A videotape of British hostage Norman Kember and two other peace activists has been broadcast on Arab satellite television station al-Jazeera.
The 74-year-old, from north-west London, was seized in Baghdad with two Canadians and an American in November.
The members of peace group Christian Peacemaker Teams (CPT) were last seen in a video clip broadcast in January.
The UK Foreign Office said it was concerned the American Tom Fox did not appear on the tape dated 28 February.
"We hope that all four are well and will be released unharmed," a spokeswoman said.
The men were kidnapped on 26 November by a previously unknown group calling itself the Swords of Truth Brigade, which has demanded all Iraqi prisoners be freed.
Canadian-based CPT, which has been calling for an end to the presence of US and British troops in Iraq, said it would continue "to pray for their safe and speedy release" of its four members.
"We do not know what to make of Tom Fox's absence from this video. However we do know what motivated Tom and his colleagues to go to Iraq," it said in a statement.
It said the four had been carrying out peaceful work on behalf of all Iraqi detainees, adding: "We believe that the root cause of the abduction of our colleagues is the US and British-led invasion and occupation of Iraq."
The video, which was broadcast with no sound, showed Mr Kember alongside Canadians James Loney, 41, a community worker from Toronto, and Harmeet Singh Sooden, 32, an electrical engineer from Montreal.
TIMELINE OF ABDUCTION
26 November: Norman Kember abducted along with three other peace workers
30 November: Video footage of the hostages aired on al-Jazeera
2 December: Second video of the hostages aired. Their captors threaten to kill them unless their demands are met by 8 December
8 December: Third video is aired on al-Jazeera showing Mr Kember and US hostage Tom Fox wearing orange jumpsuits, blindfolded and shackled. The captors' deadline is extended to 10 December
28 January: New video of hostages, dated 21 January, aired on al-Jazeera
5 March: Vigils are held across the UK to mark 100 days since the kidnappings
7 March: New video of three of the hostages, dated 28 February, aired on al-Jazeera
But Mr Fox, 54, from Clearbrook, Virginia, who has been working with CPT and Iraqi human rights organisations for the past two years, did not appear.
Paul Slattery, a member of Mr Fox's US-based support group, told the BBC: "We are all crossing our fingers and hoping for the best.
"Everything in Baghdad is so messed up at the moment, it is easy to believe that the hostages are split up and for some reason they just couldn't bring them all together for the video."
According to al-Jazeera, the three hostages appealed to Gulf leaders and their own governments to help bring about their release.
Vigils to mark 100 days since the kidnappings were held in a number of cities across the world last weekend.
Vigils for the captives took place over the weekend
The kidnappers initially threatened to kill the hostages if their demands were not met by 10 December.
In an earlier video broadcast on 29 January - the first news of the men for seven weeks - the kidnappers again threatened to kill the hostages if their demands were not met.
Mr Kember's friend, Chris Cole, told BBC News of his relief at seeing the latest video but said he remained "very worried".
"I think he looks in reasonable health in the circumstances," Mr Cole said.
"We would very much like to have them here to continue their work for peace and justice in the Middle East."
Meanwhile, Anas al-Tikriti from the Muslim Association of Britain, who went to Baghdad to try to secure Mr Kember's release in December, said it might be possible to open negotiations with the abductors.
"Rather than basically just ignore the entire world and carry out their threat regardless, the abductors are showing some sort of sense that they do want to communicate, they do want to negotiate," he said.