Two fifths of doctors, nurses and other frontline staff at Buckinghamshire Healthcare are not vaccinated against flu, figures reveal.
NHS England has urged healthcare workers to get vaccinated to protect themselves and their patients, pushing for "near universal" coverage.
By the end of December 2018, 2,363 front-line NHS workers at Buckinghamshire Healthcare NHS Trust had not had the jab, according to Public Health England.
Of the 5,958 doctors, nurses, clinical staff and support workers at the trust with direct patient contact, 3,595 were vaccinated between September and December.
That's an uptake rate of 60%, lower than the national average of 66%.
Last winter, NHS England praised trusts that achieved coverage of 90% or above at the end of the season.
Bosses have now warned that staff refusing the vaccine could be banned from treating patients.
As a result, trusts are now required to submit data on how many of their staff refuse the jab.
Buckinghamshire Healthcare NHS Trust reported that 271 staff turned down the jab - making up more than 10% of unvaccinated staff.
The other staff may not have been offered the vaccination yet.
Coverage at the trust has improved since the same period in 2017-18, when 51% of front-line staff were vaccinated.
St Helens and Knowsley Hospitals, in Merseyside, was the only trust to hit 90% coverage by the end of December.
Oxleas Trust in south east London and the Birmingham and Solihull Mental Health Trust both had an uptake rate of just 35%.
Dr Nick Scriven, president of the Society for Acute Medicine, said the variation in reported uptake was "disturbing".
He said: "I can see employers taking a firm stance with anyone who chooses not to be vaccinated and who then contracts influenza.
"The NHS workforce crisis is well-described, and anything that could further exacerbate that is to be avoided."
PHE medical director Professor Paul Cosford said: "Healthcare professionals have a responsibility to be vaccinated to protect themselves and their patients from flu.
"Front-line health and social care workers are at more risk of catching flu because of their contact with patients.
"They are also more likely to pass it on to their patients, many of whom will be at higher risk of serious complications of flu."
NHS England said staff have achieved a record level of uptake for the vaccine this winter.
Across the United Kingdom, there have been 1,684 hospital admissions and 102 deaths due to flu since October 2018.