela-dawe-ajo 86.9%, Toke-meti 84.2%, Lenca 81.82%
Am-Euras. J. Sci. Res., 11 (5): 320-331, 2016
324
Table 1: Adult bees sampling kebele with percentage bee colonies tested positive for varroa mites.
Total number of Number found positive Average number of Average number of
Sampled area bee colonies sampled to varroa mite % infestation bees sampled per colony varroa recovered per colony
Negafile 15 12 80 255 11
Melka-nega-denbi 14 13 92.3 265 11
Himela-dawe-ajo 23 20 86.9 261 4
Toke-meti 19 16 84.2 264 6
Lenca 11 9 81.82 266 3
Total 82 70 425.22 1311 35
Average 14 85.044 262.00 7
STDEV .356 4.421 22.831 6.740
Table 2: Total number of sampled bee colonies per kebele and obtained results through
Total number of Number found positive Average number of Average number of
Sampled area bee colonies sampled to varroa mite % infestation bees sampled per colony varroa recovered per colony
Negafile 15 12 80 255 11
Melka-nega-denbi 14 13 92.3 265 11
Himela-dawe-ajo 23 20 86.9 261 4
Toke-meti 19 16 84.2 264 6
Lenca 11 9 81.82 266 3
Total 82 70 425.22 1311 35
Average 14 85.044 262.00 7
STDEV .356 4.421 22.831 6.740
Table 3: Total number of sampled bee colonies per kebele and obtained results through microscopy diagnosis
Number of Number positive %of Nosemosis Number positive %of Amoeba
Sampled area samples colonies to Nosema spore within site to Amoeba cyst within site
Negafile 15 6 40.0% 9 60.0%
Melka-nega-denbi 14 5 35.7% 11 78.6%
Himela-dawe-ajo 23 9 39.1% 14 60.9%
Toke-meti 19 8 42.1% 10 52.6%
Lenca 11 4 36.4% 7 63.6%
Total 82 32 51 317
Average 39.0% 62.2%
df 4 4
P .996 .676
Brood Bee Diagnosis: From the total of 82 bee The average mite load per brood recovered range
colonies diagnosed for brood, 70 bee colonies accounting from 9 to 31 which was highest in and low in Lenca
85% of the diagnosed brood were tested positive for respectively (Table 2).
varroa mites. The result from the diagnosed brood
showed that the sampling localities were infested with Microscopy Examination and Inspection: Nosema spores
varroa mites with the infestation rate ranging from 80% to were found in five sampled sites with a total of 82 (39%)
92.3%. On the average 16 ± 8.320 bee colonies were colonies tested and Amoeba cyst were found with total of
sampled for brood diagnosis from each kebele and an 82(62.2%) colonies tested (Table 3). The %of Nosema
average of 14 bee colonies were found positive to varroa spores within sites in the honeybee colonies range from
mite. Likewise, on the average 262 ± 22.154 brood cells 36% to 42% and Amoeba range from 53% to 79%
were opened from each bee colony with an average of (Table 3).
16.15 ± 19.946 varroa mite detection. Through the brood Based on colony status of diagnosed honey bee
analysis, Melka-nega-denbi and Negafile kebele were colonies within kebele, the highest nosemosis 42.1%
found with high and low varroa mite infestation rates observed in Himela-dawe-ajo while highest amoeba
respectively (Table 2). infection 78.6% was observed in Melka-nega-denbi.
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325
Table 4: Total number of sampled bee colonies per kebele and obtained results through colony inspections
Number of Number wax
Sampled area sampled colonies moth present % of wax moth Number spider present % of spider
Negafile 15 10 66.7% 10 66.7%
Melka-nega-denbi 14 10 71.4% 9 64.3%
Himela-dawe-ajo 23 13 56.5% 14 60.9%
Toke-meti 19 12 63.2% 11 57.9%
Lenca 11 5 45.5% 5 45.5%
Total 82 50 303.44 49 295.51
Average 61.0% 59.8%
df 4 4
P .722 .854
Table 5: Percentage reduction in the mean and mortality rate of V. destructor mites by different propolis concentrations at each contact of treatment time
represented by mean ± S: D: values.
