Tracheal Mite Research Articles

Health status of honeybee colonies (Apis mellifera) and disease-associated risk factors in different agroecological zones of Southwest Ethiopia

A cross-sectional study design was conducted in different agroecological zones of southwest Ethiopia from October 2019 to October 2021. The study aimed to determine the prevalence and associated risk factors for honeybee diseases and pests, as well as the impact of these issues on honeybee colonies and their products. To identify potential risk factors for honeybee disease and pests, a multivariate random effects logistic regression analysis was used. Adult honeybee and brood samples from a total of 384 honeybee colonies were collected and tested using standard laboratory diagnostic methods. The highest prevalence (55.8%) of ants was recorded, followed by wax moths (22.5%) and hive beetles (23.3%). In the current study, the main honeybee diseases observed in the study areas were varroosis (36.5%), bee lice (5.2%), nosemosis (39.6%), amoeba (56%), and chalkbrood (4.5%). However, tracheal mites, sachbrood, and American and European foul brood, were not detected. The agroecological zone (OR = 5.2, 95% CI: 1.75–14.85), type of hive (OR = 2.9, 95% CI: 1.17–17.03), management system (OR = 4.3, 95% CI: 1.23–14.70), and the management of the colony (OR = 3.5, 95% CI: 1.31–9.14) were identified as risk factors for varroosis in these areas. The occurrence of nosemosis in colonies was also influenced by the agroecological zone (OR = 12.2, 95% CI: 3.06–48.54) and colony management (OR = 3.4, 95% CI: 1.59–7.23). The agroecological zone (OR = 10.5, 95% CI: 12.76–22.63) and hive type (OR = 3.0, 95% CI: 1.39–6.36) were the primary risk factors for the occurrence of amoeba in honeybee colonies. However, the occurrence of bee lice (OR = 34.7, 95% CI: 3.96–104.93) and chalkbrood (OR = 4.8, 95% CI: 1.44–13.16) in honeybee colonies was only influenced by the agroecological zone in the study areas. This study demonstrated that losses in honey production in the area are significantly attributed to honeybee disease and pests. Therefore, it is essential to increase public awareness of how honeybee diseases and pests affect honey production and to develop and implement appropriate control measures for these diseases and pests. Furthermore, more studies should be conducted to characterize and isolate other causes of honeybee diseases and pests in various locations.

Comparison of indoor (refrigerated) versus outdoor winter storage of commercial honey bee, Apis mellifera (Hymenoptera: Apidae), colonies in the Western United States.

Honey bees (Apis mellifera L.) are critical to the pollination of many important crops in the United States, and one crop that demands large numbers of colonies early each year is almonds. To provide adequate numbers of colonies for almond pollination, many beekeepers move colonies of bees to high-density holding yards in California in late fall, where the bees can fly and forage, but little natural pollen and nectar is available. In recent years, high colony losses have occurred in some operations following this management strategy, and alternative approaches, including indoor storage of colonies, have become more commonly used. The current study evaluated colonies kept indoors (refrigerated and/or controlled atmosphere) for the winter compared with those kept outdoors in either Washington or California. Colonies were evaluated for strength (frames of bees), brood area, lipid composition of worker bees, colony weight and survival, parasitic mites (Varroa mites, tracheal mites), and pathogens (Nosema spp.). No differences were found in colony weight, survival, parasitic mite levels, or pathogen prevalence among the treatments. Colonies stored indoors and outdoors in WA had significantly more frames of bees and less brood present after the storage period than colonies stored outdoors in CA. Lipid composition of honey bees stored indoors was significantly higher than colonies stored outdoors in WA or CA. The implications of these findings for overall colony health and improved pollination activity are discussed.

PCR-based detectionof the honeybee tracheal mite (Acarapis woodi) in Türkiye.

Acarapis woodi (Rennie 1921) (Acari: Tarsonemidae) is one of the mites that settles in the respiratory system of honeybees (Apis mellifera L. (Hymenoptera, Apidae)) and distributed throughout the world. It causes significant economic losses on honey production. In Türkiye, studies on the existence of A. woodi are very limited and so far, no studies on the molecular diagnosis and phylogenetic of it have been reported in Türkiye. This study was conducted to investigate the prevalence of A. woodi in Türkiye, especially in areas where beekeeping is intense. Diagnosis of A. woodi was performed using both microscopic and molecular methods using specific PCR primers. Adult honeybee samples were collected from 1.193 hives in 40 provinces of Türkiye between 2018 and 2019. Based on identification studies, the presence of A. woodi was detected in a total of 3 hives (0.5%) in 2018 and 4 hives (0.7%) in 2019. This is the first report for determination of A. woodi in Türkiye.

