The 82-year-old Tibetan spiritual leader, who lives in exile in India, was to speak at a three day conference in Botswana's capital Gaborone, which begins on August 17.
An official statement released late on Friday said the Dalai Lama had written to the president of Botswana and the conference organisers, the Mind and Life Institute, expressing "profound regret" at having to cancel his visit "due to exhaustion".
"His Holiness has reluctantly had to concede that his 82-year-old body was telling him to rest.
"Since his physicians have also advised him to avoid undertaking long journeys for the next few weeks, His Holiness will return to Dharamsala to recuperate," the statement said.
The Buddhist leader has lived in exile in India since he fled a failed Tibetan uprising in 1959.
Botswana said last month that the visit was "purely private" but added that the Dalai Lama would be granted the status of a foreign dignitary.
Beijing issued a warning to Botswana, asking the African nation to respect China's "core interests".
China's ruling Communist party views the Dalai Lama as a dangerous "separatist" campaigning for Tibetan independence. It consistently condemns other countries which let him visit.
The Dalai Lama says he wants more autonomy for Tibet rather than outright independence.
Chinese foreign ministry spokesperson Lu Kang last month called for Botswana to "earnestly respect" China's sovereignty, and "make the right decision" on Tibet-related issues.
"China will not interfere in other countries' internal affairs, and we will not accept that other countries do things that would harm the core interests of China," he said.
China is a key investor in Africa and the continent's largest trade partner. In Botswana it has helped build power plants, road networks, bridges and schools.