Dunedin to Brisbane is off the flight path when Virgin Australia returns to the New Zealand market later this year.
Before Covid-19, Virgin Australia flew two to three direct weekly flights from Dunedin, which were stopped when the airline suspended all New Zealand services in March 2020.
Airline spokesman Kris Taute said the Dunedin route would not be making a return in the short to medium term.
Depending on how international air travel recovered over the next few years, the company could look at a return, he said.
Air New Zealand has not ruled out taking over the route.
During a visit to Dunedin yesterday, chairwoman Dame Therese Walsh said the airline "considers everything".
Air New Zealand was flying all of the routes to Australia it did pre-Covid since the travel bubble had opened.
Dame Therese acknowledged the route was important to Dunedin.
"I know it is something that is subject to discussion in Dunedin, whether Virgin will continue and what the alternative plans might be," she said.
"So we certainly keep discussing the options."
Air New Zealand's Dunedin domestic routes were back to 90% of pre-Covid levels.
Dunedin Airport was performing "above the national average"compared to other airports the carrier flew to in the country, she said.
Air New Zealand's board had its annual out-of-Auckland board meeting in Dunedin on Wednesday and spent two days in the city, which included being hosted for a business breakfast by the Otago Chamber of Commerce.
Between 40,000 and 45,000 passengers crossed the Tasman from Dunedin Airport each year on the Virgin flight.
Dunedin Airport business development general manager Megan Crawford said the airport had worked on a transtasman strategy about getting the route up and running again last year.
The airport was hopeful Dunedin and the wider lower South Island would be serviced by the route again one day, but it was not as simple as an airline coming straight into the route and picking it up, she said.
Enterprise Dunedin director John Christie said the transtasman service had been seen as "quite desirable" by Dunedin travellers.
"We would obviously like to see a service on that route continue but it is a result of having Covid and the decisions of airlines to look at different routes which means Virgin have decided not to continue with that service which is, in the current environment, something that is understandable," Christie said.
"It will have an impact, I guess, on bringing direct passengers to and from Brisbane and vice-versa.
"The Australian market is an important market for some of our local operators.
"We'd like to see any of those routes into the east coast of Australia be part of the Dunedin offering and I'd like to think, as we see those opportunities with borders opening, airlines will respond to some of those new routes including Air New Zealand and any other airline that might looking at using our facilities."