Honey Bee Apis mellifera Parasites in the Absence of Nosema ceranae Fungi and Varroa destructor Mites

Few areas of the world have western honey bee (Apis mellifera) colonies that are free of invasive parasites Nosema ceranae (fungi) and Varroa destructor (mites). Particularly detrimental is V. destructor; in addition to feeding on host haemolymph, these mites are important vectors of several viruses that are further implicated as contributors to honey bee mortality around the world. Thus, the biogeography and attendant consequences of viral communities in the absence of V. destructor are of significant interest. The island of Newfoundland, Province of Newfoundland and Labrador, Canada, is free of V. destructor; the absence of N. ceranae has not been confirmed. Of 55 Newfoundland colonies inspected visually for their strength and six signs of disease, only K-wing had prevalence above 5% (40/55 colonies = 72.7%). Similar to an earlier study, screenings again confirmed the absence of V. destructor, small hive beetles Aethina tumida (Murray), tracheal mites Acarapis woodi (Rennie), and Tropilaelaps spp. ectoparasitic mites. Of a subset of 23 colonies screened molecularly for viruses, none had Israeli acute paralysis virus, Kashmir bee virus, or sacbrood virus. Sixteen of 23 colonies (70.0%) were positive for black queen cell virus, and 21 (91.3%) had some evidence for deformed wing virus. No N. ceranae was detected in molecular screens of 55 colonies, although it is possible extremely low intensity infections exist; the more familiar N. apis was found in 53 colonies (96.4%). Under these conditions, K-wing was associated (positively) with colony strength; however, viruses and N. apis were not. Furthermore, black queen cell virus was positively and negatively associated with K-wing and deformed wing virus, respectively. Newfoundland honey bee colonies are thus free of several invasive parasites that plague operations in other parts of the world, and they provide a unique research arena to study independent pathology of the parasites that are present.

Infra-Population and -Community Dynamics of the Parasites Nosema apis and Nosema ceranae, and Consequences for Honey Bee (Apis mellifera) Hosts

Nosema spp. fungal gut parasites are among myriad possible explanations for contemporary increased mortality of western honey bees (Apis mellifera, hereafter honey bee) in many regions of the world. Invasive Nosema ceranae is particularly worrisome because some evidence suggests it has greater virulence than its congener N. apis. N. ceranae appears to have recently switched hosts from Asian honey bees (Apis cerana) and now has a nearly global distribution in honey bees, apparently displacing N. apis. We examined parasite reproduction and effects of N. apis, N. ceranae, and mixed Nosema infections on honey bee hosts in laboratory experiments. Both infection intensity and honey bee mortality were significantly greater for N. ceranae than for N. apis or mixed infections; mixed infection resulted in mortality similar to N. apis parasitism and reduced spore intensity, possibly due to inter-specific competition. This is the first long-term laboratory study to demonstrate lethal consequences of N. apis and N. ceranae and mixed Nosema parasitism in honey bees, and suggests that differences in reproduction and intra-host competition may explain apparent heterogeneous exclusion of the historic parasite by the invasive species

Effects of Nosema apis, N. ceranae, and coinfections on honey bee (Apis mellifera) learning and memory

Western honey bees (Apis mellifera) face an increasing number of challenges that in recent years have led to significant economic effects on apiculture, with attendant consequences for agriculture. Nosemosis is a fungal infection of honey bees caused by either Nosema apis or N. ceranae. The putative greater virulence of N. ceranae has spurred interest in understanding how it differs from N. apis. Little is known of effects of N. apis or N. ceranae on honey bee learning and memory. Following a Pavlovian model that relies on the proboscis extension reflex, we compared acquisition learning and long-term memory recall of uninfected (control) honey bees versus those inoculated with N. apis, N. ceranae, or both. We also tested whether spore intensity was associated with variation in learning and memory. Neither learning nor memory differed among treatments. There was no evidence of a relationship between spore intensity and learning, and only limited evidence of a negative effect on memory; this occurred only in the co-inoculation treatment. Our results suggest that if Nosema spp. are contributing to unusually high colony losses in recent years, the mechanism by which they may affect honey bees is probably not related to effects on learning or memory, at least as assessed by the proboscis extension reflex.