Contact time
Propolis concentration (%, w/v) —————————————————————————————————————————————————-
In 55% ethanol (30s treatment) T5s T10s T20s T30s T40 T60s T75s T90s
5% Mean 3.78 3.00 2.11 1.11 .22 .00 .00 .00
Std. Deviation .441 .500 .928 1.054 .667 .000 .000 .000
7.5% Mean 3.33 3.00 2.00 .67 .00 .00 .00 .00
Std. Deviation .577 .000 .000 1.155 .000 .000 .000 .000
10% Mean 3.00 2.00 1.33 .00 .00 .00 .00 .00
Std. Deviation .000 .000 1.155 .000 .000 .000 .000 .000
15% Mean 2.50 2.00 .00 .00 .00 .00 .00 .00
Std. Deviation .548 .000 .000 .000 .000 .000 .000 .000
20% Mean 2.33 .00 .00 .00 .00 .00 .00 .00
Std. Deviation .577 .000 .000 .000 .000 .000 .000 .000
control Mean 1.00 1.00 1.00 1.00 1.00 1.00 1.00 1.00
Std. Deviation .000 .000 .000 .000 .000 .000 .000 .000
Total Mean 2.70 2.00 1.17 .60 .27 .20 .20 .20
Std. Deviation 1.088 1.050 1.053 .814 .521 .407 .407 .407
There were no significant differences in the Proportion of Narcotized Mites Just after Treatment:
infection from colony to colony, site to site in Comparison of the mean and mortality rate, from the Petri
nosema and amoeba infection rate (Pearson Chi- dish assay experiment displayed propolis concentrations
Square =.179, df = 4, P =.996 (P > 0.05) and (Pearson Chi- that show considerable effects on the mortality rate (Table
Square = 2.394, df = 4, P =.676 (P > 0.05) respectively 1). The reduction in the mean due to treatment with a
(Table 3). certain propolis concentration was with increasing
A total of 82 colonies examined were infested propolis concentrations where the mite was found out to
with Wax moth and Spider (Lactrodectus mactan) be more sensitive well as the reduction in the movement
(Table 4). Again, 61% colonies were infested with of mites and mortality achieved except for the experiments,
Wax moth and 59% colonies were infested with where the mite was found to be recovered (Table 5).
spider (Table 4). When infestation of wax moth was The effect of contact time of different propolis
calculated based on colony status of inspected hive concentrations on the mortality rate of varroa mites as can
per kebele, the highest wax moth (71.4%) was be seen from Fig. 1, mortality rate increased with
observed in Melka-nega-denbi, while the spider increasing contact time for concentrations of 5, 7.5, 10, 15
66.7(%) was observed in Negafile. In contrary lowest and 20% in 55% ethanol. In the shortest contact time, for
Wax moth 45.5% and spider 45.5% was observed in 5 with 20% propolis in 55% ethanol resulted in 100%
Lenca (Table 4). However, the difference in infestation of mortality. In the case of different propolis concentrations
wax moth and spider was not statistically significant in 55% ethanol, mortality rate, even though it was weak
among bees of different colony status of sampled site seemed to increase with increasing contact time and
(P>0.05). concentration.
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326
Fig. 1: Effect of contact time of different propolis concentrations (% w/v) in 55% ethanol on the narcotic and lethality
of varroa mites: six mites per experiment, n= 3: the bar indicate series 0 = 5%: series 1 = 7.5%: series 2 = 10%:
series 3 = 15%: series 4 = 20% concentration of propolis: Y-axis= 0—1 low movement; 2 no movement; or lethal,
3=medium movement and 4= high movement: X-axis= Treatment time
Fig. 2: Antivarroa actions of propolis from different geographic origins and the effect of contact time of different propolis
concentrations on the mortality rate of Varroa mites: Y-axis= Narcotic and letal effect of propolis=degree of
movment: X-axis= site; 1-5= Bako: 6&7= Eastern hararge: 8= Toke-Kutaye: 9&10= control: The bar indicate the
time of propolis treatment (contact time of the propolis).
However, a contact time of 5, 10, 20, 30 and 40 Treatment with various propolis concentration of
seconds resulted in a mortality in case of 20, 15, 10, 7.5 Bako at different contact times showed that both narcosis
and 5 % propolis respectively regardless of propolis immediately after treatment and lethality was achieved
origins. Even though all propolis concentration in 55% after treatment regardless of both propolis concentration
ethanol was used in these investigations after contact and contact time. Further observation of the activity of
time of 60seconds no movement of the mite was observed the mites displayed that with 5% propolis concentration
(Fig 1). of solution Bako narcosis lasted longer 5s to 30s after this
Antivarroa actions of propolis from different time it was very low movement and at 40s no movement of
geographic origins and the effect of contact time of the mites were observed and no mites were recovered
different propolis concentrations on the mortality rate of after the allocated period of time.
