According to the official results announced, conservative candidate Jill Mortimer obtained 15,529 votes, almost double those obtained by her labor opponent Paul Williams (8,589).
Hartlepool, in the north of England, has been in the hands of the Labor Party since 1964, and since the current constituency was created in 1974.
Former MP Mike Hill resigned in March after allegations of sexual harassment.
This is only the second time since 1982 that an opposition party has lost a partial legislative election to a party in government.
The result is one of the first to be announced in a series of elections held on Thursday.
In total, about 4,650 representatives from 143 counties in England, 13 directly elected mayors, 39 police commissioners and 25 deputies from the London City Assembly were voted on.
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Elections were also held to choose the 129 members of the Scottish Assembly and 60 members of the Wales Assembly, the outcome of which will dictate the respective autonomous governments.
“It is a historic result”, congratulated Amanda Milling, from the top of the party, in a note.
Among some members of the labor party, it is impossible to hide dissatisfaction with the first results of regional and local elections. “It’s terribly disappointing,” tweeted Labor MP Richard Burgon. “We went backwards in the areas where we should win. Labor leadership must change,” he said.