Effects of neonicotinoids on honey bee autogrooming behavior against the tracheal mite Acarapis woodi.

The European honey bee, Apis mellifera, is the most common and important pollinator of crops worldwide. Honey bees are damaged by destructive parasitic mites, but they also have evolved a behavioral immune system to remove them. Exposures to neonicotinoids, however, can cause significant behavioral effects because these compounds alter the central role of nicotinic acetylcholine receptor in insect brains. In this study, we assessed the effects of three neonicotinoids that have a high toxicity to bees-imidacloprid, thiamethoxam, and clothianidin-on the behavioral immune system of honey bees. We used A. mellifera and the endoparasitic mite Acarapis woodi as a behavioral immune system model because A. mellifera can effectively remove the mite by autogrooming. Our results did not demonstrate an effect of neonicotinoid application on whether bees show autogrooming or on mite removal, but the time to initial autogrooming became shorter and the number of autogrooming attempts increased. As opposed to previous studies, our findings indicate that the honey bee response to parasitic mites becomes more sensitive after exposure to neonicotinoids.Clinical Trials Registration: Not applicable.

Beekeeping Genetic Resources and Retrieval of Honey Bee Apis mellifera L. Stock in the Russian Federation: A Review.

Simple SummaryThe loss of honey bees poses a significant problem for the beekeeping industry. Opportunities to recover the stock of honey bees are crucial in areas with drastic losses. In this article, we describe known native and bred Apis mellifera L. stocks of honey bees in the Russian Federation and their characteristics, identified using morphometric and genetic methods. We review the experience of Russian breeders and other breeders in breeding initial A. mellifera Far East honey bees with inherited traits for Varroa resistance. We also describe the bred types of honey bee A. m. mellifera L., A. m. caucasia Gorb., A. m. carpatica Avet. that offer potential in the aim to recover honey bee populations after losses. Our findings indicate that it is necessary to avoid honey bee losses by breeding resistant honey bees. Several bred types of honey bees from the dataset showed high performance in terms of overwintering, resistance to Varroa destructor, Acarapis woodi, and Nosema infections, and little or no swarm conditions. This review shows the potential to increase selection efforts in the breeding of resistant Apis mellifera L. honey bee populations in the Russian Federation and throughout the world.The loss of honey bees has drawn a large amount of attention in various countries. Therefore, the development of efficient methods for recovering honey bee populations has been a priority for beekeepers. Here we present an extended literature review and report on personal communications relating to the characterization of the local and bred stock of honey bees in the Russian Federation. New types have been bred from local colonies (A. mellifera L., A. m. carpatica Avet., A. m. caucasia Gorb.). The main selection traits consist of a strong ability for overwintering, disease resistance and different aptitudes for nectar collection in low and high blooming seasons. These honey bees were certified by several methods: behavioral, morphometric and genetic analysis. We illustrate the practical experience of scientists, beekeepers and breeders in breeding A. mellifera Far East honey bees with Varroa and tracheal mite resistance, which were the initial reasons for breeding the A. mellifera Far Eastern breed by Russian breeders, Russian honey bee in America, the hybrid honey bee in Canada by American breeders, and in China by Chinese beekeepers. The recent achievements of Russian beekeepers may lead to the recovery of beekeeping areas suffering from crossbreeding and losses of honey bee colonies.

Amplicon Sequencing of Variable 16S rRNA from Bacteria and ITS2 Regions from Fungi and Plants, Reveals Honeybee Susceptibility to Diseases Results from Their Forage Availability under Anthropogenic Landscapes.

