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Combining thermopreference (T<i>_p</i>) and CO₂-gated heat-seeking assays, we studied the thermal <i>preferendum</i> and response to thermal cues in three <i>Culex</i> mosquito species exhibiting differences in native habitat and host preference (e.g., biting cold and/or warm-blooded animals). Results show that these species differ in both T<i>_p</i> and heat-seeking behavior. In particular, we found that <i>Culex territans</i>, which feed primarily on cold-blood hosts, did not respond to heat during heat-seeking assays, regardless of the CO₂ concentration, but exhibited an intermediate T<i>_p</i> during resting. In contrast, <i>Cx. quinquefasciatus</i>, which feeds on warm blooded hosts, sought the coolest locations on a thermal gradient and responded only moderately to thermal stimuli when paired with CO₂ at higher concentrations. The third species, <i>Cx. tarsalis</i>, which has been shown to feed on a wide range of hosts, responded to heat when paired with high CO₂ levels and exhibited a high T<i>_p</i>. This study provides the first insights into the role of heat and CO₂ in the host seeking behavior of three disease vectors in the <i>Culex</i> genus and highlights differences in preferred resting temperatures.