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Borrelia burgdorferi B31MI carries 18 plasmid-carried genes that form the bdr gene family. The bdr genes of B. burgdorferi encode proteins that form three distinct subfamilies, the BdrD, BdrE, and BdrF subfamilies. bdr orthologs have been demonstrated to be carried by all Borrelia species analyzed, and their widespread distribution suggests that they play an important genus-wide functional role. The biological rationale for maintaining 18 bdr alleles has not been defined. It is our hypothesis that specific paralogs function in different environments and are differentially expressed in response to environmental conditions. As a first step in testing this hypothesis, the production patterns of the Bdr proteins in spirochetes grown under a variety of conditions were assessed through immunoblot analyses. The influence of temperature, serum deprivation, tick feeding, and the mammalian environment on Bdr production was evaluated. These analyses revealed that the synthesis of some Bdr paralogs is environmentally regulated. The production of BdrF2, BdrF1, BdrE4, and BdrE5 were upregulated in host-adapted bacteria, while the production levels of other Bdr paralogs were influenced by temperature and serum starvation. These observations suggest that different Bdr paralogs function in different biological environments and provide insight into the biological basis for maintaining multiple members of this gene family.