Outgoing Liberal Concetta Fierravanti-Wells laid into the prime minister during a fiery late-night parliamentary speech on Tuesday.
The senator - who lost preselection for her NSW upper house seat at the weekend - claimed Mr Morrison was "not fit to be prime minister" hours after the federal budget was revealed.
One Nation senator Pauline Hanson on Wednesday backed up the claims made by Senator Fierravanti-Wells, calling the prime minister a bully.
But Liberal senator Jane Hume said that was "an easy accusation for someone who has it in for the government".
The senator - who is also superannuation minister - said she has never been bullied by the prime minister.
"I can safely say this prime minister is not a bully and he has been extraordinarily supportive of all the women I work with," she told ABC Radio on Thursday.
"Yet these things seem to fall by the wayside when an easy accusation is made by somebody who has it in for this government."
Independent senator Jacqui Lambie said the accusations have nothing to do with personal grievances.
"(Morrison) is probably one of the most unpleasant men I've ever had to sit in front of," she told Sky News on Thursday.
"He is a bully, he is intimidating and that's the truth of the matter and good on Connie (Fierravanti-Wells) for calling him out."
Senator Lambie says accusations about the prime minister's character have floated around parliament for a while.
"We've all been talking about Scott Morrison up here - some of us females for a long time - and it needs to come out for the election on what sort of bully this man is," she said.
"This bloke has a real problem in relating to women and when he picks people to do the job, it's not on merit or on credit, it is whether or not you bow down to Scott and kiss his backside. That is how it works in the Liberal Party these days."
Mr Morrison said he understood Senator Fierravanti-Wells' disappointment at missing out on preselection.
"But (in) politics, on occasion, people have disappointments, and so I obviously don't agree with her assessment," he told 6PR radio.
"But if she has any formal complaint she'd like to make, in the Liberal Party we have processes for dealing with that, and I'd encourage her to do so."
He said he had always sought to work "politely and co-operatively" with Senator Hanson.
Former Liberal prime minister John Howard told the Sydney Morning Herald and The Age that Mr Morrison may be "forceful" but he was not a bully.
"I haven't seen any evidence that (Mr Morrison) is a bully, that he's arrogant or any of that. Forceful? Well, anybody who gets to be the leader of a political party is forceful," he said.
But Labor frontbencher Tanya Plibersek said she was not surprised Mr Howard was defending the prime minister given an election is due to be called.
"To be honest, I don't think Scott Morrison would be bullying John Howard," she said.
"The way bullies work is not usually that they bully people who are more powerful than them - they look for people they think will respond to intimidation."
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