Effects, but no interactions, of ubiquitous pesticide and parasite stressors on honey bee (Apis mellifera) lifespan and behaviour in a colony environment

Interactions between pesticides and parasites are believed to be responsible for increased mortality of honey bee (Apis mellifera) colonies in the northern hemisphere. Previous efforts have employed experimental approaches using small groups under laboratory conditions to investigate influence of these stressors on honey bee physiology and behaviour, although both the colony level and field conditions play a key role for eusocial honey bees. Here, we challenged honey bee workers under in vivo colony conditions with sublethal doses of the neonicotinoid thiacloprid, the miticide tau-fluvalinate and the endoparasite Nosema ceranae, to investigate potential effects on longevity and behaviour using observation hives. In contrast to previous laboratory studies, our results do not suggest interactions among stressors, but rather lone effects of pesticides and the parasite on mortality and behaviour, respectively. These effects appear to be weak due to different outcomes at the two study sites, thereby suggesting that the role of thiacloprid, tau-fluvalinate and N. ceranae and interactions among them may have been overemphasized. In the future, investigations into the effects of honey bee stressors should prioritize the use of colonies maintained under a variety of environmental conditions in order to obtain more biologically relevant data.

The ectoparasitic mite Tropilaelaps mercedesae reduces western honey bee, Apis mellifera, longevity and emergence weight, and promotes Deformed wing virus infections

Historically an ectoparasite of the native Giant honey bee Apis dorsata, the mite Tropilaelaps mercedesae has switched hosts to the introduced western honey bee Apis mellifera throughout much of Asia. Few data regarding lethal and sub-lethal effects of T. mercedesae on A. mellifera exist, despite its similarity to the devastating mite Varroa destructor. Here we artificially infested worker brood of A. mellifera with T. mercedesae to investigate lethal (longevity) and sub-lethal (emergence weight, Deformed wing virus (DWV) levels and clinical symptoms of DWV) effects of the mite on its new host. The data show that T. mercedesae infestation significantly reduced host longevity and emergence weight, and promoted both DWV levels and associated clinical symptoms. Our results suggest that T. mercedesae is a potentially important parasite to the economically important A. mellifera honey bee.

The COLOSS BEEBOOK Volume I, Standard methods for Apis mellifera research: Introduction

The COLOSS BEEBOOK is a practical manual compiling standard methods in all fields of research on the western honey bee, Apis mellifera. The COLOSS network was founded in 2008 as a consequence of the heavy and frequent losses of managed honey bee colonies experienced in many regions of the world (Neumann and Carreck, 2010). As many of the world’s honey bee research teams began to address the problem, it soon became obvious that a lack of standardized research methods was seriously hindering scientists’ ability to harmonize and compare the data on colony losses obtained internationally. In its second year of activity, during a COLOSS meeting held in Bern, Switzerland, the idea of a manual of standardized honey bee research methods emerged. The manual, to be called the COLOSS BEEBOOK, was inspired by publications with similar purposes for fruit fly research (Lindsley and Grell, 1968; Ashburner 1989; Roberts, 1998; Greenspan, 2004).

The COLOSS BEEBOOK Volume II, Standard methods for Apis mellifera pest and pathogen research: Introduction

The COLOSS BEEBOOK is a practical manual compiling standard methods in all fields of research on the western honey bee, Apis mellifera. The COLOSS network was founded in 2008 as a consequence of the heavy and frequent losses of managed honey bee colonies experienced in many regions of the world (Neumann and Carreck, 2010). As many of the world’s honey bee research teams began to address the problem, it soon became obvious that a lack of standardized research methods was seriously hindering scientists’ ability to harmonize and compare the data on colony losses obtained internationally. In its second year of activity, during a COLOSS meeting held in Bern, Switzerland, the idea of a manual of standardized honey bee research methods emerged. The manual, to be called the COLOSS BEEBOOK, was inspired by publications with similar purposes for fruit fly research (Lindsley and Grell, 1968; Ashburner, 1989; Roberts, 1998; Greenspan, 2004).

The COLOSS BEEBOOK Volume II: Standard methods for Apis mellifera pest and pathogen research

n recent years, declines of honey bee populations have received massive media attention worldwide, yet attempts to understand the causes have been hampered by a lack of standardisation of laboratory techniques. Published as a response to this, the COLOSS BEEBOOK is a unique collaborative venture involving 234 bee scientists from 34 countries, who have produced the definitive guide to how to carry out research on honey bees. It is hoped that these volumes will become the standards to be adopted by bee scientists worldwide. Volume II includes approximately 600 separate protocols dealing with the study of the pests and diseases of the honey bee, Apis mellifera. These cover epidemiology and surveying techniques, virus diseases, bacterial diseases such as European and American foulbrood, fungal and microsporidian diseases such as Nosema, mites such as Acarapis, Varroa and Tropilaelaps, and other pests such as the small hive beetle.

