The concern was raised with MSPs one week after a report showed an increase in the number of patients waiting beyond four hours in A&E across Scotland.
The health service has rarely achieved the Scottish Government's 98% standard, although there are signs of improvement.
A lower interim target of 95% was set in April last year in an attempt to boost performance by September this year.
MSPs on Holyrood's Public Audit Committee quizzed the Auditor General on her findings in a meeting today.
"What we are seeing, and what we say in the report, is that there are indications of pressure in the system," she told them.
Fourteen of the 31 A&E departments met the four-hour target in December last year.
"We believe a number of boards will find it hard to meet the 95% target by September of this year," she told the committee.
Further concern was raised about a spike in activity across hospitals in the last 10 minutes of the four-hour period.
More than 18% of admissions from A&E at the Royal Infirmary of Edinburgh were recorded just before the time limit passed.
Ms Gardner said extra work will be carried out to explore what is happening to patients during the course of their wait.
"We are planning some work on unscheduled work more generally," she said.
"As part of that we will want to talk to some patients again."
Scotland has 31 A&E departments which saw about 1.35 million patients in 2012-13 at a cost of about £163 million, Audit Scotland said in its report on May 8.
About 104,000 people waited beyond the standard four-hour target in 2012-13, compared with about 36,000 in 2008-09.
The proportion of people being seen within the four-hour target fell from 97.2% at the end of 2009 to 93.5% by December last year.
The report shows variation in demand at A&E across the country.
Gilbert Bain Hospital in Lerwick, Shetland, and the Royal Aberdeen Children's Hospital were the only two to meet the 98% target in each month of the financial year 2012-13. Most other hospitals failed in every month.
The report noted action is being taken by the Scottish Government and that the situation has improved in recent months.
The Government announced a national plan in February last year to improve emergency care.
The NHS plans to invest about £50 million in unscheduled care between 2013-14 and 2015-16.
Health Secretary Alex Neil said there has been an 87% reduction in patients waiting over 12 hours.