Varroa mites. Antivarroa actions of propolis from Bako Treatment of Varroa mites with 5% propolis
were investigated at of 5, 7.5, 10, 15 and 20%, Eastern- concentration of solution East hararge the proportion of
harage 5 and 15% and whereas propolis from Toke-kutaye narcotized mites just after treatment ranged from 37.8 ±
at 5% propolis concentrations in 55% ethanol were 4.41% to 30 ± 5% at and 10s contact times respectively
investigated at contact time of 5, 10, 20, 30, 40, 60, 75 and and 100% mortality at 20s. In case of 5% propolis
90 seconds (Fig 2). concentration of solution Bako the proportion of
Am-Euras. J. Sci. Res., 11 (5): 320-331, 2016
327
narcotized mites just after treatment from 37.8 ± 4.41% to established more 10 mites natural drop per day can cause
22 ± 6.67% at 5s and 40s contact times respectively and colony collapse [28].
100% mortality at 60s (Table 1 and Fig 2 and 3). On the other hand, the wide distribution of the
Treatment time played a role in case of treatments varroa mites in all the surveyed areas and in all inspected
with high concentration of propolis for both solutions bee colonies indicates longtime introduction [29,30].
(Bako and East hararge). Treatment of mites with Higher infestation rate of varroa mite observed in the
15%(w/v) propolis in 55% of solution East harage the apiary might be associated with contact among colonies.
proportion of narcotized mites just after treatment was 25 Colonies in the apiary found close to each other, hence,
± 5.48% at 5s and 100% lethal which was achieved at 10s. facilitate transmission of varroa mite among the colonies
Even though Treatment of mites with 15% (W/V) propolis through swarming and drifting [31]. Chance of bees in
in 55% solution of Bako narcotized mites just after apiary to visit same flower is higher than bees in
treatment were 25 ± 5.48% at 5s and 100% mortality were backyard. However, the causes of variations in infestation
achieved at 10s as there was no activity of mites observed rates among the studied districts, it cannot be established
after this contact time (Table 1 and Fig 2 and 3). whether the infestation was due to poor hive management
Treatment of Varroa mites with 5% propolis or climatic factors or related to the geographical location
concentration solution of Toke-kutaye proportion of and service level of the places as bee colony marketing
narcotized mites range from 37.8 ± 4.41% to 30 ± 5% at 5s points. On the other hand, it is not certain how and when
and 10s contact times respectively and 100% mortality at these mites invaded the honeybee colonies of Apis
20s. resulted in an initial narcosis at 5s which was seems mellifera in the country especially in the districts where
similar in comparison with that of solution Bako but mites were found.
narcosis lasted within the first 10s of contact time and The presence of Varroa destructor, a parasitic mite
mortality achieved at 20s exactly as of East harage of the could major honey bee health problem cause the decline
same concentration 5% (Table 1 and Fig 2 and 3). in colony establishment and are a major problem for kept
Treatments with higher concentration of propolis bees in apiaries [32-35].
(20% (W/V) propolis) in 55% of Bako resulted 100% In the present study Nosema spores were found in
mortality, regardless of contact time, indicating its high five sampled sites with a total of 39% infestation rate
toxicity with the slightest contact time, as low as 5s observed in the current study was much lower than other
(Fig 2:5). previous reports in Ethiopia. Diagnosis made on 152
The control treatments of the experiment solutions of colony bees in field and laboratory at Addis Abeba
distilled water and 55% ethanol have not shown lethality reported prevalence rate of 53.3% [36]. nosema was also
effects after treatment of specified contact time all the mite reported from different regions with varying prevalence
was recovered after 90s (Fig 2; 9:10). The control mites of rate such as 58% in Oromia, 60% Benishangul-Gumuz and
both solutions after treatment their movement activity 47% in Amhara regions [37]. However higher than when
were not affected. Hence the experimental control used compared with survey conducted in Ethiopia regional
(distilled water and 55% ethanol) did not show in lethality state of Oromia, Amhara, Southern Nations and
effect with increasing contact time (Fig 2; 9:10). Nationality and Peoples (SNNP), Tigray, Gambella,
DISCUSSIONS [38]. While in the present diagnostic survey of Amoeba
All the surveyed areas tested positive to varroa mite is unequal when compared with diagnosis made on honey
with infection levels ranging 80 % to 92.3%. Which was bees in field and laboratory at Addis Abeba which
concise much greater with recent reported in Ethiopia by reported a prevalence rate of 73% of amoeba infestation
Begna [27] reported that varroa mite with infestation level [36]. it was also lower than Diagnosis made in Ethiopia
ranging 37.5 % to 100 % in Tigray Region, Ethiopia. regional state such as; Oromia region with prevalence rate
The average number of varroa mite recovered from a (88%), Amhara region (95%) and Benishangul- Gumuz
single bee colony through adults and brood’s analyses with 60% [37], this suggest the unequal distribution of the
were 7 and 16 mites, respectively observed in the current disease. Study on annual cycle and seasonal dynamics of
study suggesting Varroa mites have led to the virtual amoeba from the Holeta research center apiary [39]
elimination of feral bee colonies and the honey bees are reported, amoeba cysts were reported throughout the year
close to collapse concurring with the finding that regardless of hive type.