European Apis mellifera and Asian Apis cerana honeybees are essential crop pollinators. Microbiome studies can provide complex information on health and fitness of these insects in relation to environmental changes, and plant availability. Amplicon sequencing of variable regions of the 16S rRNA from bacteria and the internally transcribed spacer (ITS) regions from fungi and plants allow identification of the metabiome. These methods provide a tool for monitoring otherwise uncultured microbes isolated from the gut of the honeybees. They also help monitor the composition of the gut fungi and, intriguingly, pollen collected by the insect. Here, we present data from amplicon sequencing of the 16S rRNA from bacteria and ITS2 regions from fungi and plants derived from honeybees collected at various time points from anthropogenic landscapes such as urban areas in Poland, UK, Spain, Greece, and Thailand. We have analysed microbial content of honeybee intestine as well as fungi and pollens. Furthermore, isolated DNA was used as the template for screening pathogens: Nosema apis, N. ceranae, N. bombi, tracheal mite (Acarapis woodi), any organism in the parasitic order Trypanosomatida, including Crithidia spp. (i.e., Crithidia mellificae), neogregarines including Mattesia and Apicystis spp. (i.e., Apicistis bombi). We conclude that differences between samples were mainly influenced by the bacteria, plant pollen and fungi, respectively. Moreover, honeybees feeding on a sugar based diet were more prone to fungal pathogens (Nosema ceranae) and neogregarines. In most samples Nosema sp. and neogregarines parasitized the host bee at the same time. A higher load of fungi, and bacteria groups such as Firmicutes (Lactobacillus); γ-proteobacteria, Neisseriaceae, and other unidentified bacteria was observed for Nosema ceranae and neogregarine infected honeybees. Healthy honeybees had a higher load of plant pollen, and bacteria groups such as: Orbales, Gilliamella, Snodgrassella, and Enterobacteriaceae. Finally, the period when honeybees switch to the winter generation (longer-lived forager honeybees) is the most sensitive to diet perturbations, and hence pathogen attack, for the whole beekeeping season. It is possible that evolutionary adaptation of bees fails to benefit them in the modern anthropomorphised environment.

Range expansion of the tracheal mite Acarapis woodi (Acari: Tarsonemidae) among Japanese honey bee, Apis cerana japonica, in Japan

Acarapis woodi, a parasitic mite of honey bees, was first detected in Japan in 2010. Infestation was mostly observed in the Japanese honey bee (Apis cerana japonica) and was rare in the European honey bee (Apis mellifera). By 2014, the mites had spread throughout central and eastern Japan. In the current study, we investigated the subsequent expansion of the mite to western Japan. Our research revealed that the mites were distributed across most of Japan by 2018, except for Wakayama and Kochi prefectures. Many small remote islands more than 20km away from mainland Japan are still free of A. woodi, but bees on some of these islands were infested. About 40% of colonies of the Japanese honey bee in Japan were infested by the mites, and average mite prevalence of the infested colonies was about 50% during the 6-year study. There was no trend of decline in the infested colony proportion or in the mite prevalence. In addition, the observation of Japanese honey bee colonies by hobby beekeepers for two signs of mite infestation, K-wing and crawling bees, was an effective means for estimating infestation by tracheal mites.

Differential autogrooming response to the tracheal mite Acarapis woodi by the honey bees Apis cerana and Apis mellifera

The infestation of honey bees by the endoparasitic tracheal mite Acarapis woodi was first discovered in Apis mellifera on the Isle of Wight, England, and the mite has since spread to all continents except Australia. Since 2010, this tracheal mite has spread rapidly in the Japanese honey bee, Apis cerana japonica, of mainland Japan, causing considerable colony mortality. In contrast, infestations by the mites in the imported and managed European honey bee, A. mellifera, have rarely been observed in Japan. A previous laboratory experiment revealed a difference in susceptibility by demonstrating that the tracheal mite more frequently enters tracheae of A. cerana than those of A. mellifera. In this study, we compared autogrooming responses of A. cerana and A. mellifera by depositing a mite on each honey bee’s mesoscutum, and we then assessed the efficacy of autogrooming to remove the mite. The bees that received mites more frequently showed an autogrooming response compared to unchallenged bees in both bee species. However, a significantly greater proportion of A. mellifera individuals autogroomed compared to A. cerana. In addition, when bees autogroomed, A. mellifera removed the tracheal mite more effectively than A. cerana. When considering all bees in the mite-deposited group, the proportion of mite removal in A. mellifera was almost twice as high as that in A. cerana. Thus, the difference in susceptibility to the tracheal mite between these two bee species is attributed to the difference in the behavioral response threshold to mites and the effectiveness of mite removal by grooming.