The COLOSS BEEBOOK Volume I: Standard Methods for Apis mellifera Research

In recent years, declines of honey bee populations have received massive media attention worldwide, yet attempts to understand the causes have been hampered by a lack of standardisation of laboratory techniques. Published as a response to this, the COLOSS BEEBOOK is a unique collaborative venture involving 234 bee scientists from 34 countries, who have produced the definitive guide to how to carry out research on honey bees. It is hoped that these volumes will become the standards to be adopted by bee scientists worldwide. Volume I includes approximately 1,100 separate protocols dealing with the study of the honey bee, Apis mellifera. These cover anatomy, behavioural studies, chemical ecology, breeding, genetics, instrumental insemination and queen rearing, pollination, molecular studies, statistics, toxicology and numerous other techniques

Potential for virus transfer between the honey bees Apis mellifera and A. cerana

Viruses seem to play a key role in European honey bee, Apis mellifera health, and have a much broader host spectrum than previously thought. Few studies have investigated interspecific virus transfer within the genus Apis. The introduction of A. mellifera into Asia exposed endemic Apis species to the risk of obtaining new viruses or viral strains and vice versa. To investigate the potential for host shifts, virus prevalence and sequences were monitored over three years in single and mixed-species apiaries hosting introduced A. mellifera and endemic Apis cerana. Deformed wing virus (DWV), Israeli acute paralysis virus (IAPV), black queen cell virus (BQCV), and sacbrood virus (SBV) were found, but not KBV, VDV-1, ABPV, or CBPV. Virus infections and prevalence were generally lower in A. cerana compared to A. mellifera, and varied over the years. The sequence data provided evidence for interspecific transfer of IAPV, BQCV, and DWV, but SBV strains seem to be species specific. Prevalence and sequence results taken together indicate that interspecific transfers of viruses are rare, even if honey bees are kept in close proximity. We discuss the pattern observed in the context host specificity and resistance. Our understanding of the extent of these exchanges is limited by a lack of knowledge on the mechanisms of adaptation of viruses to different hosts.

Virus infections and winter losses of honey bee colonies (Apis mellifera)

Apiculturists have recently been confronted with drastic and inexplicable winter losses of colonies, and virus infections may be involved. Here, we surveyed 337 Swiss honey bee colonies in the winter of 2005 and 2006 and categorized their health status as: 1. dead (= no or few live bees left); 2. weak (= dwindling, high mortality of adult bees); or 3. healthy (= normal overwintering colony). From each colony, pooled adult workers were analyzed for deformed wing virus (DWV), acute bee paralysis virus (ABPV), chronic bee paralysis virus (CBPV) and Kashmir bee virus (KBV). Neither KBV nor CBPV were found, but significantly higher ABPV and DWV infections were found in dead vs. weak vs. healthy colonies (except DWV in 2006 between weak and healthy). Moreover, ABPV and DWV loads were positively correlated with each other. This is the first report demonstrating statistically significant correlations between viruses associated with Varroa destructor and winter mortality.

Genetic characterization of slow bee paralysis virus of the honeybee (Apis mellifera L.)

Complete genome sequences were determined for two distinct strains of slow bee paralysis virus (SBPV) of honeybees (Apis mellifera). The SBPV genome is approximately 9 5 kb long and contains a single ORF flanked by 5'- and 3'-UTRs and a naturally polyadenylated 3' tail, with a genome organization typical of members of the family Iflaviridae The two strains, labelled `Rothamsted' and 'Harpenden', are 83% identical at the nucleotide level (94% identical at the amino acid level), although this variation is distributed unevenly over the genome. The two strains were found to co-exist at different proportions in two independently propagated SBPV preparations The natural prevalence of SBPV for 847 colonies in 162 apiaries across five European countries was <2%, with positive samples found only in England and Switzerland, in colonies with variable degrees of Varroa infestation

Standard methods for maintaining adult Apis mellifera in cages under in vitro laboratory conditions

Adult honey bees are maintained in vitro in laboratory cages for a variety of purposes. For example, researchers may wish to perform experiments on honey bees caged individually or in groups to study aspects of parasitology, toxicology, or physiology under highly controlled conditions, or they may cage whole frames to obtain newly emerged workers of known age cohorts. Regardless of purpose, researchers must manage a number of variables, ranging from selection of study subjects (e.g. honey bee subspecies) to experimental environment (e.g. temperature and relative humidity). Although decisions made by researchers may not necessarily jeopardize the scientific rigour of an experiment, they may profoundly affect results, and may make comparisons with similar, but independent, studies difficult. Focusing primarily on workers, we provide recommendations for maintaining adults under in vitro laboratory conditions, whilst acknowledging gaps in our understanding that require further attention. We specifically describe how to properly obtain honey bees, and how to choose appropriate cages, incubator conditions, and food to obtain biologically relevant and comparable experimental results. Additionally, we provide broad recommendations for experimental design and statistical analyses of data that arises from experiments using caged honey bees. The ultimate goal of this, and of all COLOSS BEEBOOK papers, is not to stifle science with restrictions, but rather to provide researchers with the appropriate tools to generate comparable data that will build upon our current understanding of honey bees.