The search team looking for the remains of Ben Needham have been digging around a tree which was not there when the tot went missing.
Detectives analysed a picture taken shortly after Ben vanished in July 1991 and realised that a tree which is close to the spot where the toddler was last seen was not there when he vanished.
There is some speculation the 25ft tree may have been planted afterwards.
Detective Inspector Jon Cousins, of South Yorkshire Police, said: “The only record we have of the time that Ben disappeared in 1991 is a photo.
“It was taken around 10 to 14 days after his disappearance, when members of the media arrived here on Kos to have a look at the investigation.
“It is the earliest record we have and the closest record of the time Ben disappeared. There is an indication that a tree that is here now clearly wasn’t there at the time.
“That is why we are concentrating our effort around it to make sure we have covered it.
“We have been able to analyse that area with the experts who are here now and, of course, we are able to look at all the fauna and tree growth around it.”
Detectives are working on the theory that Ben may have been accidentally run over and buried by a digger clearing a building plot close to where the 21-month-old was last seen.
He was playing outside a farmhouse his grandfather was renovating when he disappeared.
A 19-strong search team is excavating one of two sites where the digger dumped material on the day Ben vanished.
They will move onto a second site if nothing is found at the first.
Some items of fabric are being examined to establish whether they match the clothing Ben was wearing when he went missing.
Experts are also examining part of a sandal which was found when some initial excavation work was carried out in 2012.