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The objectives of this retrospective study were to investigate the relationship between temperature-humidity index (THI) and conception rate (CR) of lactating dairy cows, to estimate a threshold for this relationship, and to identify periods of exposure to heat stress relative to breeding in an area of moderate climate. In addition, we compared three different heat load indices related to CR: mean THI, maximum THI, and number of hours above the mean THI threshold. The THI threshold for the influence of heat stress on CR was 73. It was statistically chosen based on the observed relationship between the mean THI at the day of breeding and the resulting CR. Negative effects of heat stress, however, were already apparent at lower levels of THI, and 1 hour of mean THI of 73 or more decreased the CR significantly. The CR of lactating dairy cows was negatively affected by heat stress both before and after the day of breeding. The greatest negative impact of heat stress on CR was observed 21 to 1 day before breeding. When the mean THI was 73 or more in this period, CR decreased from 31% to 12%. Compared with the average maximum THI and the total number of hours above a threshold of more than or 9 hours, the mean THI was the most sensitive heat load index relating to CR. These results indicate that the CR of dairy cows raised in the moderate climates is highly affected by heat stress.