VIRAL DISEASE SUSPECTIBILITY AND TRANSMISSION
Transmission among colonies is a central feature for the epidemiology of honey bee pathogens..
Honey bees are social insects living in densely packed colonies of related individuals. Their homogeneity, in terms of physical environment and genetic composition, and the close contact among nestmates makes them particularly susceptible to disease infections because transmission among individuals within the colony is markedly facilitated in such a setting (Forfert et al. 2016). Not only must honey bee pathogens ensure transmission within the colony (intra-colonial transmission), they must also be transmitted between different colonies (inter-colonial transmission) to maintain themselves in a population (Forfert et al. 2016; Fries and Camazine 2001). This represents a critical step in the life of many honey bee pathogens and pests (Fries and Camazine 2001). Transmission can occur via various routes: 1) the flight of an (infected) drone or worker from its own into another colony (drifting); 2) the return of a worker bee after robbing an infected colony; 3) contact of infected and uninfected workers from different colonies during foraging; 4) contact with infected material from the environment; or 5) sexual contact of infected drones and queen (Fries and Camazine 2001; Yue et al. 2006; Yañez et al. 2012).
Transmission among colonies is a central feature for the epidemiology of honey bee pathogens. High colony abundance may promote transmission among colonies independently of apiary layout, making colony abundance a potentially important parameter determining pathogen prevalence in populations of honey bees. To test this idea, Forfert et al. (2016) sampled male honey bees (drones) from seven distinct drone congregation areas (DCA), and used their genotypes to estimate colony abundance at each site. A multiplex ligation dependent probe amplification assay (MLPA) was used to assess the prevalence of ten viruses, using five common viral targets, in individual drones. There was a significant positive association between colony abundance and number of viral infections. This result highlights the potential importance of high colony abundance for pathogen prevalence, possibly because high population density facilitates pathogen transmission. Pathogen prevalence in drones collected from DCAs may be a useful means of estimating the disease status of a population of honey bees during the mating season, especially for localities with a large number of wild or feral colonies.
Two main routes of chronic bee paralysis virus (CBPV) transmission are known: contact between adult bees when healthy bees are crowded together with infected individuals (Bailey et al. 1983), and spread of infectious particles in the feces of paralyzed bees that are taken up orally by healthy nestmates (Ribiere et al. 2007). Crowded conditions favoring these two routes occur frequently in growing bee colonies during the Spring and early Summer. In experimental studies, bees can be successfully infected by feeding the virus mixed into sugar solution, by topical application of viral particles, or by direct injection (Bailey et al. 1963; Bailey 1965a; Rinderer and Rothenbuhler 1976). Experimentally infected adult bees show symptoms after about five to six days, similar to those of naturally infected bees (Rinderer and Rothenbuhler 1976; Bailey 1965b; Chevin et al. 2012). A higher number of virus particles is needed to infect bees by topical application and by oral infection, as compared to injection (Bailey et al. 1963; Bailey et al. 1983; Bailey 1965b). A recent study showed that CBPV replication was more effective in individual bees inoculated per os compared to bees inoculated via the cuticle (Toplak et al. 2013).
Chronic bee paralysis virus (CBPV) is known as a disease of worker honey bees.
Chronic bee paralysis virus (CBPV) is known as a disease of worker honey bees. To investigate pathogenesis of the CBPV on the queen, the sole reproductive individual in a colony, Amiri et al. (2014) conducted experiments regarding the susceptibility of queens to CBPV. Results from susceptibility experiment showed a similar disease progress in the queens compared to worker bees after infection. Infected queens exhibit symptoms by day six post infection and virus levels reach 1011 copies per head. In a transmission experiment they showed that social interactions may affect the disease progression. Queens with forced contact to symptomatic worker bees acquired an overt infection with up to 1011 virus copies per head in six days. In contrast, queens in contact with symptomatic worker bees, but with a chance to receive food from healthy bees outside the cage appeared healthy. The virus loads did not exceed 107 in the majority of these queens after nine days. Symptomatic worker bees may transmit sufficient active CBPV particles to the queen through trophallaxis, to cause an overt infection.
Bee viruses (Kashmir bee virus (KBV) and sacbrood virus (SBV)) can potentially be transmitted horizontally via worker secretions. Shen et al. (2005) demonstrated that bee viruses (KBV and SBV) were detected in worker bees, brood food, honey, pollen and royal jelly. These bee viruses may be transmitted from worker bees to larvae or to other adult bees (queen, other workers or drones) via food resources in the colony. It is also possible that bee viruses might be transmitted among colonies by feeding bees the honey or pollen gathered from diseased colonies. This is a practice used by some beekeepers to help colonies survive during times of low flowering. Also, dead or weakened colonies can be raided by worker bees from other colonies and honey and pollen taken back to their colonies. The importance of these food sources in viral transmission between colonies needs to be defined. Support for food being a route of transmission comes from studies done by Bailey (1969), who injected SBV preparations into adult bees and fed SBV to larvae, and then detected viruses in different tissues (head, abdomen, midgut and hypopharyngeal glands) by immunodiffusion. This experiment demonstrated virus in the hypopharyngeal glands of bees that had been injected with or fed SBV. The results also showed that more SBV accumulated in the heads of infected bees than in other regions.