Benishangul–gumuz, Somale with 37.3% of infection rate
cyst were found with total of 51(62.2%) colonies tested it
Am-Euras. J. Sci. Res., 11 (5): 320-331, 2016
328
The infestation of Nosema spore and amoeba cyst destructor mites that the length of narcosis and mortality
observed in the area pointed out that the late detections rate depended on propolis concentration, solvent of
of adult honeybee diseases Nosema and Amoeba in 1989 extraction and length of contact time.
in the country and the follow-up investigations on its As seen from the present experimental results,
wide distribution suggests, less attention given to treatment of mites with 5% of propolis (from Easternmonitoring of local honeybee diseases [38]. hararge) and (from Toke-kutaye) in 55% ethanol (w/v %)
The presence of pests on the colonies confirmed that concentration was more effective than the corresponding
pest infestation is a major honey bee health problem in treatment with 5% of propolis (from Bako). The most
beekeeping in the tropics [40-43]. The pests are plausible explanation for these differences that origin of
responsible for the destruction of the colony and decline propolis from diverse agro-ecolological zones affected by
in its establishment [40]. temperature. Most of the biological active hydrophopic
The wax moth that is one of the most important pests components of propolis might differ from area to area.
of honeybee colony with worldwide distributions is also This means that propolis from Eastern-harage and Tokeidentified as one of the serious local honeybee pest in kutaye was quantitate component of biological soluble
the country [44]. The infestation of wax moth in the active ingredient possibly higher than propolis from Bako.
studied district 61% was higher than the previous studies Even though propolis in general and most of its
The study that was conducted in three zones of the components in particular, are water insoluble. The water
country investigated wax moth prevalence variations from soluble components of propolis comprise about 2.5–6.5%
zone to zones with South west Shoa zone having high only depending on the origin of propolis [47]. This is also
infestation level (26.66%) followed by West and East shoa possible explanation why propolis does not kill Varroa
zones 22.85% and 26.66%, respectively [45]. mites in the beehive while mite walking on thin propolis
Also, the same study indicated that about 56%-75% layers throughout the hive in the beehive’s interior. Since
of the wax moth infected honeybee colonies absconded most of its components in particular are water insoluble in
and the remaining dwindled. So this pointed out the the beehive’s interior [47]. The mortality rate of Varroa
presence of wax moth one of major honeybee’s health mites with propolis, as seen from present result depended
problem in the studied localities district. In the on concentration, contact time and origins of propolis.
considerations of its widely distributions and serious The 20% Propolis extracted in 55% ethanol was superior
effects, practical experiment was designed and identified to the 5% Propolis extracted in 55% ethanol. The 5s
effective preventive and/or control management practice contact time of Varroa mites with 20% propolis in 55%
[46]. According to the results of this experiment, 82.3% ethanol resulted in 100% mortality indicating that it is
effective measures that restrain the pest entry into highly toxic. In the case of treatments with propolis 5, 7.5,
beehives have been developed. The study recommended 10 and 15 % propolis in 55% ethanol also the mortality
management techniques of strengthening honeybee rate of Varroa mites rose with increasing contact time. But
colonies via feeding, removing unoccupied suppers and in comparison of mortality rate of varroa mite with
combs, combination of these practices and trapping adult experimental work of Garedewa et al. [26] the treatment
wax. with 10% propolis in 55% ethanol resulted in 100%
Proportion of narcotized mite just after treatment and mortality, regardless of contact time but in the case of
mortality rate and effect of different concentration of treatments with propolis 5 and 7.5% propolis in 55%
propolis from the Petri dish assay in this experimental ethanol the mortality rate Varroa mites rose with
study displayed propolis concentrations that showed increasing contact time seems in agreement with present
considerable effects on the mortality rate of varroa mites result. Hence length of mortality depended on propolis
and propolis from diverse geographical origins displayed concentration, solvent of extraction and length of contact
the differences in the antivarroal activities (Table 1, fig 3). time [26].
