BOSTON—In his first public appearance since the Boston Marathon bombing, suspect Dzhokhar Tsarnaev pleaded not guilty in a packed federal courtroom Wednesday to a host of charges stemming from the attack that left three dead and hundreds wounded.

Mr. Tsarnaev wore an orange jumpsuit unbuttoned over a black shirt and the same shaggy hairstyle seen in many photographs that emerged after his arrest. He sat low in his chair and repeatedly touched his face.

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