| Summary: | The strong association between <i>Varroa destructor</i>, deformed wing virus (DWV), and high overwintering colony losses (OCL) of honey bees is well established. Three DWV master variants (DWV-A, -B, and -C) have been described, and their role in colony mortality remains an open question. Therefore, the aim of this study is to investigate the seasonal prevalence, viral load, and changing distribution of the three DWV master variants within honey bee colonies from England, Wales, and 32 states across the United States. Here, we report that in 2016, DWV-B was prevalent (100%, <i>n</i> = 249) and dominant (95%) in England and Wales, compared to the US. (56%, <i>n</i> = 217 and 23%, respectively), where DWV-A was prevalent (83%, <i>n</i> = 217) and dominant (63%). DWV-C was regularly detected in low viral loads (<1 × 10<sup>7</sup> genome equivalents per bee) and at lower prevalence (58% in England and Wales, <i>n</i> = 203, and 14% across the United States, <i>n</i> = 124) compared to DWV-A and -B. DWV-B prevalence and dominance in England and Wales coincided with low OCL (6%). Meanwhile, a 60% loss was reported by participating U.S. beekeepers. In the United States, DWV-A prevalence (89%, <i>n</i> = 18) and viral load were significantly (<i>p</i> = 0.002) higher (1 × 10 <sup>8</sup>−1 × 10<sup>11</sup>) in colonies that died when compared to the surviving colonies (49% (<i>n</i> = 27), 1 × 10<sup>6</sup>−1 × 10<sup>10</sup>). DWV-B had low prevalence (56%, <i>n</i> = 18) in the colonies that died with viral loads of <1 × 10<sup>10</sup>. However, DWV-B was routinely detected in high viral loads (>1 × 10<sup>10</sup>) in surviving colonies from all sample locations, providing further supporting evidence of DWV-A exhibiting increased virulence over DWV-B at the colony level.
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