Accordingly proportion of narcotized mite just after The control experiments did not have lethal effects at
treatment decreases and mortality rate increased with all. This may show that, in order to detect the influence of
increasing contact time for propolis concentrations of 5, propolis concentrations of varroacidal agents on the
7.5, 10, 15 and 20% in 55% ethanol used in this experiment. narcotic and mortality of Varroa mites. One needs a higher
Remarkably propolis in previous experimental concentration of propolis to observe a remarkable
laboratory by the work of Garedewa et al. [26] proved that varroacidal effect when propolis was extracted and used
propolis has a strong narcotic and lethal effect on V. in 55% ethanol, where most of the water-soluble and some
Am-Euras. J. Sci. Res., 11 (5): 320-331, 2016
329
water insoluble components were extracted. This shows acaricides and if it is effective in the field experiment
that even if some of the components of propolis are and if it has no negative effects on bee themselves it
solubilised in the high humidity of the hive’s interior, their may has the possibility to shed light on the use of
concentration is too weak to kill the mites [26]. Most of propolis as in hive treatment with possible natural benefit
the varroacidal agents of propolis are water insoluble of propolis used by honeybees and minimize the cost of
leading to the dominance of propolis extracted in 55% beekeeping.
ethanol. Based on the above conclusions the following
CONCLUSIONS AND RECOMMENDATIONS
This preliminary work was focused on the narcotic as this will help to sustain the health of honey bee
and lethality efficacy of propolis of different origins and (Apis mellifera) already established in the bee
the pest’s diseases affecting bee health. To gain a better keeper’s apiary in the district.
view of what is affecting the managed honey bee health, International trade in these commodities has been
it is important to understand the key pests and diseases mainly responsible for distributing most of the known
affecting bee health. The world distribution of honeybee pests and diseases of bees. Authorities, should
disease is a great importance to bee keepers as there is a responsible for monitoring the spread of animal
significant relationship between the success of apiculture diseases and for regulating international trade in bees
industry and the control of honey bee health problems. and bee products.
This is because if it once occurs in the colony, they cause To fully exploit opportunities in beekeeping
partial or total loss of colonies and most of them spread subsector, interventions to address constraints and
very quickly and difficult to treat. Varroa, nosema, detecting the occurrence of honey bee diseases and
amoeba, pests like wax moth and spider could be the major health problem at apiaries level is the key step to
honey bee health problems responsible for endanger prevent their harmful effects as early as possible.
honey bee health within the hive in Toke-kutaye districts. Nosema and Amoeba and require careful monitoring
it is difficult to trace how and when varroa mite was and management to ensure losses are kept to a
introduced, Ethiopia has been tested positive to varroa minimum.
mite. The presence of varroa mite in the country is highly Authorities are advised to come up with an urgent
significant and market oriented might be due to high bee monitoring programme to determine mite infestation
colony mobility coupled with lack of awareness neither levels and its effects to honeybee colonies
from the beekeepers nor from the experts enhanced its It is therefore very important that the existing
high rate of distributions. This study for second time problems are well managed to maintain bee health
reported the presence of varroa mite in Ethiopia and the and that the risks and consequences of pests and
first finding in the study area. Furthermore, it advanced diseases are well understood and appropriate plans
understanding of the existences of varroa mite as in place to deal with any such honey bee health
implicated with honeybee colony losses and economic problem.
significance to the countries in general. Even though Propolis from different geographic origins of the
Nosema and amoeba their presence in honeybee colonies country should further investigated effect of different
could result in shortening of honey bee live and concentration of propolis extracted in different level
determinately reduces honeybee colonies strength of ethanol and type of effective propolis identified
predispose bees to pest and predators which directly and The varroacidal component of propolis should
indirectly affecting the beekeeping sector and honey extracted and field trial in the beehives as soon as
production in the woreda. Associated with the current possible
problematic varroa mite in beekeeping subsector and
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