Honey bees can be exposed to several viruses and transmission can occur both horizontally and vertically (Chen et al. 2006; Chen and Siede 2007). In horizontal transmission, viruses are transmitted among individuals of the same generation. Vertical transmission occurs from queens to their offspring and can have two main causes: (I) infected sperm originating from the drones and (II) contaminated eggs originating from infected spermatheca and/or ovaries of the queen. Ravoet et al. (2015) investigated vertical transmission of viruses within a Belgium queen bee breeding program. They found a high prevalence of different honey bee viruses in eggs used in queen breeding operations, with more than 75% of the egg samples being infected with at least one virus. The most abundant viruses were deformed wing virus (DWV) and sacbrood virus (SBV) (> 40%), although Lake Sinai virus (LSV) and acute bee paralysis virus (ABPV) were also occasionally detected (between 10-30%). In addition, aphid lethal paralysis strain Brookings, black queen cell virus, chronic bee paralysis virus and Varroa destructor Macula-like virus occurred at very low prevalences (< 5%).
Under field conditions, Varroa jacobsoni were shown to be highly effective vectors of deformed wing virus (DWV) between bees.
Amiri et al. (2016) investigated whether sexual transmission during multiple matings of queens is a possible way of obtaining virus infection in queens. In an environment with high prevalence of deformed wing virus, queens were trapped upon their return from natural mating flights. The last drone’s endophallus, if present, was removed from the mated queens for deformed wing virus quantification, leading to the detection of high-level infection in three endophalli. After oviposition, viral quantification revealed that seven of the 30 queens had high-level deformed wing virus infections, in all tissues, including the semen stored in the spermathecae. Two groups of either unmated queens with induced egg laying, or queens mated in isolation with drones showing comparatively low deformed wing virus infections served as controls. None of the control queens exhibited high-level viral infections. Their results demonstrate that deformed wing virus infected drones are competitive to mate and able to transmit the virus along with semen, which occasionally leads to queen infections. Virus transmission to queens during mating may be common and can contribute noticeably to queen failure.
Under field conditions, Varroa jacobsoni were shown to be highly effective vectors of deformed wing virus (DWV) between bees. Adult female mites obtained from honey bee pupae naturally infected with DWV contained virus titers many times in excess of those found in their hosts and, beyond that, which might be expected from a concentration effect. It is therefore possible that DWV may be capable of replicating within the mite. Bees which tested positive for DWV exhibited characteristic morphological deformity and/or they died during pupation. Asymptomatic bees had much lower virus titers than those which were deformed or had died during pupation. It is therefore suggested that for DWV to cause pathology it must be present in pupae above a certain concentration. The amount of DWV vectored by V. jacobsoni will depend on the mites’ level of infection, which will in turn depend on whether they had fed previously on dead or deformed bees and also on the rate of replication of the virus within the mites. Consequently, developing bees infested with large numbers of mites could suffer a high incidence of deformity if the mites are heavily infected or harbor an especially virulent strain of virus. A positive relationship was found between increasing numbers of mites on individual bees and the incidence of morphological deformity and death. This probably reflected the large number of viral particles transmitted by the mites, which resulted in many multiply infested bees dying before emergence. These results demonstrate the importance of the role of viruses when considering the pathology of V. jacobsoni and that much of the pathology previously associated with the effects of mite feeding could be attributed directly to secondary pathogens vectored by V. jacobsoni (Bowen-Walker et al. 1999).
Honey bee queens are the main reproductive female in the colony and therefore the development of a new colony and the replenishment of old workers depend on an egg-laying queen. Queens seldom show symptomatic disease, although she is the longest-living member of colony. Francis et al. (2013) evaluated 86 queens for viruses that were collected from beekeepers in Denmark. All queens were tested separately by two real-time PCRs: one for the presence of deformed wing virus (DWV), and one that would detect sequences of acute bee-paralysis virus, Kashmir bee virus and Israeli acute paralysis virus (AKI complex). Worker bees accompanying the queen were also analyzed. The queens could be divided into three groups based on the level of infection in their head, thorax, ovary, intestines and spermatheca. Four queens exhibited egg-laying deficiency, but visually all queens appeared healthy. Viral infection was generally at a low level in terms of AKI copy numbers, 134/430 tissues (31%) showing the presence of viral infection ranging from 101 to 105 copies. For DWV, 361/340 tissues (84%) showed presence of viral infection (DWV copies ranging from 102 to 1012), with 50 tissues showing viral titers > 107 copies. For both AKI and DWV, the thorax was the most frequently infected tissue and the ovaries were the least frequently infected. Relative to total mass, the spermatheca showed significantly higher DWV titers than the other tissues. The ovaries had the lowest titer of DWV. No significant differences were found among tissues for AKI. A subsample of 14 queens yielded positive results for the presence of negative-sense RNA strand, thus demonstrating active virus replication in all tissues.
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Clarence Collison is an Emeritus Professor of Entomology and Department Head Emeritus of Entomology and Plant Pathology at Mississippi State University, Mississippi State, MS.