At least 3,958 members of the US military have died since the beginning of the Iraq war in March 2003, according to an Associated Press news agency count.
For their part, US troops killed eight suspected fighters and captured 26 people - including an alleged Shia militia leader - in two days of raids across Iraq, the US military said.
In Diyala province, a curfew has been imposed after the Sunni Awakening Councils ended patrols of towns and neighbourhoods.
The councils, comprised of mostly Sunni tribesmen who have worked with American troops to force al-Qaeda and other fighters from their home towns, are refusing to return to the streets unless the chief of police resigns.
Tensions began mounting after two girls were kidnapped and killed last week by men dressed in Iraqi security forces uniform. Their bodies were later found stripped naked.
The armed groups gave the chief of police until midday on Friday to apologise and arrest the men, who they say are Shia militiamen in the Iraqi security forces.
"We hereby declare suspension of all co-operation with both US military, Iraqi security forces and the local government," Abu Abdullah, spokesman for Diyala's Awakening Council, announced after the deadline passed.
The fallout between the councils and Iraq's government is likely to impede US efforts to gain full control of the region.
Hoda Abdel-Hamid, Al Jazeera's correspondent in Iraq, said: "As US forces push into Diyala as part of a massive operation to clear up the province, they need these people.
"And once you have one Awakening Council who is going to retract, it might be more difficult to convince others citizens anywhere else to be part of these Awakening Councils."
There are now more than 130 Awakening Councils across Iraq and financially supported by the US military.
The crisis threatens to spread as Awakening Councils in Falluja, Ramadi and certain neighbourhoods in Baghdad, complain of a lack of support from the government.
Targeted by both al-Qaeda fighters and Shia militias, a roadside bomb killed three Awakening Council members and wounded eight others south of Baghdad on Thursday. About 100 members were killed in January alone.
"The Iraqi government hasn't been too forthcoming and we'll see more and more of these problems bubbling up unless the US manages to convince the Iraqi government to act a bit more quickly and to integrate them into the Iraqi security forces," Abdel-Hamid said.
"Most of these Awakening Councils say 'We are securing our area, we want to be Iraqi security forces and we want to be officially recognised by the Iraqi government.'"
The government has said it will would integrate only a portion of them into the security forces, and try to allocate civilian jobs for the rest.
In other violence in Iraq, a university student was shot dead in the centre of Mosul, in the north of the country.
In Hawija town, three policemen were injured when a bomb targeting their patrol exploded.
In the southern Iraqi city of Wasit, the US army arrested a leader of a one of groups that split from the Mahdi Army militia and three other suspects during a raid.