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The close symbiotic relationships between vertebrates and their naturally-occurring bacterial microbiomes are an increasingly appreciated field of research for macro- and microbiologists alike. In many marine fish of the meso- and aphotic zones, bioluminescence is mediated by such symbioses, but the fragility, rarity, and physical inaccessibility of such taxa has prevented close analysis in the lab environment. The speciose fish genus Siphamia (Kurtiformes, Apogonidae), is an outlier among bioluminescent fish taxa in that members can be found in shallow reefs of both temperate and tropical ecoregions, making it far more accessible to research. In this study, we examine nuance in the association of Siphamia and its sole bioluminescent symbiote, the gram-negative proteobacterium Photobacterium mandapamensis. Colonies of P. mandapamensis were isolated from the ventral light organs of three different Siphamia species occupying three different climates. These bacterial isolates were screened using a PCR fingerprinting approach and unique genotypes were sequenced using Oxford Nanopore Technology. Our findings will likely shed new light on the specificity and evolution of this host-bacterium symbiosis, with temperate and tropical fish species potentially hosting different substrains of P. mandapamensis. Moreover, new substrains of P. mandapamensis - and potentially even a new species of the same genus - may be described from the light organs of previously-unexamined Siphamia species from temperate habitats. The close pairing between Siphamia and P. mandapamensis represents the first one-to-one vertebrate symbiosis to be studied extensively in the lab environment, and may soon become a model system for other vertebrate symbioses, especially in the deep ocean.