North Lanarkshire Council, one of Scotland's biggest local authorities, has confirmed traces of horse DNA were detected in a frozen burger removed for testing last week from a school kitchen.
It immediately withdraw all similar frozen burgers from its 122 primary schools, 24 secondaries and 11 schools for children with special needs.
It is the first local authority north of the Border to confirm a beef product in a school kitchen contained horsemeat, raising concerns about other foods being tainted.
North Lanarkshire and Renfrewshire councils had already withdrawn some beef products supplied by food distribution firm Brakes Group to carry out tests. It came after the group confirmed horse DNA was detected in lasagne the firm produces for Whitbread, which owns Premier Inn, Beefeater Grill and Brewers Fayre.
Rural Affairs Secretary Richard Lochhead said it was unacceptable the tainted burger had been supplied to the schools. He added: "However, North Lanarkshire Council has taken immediate action to withdraw the product from the schools and as a precautionary measure all Scottish schools have been advised to put a hold on the use of frozen burgers.
"Of the thousands of tests, this is the first positive result in our schools but it is one too many. No company should be supplying our schools with beef products that contain traces of horsemeat."
On Thursday, the Scottish Government dismissed concerns that horsemeat might find its way into school meals, after a campaign group led by the chef Nick Nairn called for greater transparency.
North Lanarkshire said it had notified the Food Standards Agency (FSA) of the discovery and said stocks of beefburger products had been removed from school kitchens as a precaution.
The investigation is focusing on the use of frozen burger supplies during the past three months, the maximum length of time it said they would be held in storage.
A council spokesman said: "We are working closely with the FSA and Scotland Excel [the local government procurement centre] and will continue to take any action necessary to ensure the integrity of foods used in our establishments."
On Thursday, the Scottish Government responded to Nairn's demands for reassurances over the horsemeat scandal saying the authorities must come clean about what has gone into school food.
It had said: "There is no evidence that any horsemeat has found its way into any school meals in Scotland and the excellent statutory nutritional standards for food do not lend themselves to using processed products.
"Beef purchased for schools is good quality, fresh or frozen, 100% traceable and requires independent inspections accredited to high industry standards.
"The relatively modest quantities of processed meat such as sausages or burgers used have a good specification and the necessary traceability."
Brakes had insisted its beef ranges supplied to schools had tested negative for horsemeat.
North Lanarkshire said it had disposed of 20 portions of Brakes lasagne last week after they were recalled by the catering firm and that separate tests on mince were all clear.
In Renfrewshire, tests on beef lasagne found no trace of horsemeat.
Scottish Labour's rural affairs and environment spokeswoman Claire Baker said: "We need greater transparency in our supply chain and a Government will ing to be proactive not just reactive.
"The Scottish Government were caught sleeping on the job over the horsemeat scandal and either still don't have the full facts or are simply refusing to divulge them."
A Brakes spokesman said: "We have recently been informed of a positive test by the Food Standards Agency, but at the moment it is not known who supplied the burger. We are aware that we supply Scotland Excel, so, as a precaution, until we are able to ascertain the facts, we will be placing the burger on hold.
"We will be working with the Food Standards Agency to identify what has happened as a matter of urgency.
"We apologise for any inconvenience this will cause."
Meanwhile, Birds Eye is withdrawing three beef ready meals from sale in the UK and Ireland.
The move follows tests that found 2% of horse DNA in a chilli con carne dish which is sold by Birds Eye in Belgium.
Its spaghetti bolognese, shepherd's pie and lasagne are made by the same Belgian manufacturer, Frigilunch N.V., and are being withdrawn "as a precautionary measure".
In a statement, Birds Eye said: "We want to reassure you from the testing we have completed that all Birds Eye beef burgers, beef pies and beef platters do not contain horse DNA.
"Regrettably, we have found one product, chilli con carne, produced for us by Frigilunch N.V. and sold in Belgium, that has tested positive for horse DNA at 2%.
"Whilst this is not a food safety issue, it is clearly unacceptable. In accordance with our high standards, we are immediately withdrawing this product from sale.
"As a precautionary measure in the UK and Ireland we will withdraw all other products produced by the same supplier, namely traditional spaghetti bolognese 340g, shepherd's pie 400g and lasagne 400g."
Birds Eye's parent company Iglo Foods Group, which also owns Findus in Italy, said it had been carrying out checks on all of its beef products after other manufacturers reported their foods had been contaminated with horsemeat.
The chilli con carne is the only product that they have found to contain equine DNA.
The Birds Eye products taken off UK supermarket shelves will not be replaced until further tests have been carried out, the company said.
Customers who purchased any of the products affected will be given a refund if they contact Birds Eye consumer services.
Birds Eye said in the statement: "The quality of our food is of the utmost importance to us. We know that our consumers rely on us to be certain that they are eating only what is labelled on the packaging and that they can always rely on us to act responsibly.
"Iglo Foods Group has introduced an ongoing DNA testing programme and we have enhanced our normal quality assurance procedures. This will help us ensure that we continue to reach the standards that all our consumers expect from our products.
"We want to apologise to consumers and reassure them that we will keep them fully informed and that we are taking action to deal with this issue."
The Birds Eye announcement comes as the Food Standards Agency prepares to publish the results of its latest round of industry tests on meat products later today.
The FSA has told the food industry to share the results of product tests, which look for horse DNA down to a level of 1%, with the aim of increasing consumer confidence.
Results of the industry tests on 2,501 beef products collated by the FSA last week revealed 29 positive results, relating to Aldi's special frozen beef lasagne and special frozen spaghetti bolognese, Co-op frozen quarter-pounder burgers, Findus beef lasagne, Rangeland's catering burger products, and Tesco value frozen burgers and value spaghetti bolognese.
Pub and hotel group Whitbread has also pulled lasagne and burgers from its menus after admitting horse DNA had been found in its food.
Horse meat has been discovered in school dinners, with cottage pies testing positive for horse DNA sent to 47 Lancashire schools before being withdrawn.
The FSA said this week that its own tests for horse meat in processed meat products were being expanded to ensure a wide range of products was sampled.
The first phase of tests concentrated on samples of minced beef products being checked for horse and pork DNA, while the second phase involved samples of beef-based ready meals.
The third phase will include products marketed or labelled as containing beef as a major ingredient. Products including gelatine, beef dripping, stock cubes, steak, stewing steak as well as ready meals which contain beef that is not minced are included. Testing on the third phase is due to begin next week.