Avoyelles Parish District Attorney Charles Riddle filed a motion Monday for recusal, citing his relationship with one of the officer's families.
Louisiana City Marshals Norris Greenhouse Jr., 23, and Derrick Stafford, 32, have been arrested and charged with second-degree murder and attempted second-degree murder.
According to the motion, Greenhouse's father, Norris Greenhouse Sr., is an assistant district attorney under Riddle.
Louisiana Attorney General James "Buddy" Caldwell said in a statement that he'd received the recusal order and "we will begin a detailed and thorough investigation, leaving no stone unturned."
"I can simply assure you that at the end of the process, justice will be served," he added.
In a court hearing Monday, a Louisiana judge set a $1 million bond for each of the two officers. The court has ordered both men to surrender their firearms and badges.
Nearly a week has passed since the incident occurred, but answers to disturbing questions are slow in coming.
A source close to the case information tells CNN that on the evening of November 3, marshals began pursuing the father, Chris Few, after they witnessed an argument between Few and his girlfriend in front of a local bar.
The source says the marshals alleged they witnessed "domestic abuse" and moved in to detain Few, who instead took off in his SUV.
The police pursuit ended in a hail of gunfire that left Few critically wounded and his young son dead.
Still unknown: Why did they open fire on 6-year-old Jeremy Mardis?
The first-grader was buckled in the front passenger seat of his father's truck when he was shot five times in the head and chest. At least 18 rounds were fired at the father and son from two firearms.
The boy's father was also struck. A hospital representative told CNN on Monday that authorities had upgraded Few from serious to fair condition.
The shooting happened on a dead-end street at the end of the chase in Marksville, a town of about 5,500 about 90 miles northwest of Baton Rouge, authorities said.
Investigators said there were no outstanding warrants against the father, and no firearm was found in his vehicle.
Greenhouse knew Few before the deadly encounter, a source familiar with the investigation told CNN. Investigators are looking into the extent of their relationship.
In an interview with CNN "New Day's" Alisyn Camerota, Louisiana State Police Superintendent Col. Michael Edmonson stressed that Marksville is a small town.
"We believe that they had some type of relationship where they met each other, knew each other," Edmonson said. "As this progresses, we'll certainly find out more and more information. But I think in a town like this, everyone knows each other."
At least one of the two officers was accused of a crime before.
According to The Town Talk newspaper in Alexandria, Louisiana, Stafford was indicted in 2011 on two counts of aggravated rape. The Town Talk reported that both cases were dismissed without prejudice in 2012. One of the incidents dated back to 2004. The other occurred while Stafford was employed by the Marksville Police Department, where he has worked for eight years.
Neither the Marksville Police Department nor Stafford's family has responded to CNN's requests for comment regarding those cases.
Stafford has been named as a defendant in five civil lawsuits, including at least one related to the use of excessive force. That particular case is still pending.
Greenhouse has also been named in a civil lawsuit related to the use of excessive force. That case is still pending.
Both men were working second jobs as city marshals when the shooting happened. Stafford is a full-time Marksville police lieutenant, and Greenhouse is a marshal in Alexandria, Edmonson said.
According to a CNN review of both officers' records, neither has ever been convicted of a crime.
The head of Louisiana's state police said "disturbing" body camera footage helped
build the case against the two officers accused of shooting the boy.
"I've been a police officer for 35 years, but as a father -- much less as a state police -- it was a disturbing, disturbing video that I watched, and that really helped move us forward," Edmonson said Sunday.
"I've got to deal in facts. What's important to me is what caused those police officers to pursue," Edmonson said. "What caused them to open fire?"
"He (Jeremy) didn't deserve to die like that. We need to find out why."
The town's mayor, John Lemoine, told CNN's Brianna Keilar on Monday that the body cameras were a recent addition for local police.
"We just went ahead and got these body cameras for these officers, and I'm glad that we did," Lemoine said. "It ... shed a lot of light on what happened the other night."
On Monday afternoon, a small prayer vigil was held at the site of the shooting.
About a dozen people joined hands in a circle around a small collection of mementos left for Jeremy. People have been leaving multicolored balloons, flowers, stuffed animals and notes around the dead-end street.
Broken glass is still strewn across the ground. Just steps away, at least one bullet hole has been marked with orange paint in a split-rail fence.
Some of the people gathered for the vigil were from Marksville, and others came from the surrounding area. Several told CNN they were unable to travel the distance to Hattiesburg, Mississippi, for Jeremy's funeral and burial, held Monday afternoon.
Jeremy Whittington was among those who attended the vigil. He told CNN's Martin Savidge he works with Few, and has been visiting him at the hospital.
He said he last visited Few on Sunday evening.
Whittington said Few was awake but drifting in and out of consciousness. He had difficulty speaking because he was connected to breathing tubes.
Jeremy's first-grade special needs teacher Roxanne Couvillion said the boy loved class, playing and alphabet puzzles. Jeremy was autistic and did not communicate verbally, she said. Couvillion said he was one of her favorite students.
She said she especially loved the way his eyes lit up when he smiled.
"We're just heartbroken," she said. "He always was an angel, and we know that he's watching over us."