Comparison of qPCR and Morphological Methods For Detection of Acarapis Woodi in Honey Bee Samples

Abstract The honey bee tracheal mite Acarapis woodi is an internal obligate parasite of adult honey bees (Apis mellifera). The small size of the mites and location within the bee’s trachea create a challenge for diagnostic identification. Detailed assessment of low-level mite infestation involves microscopic examination of the bee’s tracheae, but this traditional diagnostic method takes a long time. In recent years, new molecular diagnostic techniques have been developed to identify A. woodi with the use of PCRs. The purpose of this study was to compare the diagnostic sensitivity of microscopic examination and qPCR method for the detection of A. woodi in honey bee samples. Thirty-six samples were analyzed with both methods and qPCR was more sensitive than the microscopic method to detect A. woodi. This work demonstrates the practical application of molecular technology as a support tool for surveys and contingency management and to provide robust surveillance data on the presence or absence of A. woodi in honey bee colonies.

기문응애(Acarapis woodi) 특이 유전자 검출을 위한 초고속 nested PCR법 개발

기문응애(Acarapis woodi) 특이 유전자 검출을 위한 초고속 nested PCR법 개발

Prevalence of honeybee tracheal mite disease and infestation levels of Acarapis woodi in honeybee colonies of Morelos, Mexico

Una de las principales enfermedades que afectan a las abejas melíferas es la acariosis traqueal causada por el ácaro Acarapis woodi. El objetivo de este estudio fue estimar la prevalencia de la acariosis traqueal y los niveles de infestación de Acarapis woodi en colonias de abejas del estado de Morelos, en donde la apicultura es un actividad pecuaria importante que se realiza bajo diversas condiciones climáticas y bajo diferentes sistemas de producción. El estudio se llevó a cabo utilizando muestras de abejas de 1,198 colonias que estaban ubicadas en 233 apiarios distribuidos a lo largo del territorio estatal, que son propiedad de 96 unidades de producción apícola. Los apiarios representan el 21.5 % del total de apiarios que existen en el estado y las unidades de producción representan el 13.7 % del total de unidades de producción registradas en Morelos. Los resultados indican que la prevalencia estimada para la acariosis traqueal en Morelos es de 0.02 y que el 10.30 % de los apiarios incluidos en el estudio tuvieron al menos una colonia positiva a esta enfermedad. El porcentaje de infestación promedio en las colonias positivas fue de 7.32 ± 0.75, con un nivel mínimo de infestación de 5 % y un nivel máximo de 20 %. Estos resultados indican que actualmente la acariosis traqueal tiene una prevalencia relativamente baja en Morelos y el porcentaje de infestación promedio estimado en las colonias positivas también es bajo, asimismo indican que la prevalencia estimada para esta enfermedad en este estudio es menor a la reportada hace 34 años.

Investigation on the relationship between population synthesis of the honeybee stocks reared in Crete and their resistance to the tracheal mite Acarapis woodi, Rennie

The population synthesis of honey bee breeds reared in Crete by crossbreeding among the native race Apis mellifera adami, Ruttner, the Macedonian bee Apis mellifera macedonica, Ruttner, and the Italian bee Apis mellifera ligustica, Spinal, was determinated. The probable relationship between population synthesis and resistance to tracheal mite Acarapis woodi, Rennie, of these breeds was also investigated. Samples of 50 bees were taken from 45 colonies of an apiary, infested with acarine disease, in which the bees showed a great diversity in their morphological characteristics and were examined as regards: a) the morphological and morphometric characteristics of the sampled bees, and b) the number of infested colonies, the prevalence, the unilateral or bilateral infection and the parasite intensity of thoracic tracheas, from every sample infested by Acarapis woodi. After examination the colonies were classified in four groups, according to morphological and morphometric characteristics that hade the majority of the bees in each sample: Group I, hybrids of Apis mellifera macedonica, group II, hybrids of Apis mellifera ligustica, group III, hybrids of "bright yellow" type of Apis mellifera ligustica, and group IV, diversiform hybrids. From the tracheas examinations it was detected that: the population synthesis affects the resistance of bee colonies. Bee colonies of group I showed the highest resistance. On the contrary, those of group III showed the lowest. Bee colonies of the two other groups showed relatively high resistance but less high than that of A. m. macedonica.

Molecular methods indicate lack of spread of Acarapis woodi introduced to honey bees in western Norway.

