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The two groups threw flares and other objects at each other in Novi Sad, prompting riot police to intervene, local media reported.
The collapse of a railway station roof in Novi Sad in November last year triggered a wave of protests across Serbia, condemning institutional corruption and heaping pressure on right-wing President Aleksandar Vucic.
Simultaneous protests were held across Serbia on Wednesday, mostly outside local headquarters of Vucic's Serbian Progressive Party.
In the capital, Belgrade, there was a large police presence outside the parliament, where supporters of both sides traded insults and threw objects at each other.
Riot police were deployed elsewhere as tensions ran high, prompting Vucic to call on his supporters for calm.
On Tuesday evening, large groups of masked men, some wielding batons, confronted anti-graft demonstrators outside Vucic's party offices in the town of Vrbas, about 100 kilometres (60 miles) northwest of Belgrade.
Footage shared on social media and by local broadcasters showed rocks and bottles being thrown.
Other videos appeared to show pro-government supporters with their faces covered throwing fireworks directly at anti-government protesters.
Serbia's political opposition has accused police of doing little to stop the violence, but officers denied the claim and said 16 officers were injured trying to keep the peace.
The Novi Sad railway station roof collapse, which killed 16 people, has been widely blamed on corruption.
For nearly nine months, large rallies -- some drawing hundreds of thousands to the streets -- have demanded a transparent investigation into the disaster and are now calling for snap elections.
But Vucic remains defiant and has repeatedly rejected calls to call early elections, saying the protests are part of a foreign plot to overthrow his government.
The protests, which have remained largely peaceful, have led to the resignation of the country's prime minister and the collapse of his ministry.
But Vucic remains in power at the helm of a reshuffled government.