The letter, which is stamped by the Chinese embassy in Prague, also criticises the late Senate speaker, Jaroslav Kubera.
A popular right-wing figure, Kubera had planned to lead the Taiwan business trip but died of a heart attack on January 20.
"Czech companies whose representatives visit Taiwan with Speaker Kubera will not be welcomed in China or by the Chinese people," reads the letter, published online by the Czech Aktualne.cz news website.
"Czech companies with economic interests in China will have to pay for the visit," says the letter dated January 10, adding that China hoped the Czechs would adhere to its One-China policy and cancel the trip planned for later this year.
The document singles out several Czech companies with interests in China, including piano maker Petrof and carmaker Skoda Auto, the Czech division of Germany's Volkswagen Group.
It also mentions Home Credit Group, a unit of the financial PPF Group owned by the wealthiest Czech, Petr Kellner, a major player on China's lucrative loan market.
Czech ties with China suffered a blow last October when Prague city hall, run by a mayor from the anti-establishment Pirate Party, pulled out of a sister deal with Beijing over its insistence on the One-China policy.
Prague then signed a partnership agreement with Taipei in January, triggering outrage in Beijing.
Taiwan has been ruled separately from China since the end of a civil war in 1949, but under its "One-China" policy, Beijing considers it a part of its territory, with reunification by force an option.
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