The tracheal mite, Acarapis woodi, may be one of many factors contributing to the decline in honey bee (Apis mellifera) populations. Databases on the widespread distribution of A. woodi exist, but the data seem patchy. Norway is not listed as being infested, although there have been at least two separate introductions of the parasite. Investigations in 2003, 2006, and 2009 using standard microscopy methods indicated persistence of A. woodi in honey bees in this region. In 2013, we conducted another survey. Samples were sent in from 335 beehives belonging to 39 apiaries, and all were asked to complete a questionnaire. Analysis for A. woodi in the submitted samples was by PCR, with sequencing of positive results. The results described in this article indicate that this parasite still persists in some apiaries in this region, but at a low, and possibly decreasing, level, with positive results obtained from just two (5.1%) of the apiaries. Of the 17 beekeepers that answered the questionnaire, none reported symptoms of infestation with A. woodi. Sequencing of PCR products indicated a difference between the two A. woodi isolates. Our results were generally encouraging regarding the apparent lack of spread of A. woodi, within the County. Furthermore, the sequencing results may indicate two separate introductions rather than spread. Nevertheless, the scarcity of data, the vulnerability of honey bee populations globally, and the potential contribution of this parasite to reduced survival, indicate that the situation should be continued to be monitored. In addition, Norwegian beekeepers should be made aware of, and follow, restrictions regarding import and transport of bees, both nationally and internationally.

Is Acarapis woodi mite currently infesting Apis mellifera colonies in Argentina?

The honey bee tracheal mite Acarapis woodi (Rennie) (Acari: Tarsonemidae) is an internal obligate parasite of the respiratory system of adult honey bees (Apis mellifera), which may be one of the factors causing the loss of bee colonies and the decrease in the beekeeping industry. In Argentina, the last prevalence study of A. woodi was in 1998, and therefore there is no current data on the prevalence and impact of the mite in colonies of A. mellifera in this country. For this reason, the objective of this study was to determine the prevalence of A. woodi in honey bee colonies of apiaries belonging to different geographic regions of Argentina (Northwest, Chaco, Mesopotamic, Pampean and Patagonic) using two different methods of diagnosis. One method was traditional microscopy, which is based on direct visualization of A. woodi in the prothoracic tracheas of bees, and the other technique was molecular analysis using real-time PCR with specific primers to amplify CO1 gene of A. woodi. To perform the analysis of prevalence, bees were collected during winter, because during this season, populations of A. woodi are more abundant in bee colonies. The results were negative for both methods of diagnosis, with prevalence equal to 0%. Considering that in recent years it has been reported an increased mortality in bee colonies in Argentina, we note that acarapisosis is not currently impacting on the disappearance of A. mellifera colonies in Argentina.

Differential susceptibility to the tracheal mite Acarapis woodi between Apis cerana and Apis mellifera

In Japanese honey bees Apis cerana japonica, infestations of the tracheal mite Acarapis woodi have spread rapidly over the mainland of Japan, causing damage and the collapse of colonies. Meanwhile, infestations by mites in Apis mellifera have hardly been observed in Japan. In this study, we assessed and compared the susceptibility of the two species, A. cerana and A. mellifera, using an inoculation assay. We found that migrating female mites entered the tracheae of more newly emerged bees in both species but more frequently in A. cerana than in A. mellifera. Hence, the higher susceptibility in A. cerana is proposed as a factor causing the explosive epidemic of tracheal mites in only A. cerana in Japan. Moreover, we compared grooming behaviors between the two bee species using an observation assay as a preliminary experiment, although the bees were not exposed to the presence of tracheal mites. From these observations, the frequency of autogrooming (self-grooming) on the thorax in A. cerana was lower than that in A. mellifera. The difference in susceptibility to the mite between these two species may be due to the difference in grooming behavior frequency.

Molecular detection of the tracheal mite Locustacarus buchneri in native and non-native bumble bees in Chile

Bumble bees (Bombus spp.) are considered one of the most important pollinators of native, non-native, and domesticated plants. Pathogens and parasites, such as the tracheal mite Locustacarus buchneri (Podapolipidae), are harmful to bumble bees. We developed primers based on the mitochondrial CO1 gene for the specific detection of L. buchneri directly from samples of native Bombus dahlbomii and non-native B. terrestris and B. ruderatus. PCR products amplified and sequenced for these primers shared 100 % identity among themselves and around 99–100 % identity with other sequences of L. buchneri reported in Belgium, the Netherlands, and Japan. Bombus terrestris was the most infected species (41 %), followed by B. ruderatus (31 %) and B. dahlbomii (23 %). This is the first report involving L. buchneri in native and non-native bumble bees in Chile. Our described PCR-based detection offers a feasible, specific, and rapid method for the screening of L. buchneri in any bumble bee species.

