Firms will be asked to use judgement to ensure their premises are not crowded and avoid unnecessary interaction between customers and staff under the new rules, which come into effect from one minute past midnight.
The regulations for Scotland’s businesses are a bid to combat the spread of the virus and have come amid criticism that firms were given less than 24 hours to implement changes.
The Scottish Chambers of Commerce said the short notice would put businesses “under immense strain” as they raced to put in mitigations which many have not used since the early days of the pandemic.
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The new rules stated that everyone who was able to work from home during the first lockdown should continue to do so, while businesses should reintroduce measures such as protective screens to separate customers and staff.
All staff who have to work in person should take a lateral flow test at least twice a week and more often if they are socialising outside of work, the guidance added.
The document published on Thursday afternoon by the Scottish Government also said businesses have a responsibility to increase the flow of air into their premises and consider maximum capacities – although formal physical distancing requirements are not in place.
Shops should control the flows of customers entering and exiting and in-store to minimise risk of transmission.
Speaking at First Minister’s Questions, Scottish Conservative leader Douglas Ross said the Government had issued “confusing and inaccurate advice to businesses”.
He warned clarity was needed for firms amid confusion from different government ministers as to when the guidance would be published.
Deputy First Minister John Swinney issued a correction after telling Holyrood’s Covid-19 Committee on Thursday morning the guidance would not be published until Friday.
Mr Ross said: ““The SNP Government have left business guidance to the last minute again and created unnecessary confusion with their mixed messaging.
“Businesses have been told one thing by the First Minister on Tuesday and another thing today. They now need to comply with this guidance from Friday, 24 hours earlier than they were expecting.
“We all understand the situation is moving quickly. However, if the public are expected to adapt quickly, then it’s right we demand the Government adapts quickly too and gives people the information they need on time.
“This guidance has now, finally, been published. But there’s still no word on when struggling small businesses will get the funding they desperately need to survive this difficult period.”
Ms Sturgeon said she had signed off the guidance minutes before the beginning of FMQs.
She said: “Omicron is currently raging around this country. It is doubling every two to three days.
"Every 24 hours matters in terms of saving people's lives and protecting the health service, so that is why we're going really fast and we are communicating with business organisations as we go.”
The guidance also says businesses should be encouraged to consider approaches which would reduce physical contact, such as introducing one-way systems and electronic ticketing.
Dr Liz Cameron, chief executive of the Scottish Chambers of Commerce, said: “Scotland’s businesses are continuing to do everything they can to keep their employees and customers safe from the ever-present threat of Covid-19 and the emergence of the new Omicron variant.
“We are now in the busiest time of year for hospitality and retail businesses, many of which are already having to deal with increased staff shortages and a deluge of customer cancellations.
"Whilst we welcome the publication of the updated guidance today, to expect these hard-pressed business to review, digest and enact it before it comes into effect tomorrow, will place immense strain on business owners and our employees.”
She added: “Talk of additional or extended restrictions is damaging consumer, business and employee confidence. What businesses need to hear is a clear UK-wide plan of when restrictions will end and a clear package of financial support to compensate for loss of trade.”
The document said: “In line with Fair Work principles, businesses and service providers should discuss working arrangements with their staff and any representatives, and take every reasonably practical step to facilitate working from home and make working from home the default position.
"Where it is reasonably practicable for a person to perform their work from home, they should do so. This is especially expected of those roles that were done at home at the beginning of the Covid-19 pandemic in March 2020.
"From December 17, 2021, by law, businesses, places of worship and service providers must take reasonable measures to minimise the risk of the incidence and spread of coronavirus. Supporting employees to work from home where reasonably practical is an important part of that.”
Detailed guidance was given to businesses which require employees to share transport, suggesting that no more than two people should be together in a car, while windows should be kept open and face coverings worn.
The document said while some measures, such as wearing face coverings, are enforceable by law, authorities would continue to take a flexible approach.
The guidance said: “Regulators and police will continue their current enforcement role for these requirements under the regulations following the 4 E’s approach – engage, explain, encourage and, as a last resort, enforce.”
Ewan MacDonald-Russell, Scottish Retail Consortium head of policy, said: “Despite the unnecessarily short notice, retailers will do everything they can to try to follow the updated guidance, despite having to bring in these measures ahead of the busiest week of the year.
"The main change for shoppers will be the requirement to maintain physical distancing in stores, which may limit the number of customers who can visit at any one time.
"Retail workers will do everything they can to manage this, but we would ask customers to show some consideration and patience as well.”
Scottish Liberal Democrat leader Alex Cole-Hamilton said: "This new guidance is long on advice and recommendations that will make life very hard for hospitality businesses, but short on measures to help firms whose business is collapsing in front of their eyes.
"Taking measures to limit gatherings and encourage caution are all very well, but they need to be matched pound for pound with financial support to stop firms going to the wall this winter."
He added: "As usual this guidance arrives less than 24 hours before it is due to come into force. That's bad for firms who have to schedule staff and order produce. It's also terrible for proper parliamentary scrutiny. The government must do better."