Field application of menthol for Japanese honey bees, Apis cerana japonica (Hymenoptera: Apidae), to control tracheal mites, Acarapis woodi (Acari: Tarsonemidae)

The first record of tracheal mites, Acarapis woodi, in Japan was made in 2010. These mites have since caused serious damage to the colonies of Japanese honey bees, Apis cerana japonica. In the present study, to control the mites on Japanese honey bees with l-menthol, an agent used for European honey bees, Apis mellifera, we investigated (1) the seasonality of menthol efficacy, (2) the overwintering mortality of menthol-treated colonies, and (3) the menthol residue in honey under field conditions in cooperation with private beekeepers of Japanese honey bees. Seasonal menthol efficacy was tested by applying 30g of l-menthol for 1month in different seasons. Mite prevalence was measured by dissecting the honey bee thorax. Overwintering mortality was monitored during winter after checking the mite prevalence in autumn, and was compared with that of untreated colonies reported in our previous study. The residual level of menthol in honey was measured by GC-MS. The results showed that the menthol-treated colonies had a smaller rate of increase in mite prevalence than the untreated colonies. The effects of menthol were highest in March and April. The winter mortality was depressed by menthol treatment. Honey samples extracted from the menthol-treated colonies included 0.4ppm of menthol residue on average. Our findings suggest that menthol treatment is effective for controlling the tracheal mites on Japanese honey bees.

Molecular genetic analysis of tracheal mite resistance in honey bees

Honey bee resistance to the parasitic tracheal mite Acarapis woodi is known to be mediated by autogrooming. During autogrooming, bees use their midlegs to remove migrating foundress mites, thereby reducing invasion into their tracheae. We investigated the relationship between markers identified by single nucleotide polymorphism association and variation in average colony infestation rates, and in individual bees from a backcrossed mapping population combining tracheal mite-resistant and susceptible lines. We identified a genetic interval that may confer resistance and postulate on potential candidate genes coded in the interval. Three genes, ionotropic glutamate receptor (iGluR), tyrosine hydroxylase (TyHyd), and Gemin3, are involved in primarily neural pathways and may impact autogrooming. However, inability to validate marker segregation with infestation phenotype in an unrelated population with clear but small differences in tracheal mite resistance shows that SNPs or other genetic markers cannot necessarily be presumed to be functional for marker-assisted selection.

Tracheal mites, Acarapis woodi, greatly increase overwinter mortality in colonies of the Japanese honeybee, Apis cerana japonica

AbstractThis study evaluated the damage caused by the tracheal mite Acarapis woodi to colonies of the Japanese honeybee Apis cerana japonica, by investigating mite prevalence and colony death of the honeybees during winter. Honeybees of 114 colonies were sampled from October to December to detect mite prevalence. The colony death was monitored up to the end of April. The observed mortality due to mite infestation was compared with data from two previous studies about European honeybees. Under normal conditions in the absence of mites, 21 % of intact colonies died during the winter season. However, once the honeybee colonies were infested with A. woodi, the overwinter mortality was drastically increased. The mortality of Japanese honeybee colonies infested with the tracheal mites was higher than that reported historically in European honeybee colonies infested with the mites. Investigation of colony mortality at the apiary level showed that mortality was significantly higher in mite-infested than non-infested apiaries, but there were no significant differences related to the estimated winter air temperature. These results suggest that tracheal mite infestation seriously increases the overwinter mortality of Japanese honeybees.

Development and evaluation of a real-time PCR assay for the detection of Acarapis woodi (tracheal mites) in Apis mellifera

The tracheal mite, Acarapis woodi, is an obligate endoparasite of honeybees and a regulated pest in countries where these mites are absent. This work describes the development of a real-time PCR method for detecting tracheal mites in honeybees. The real-time PCR was evaluated for specificity, sensitivity and speed to detect A. woodi, compared to the standard manual thoracic disc method (TDM). The assay detected A. woodi down to a 1 % incidence level in bees and 1000 copies of the target DNA when using plasmid standards. Initial testing showed no cross-reaction with the other two Acarapis species from different geographical regions or with other species of mites associated with bees. However, during extensive testing of bees, a rare population of Acarapis externus mite was identified that did cross-react with the assay. Despite this cross-reaction, the assay has been shown to be a useful screening tool and results are reliable if the TDM is used as a backup to screen hives where a positive signal is obtained.