In Rome about 200 flights never got off the ground on Thursday, while the action disrupted 196 at Milan's Malpensa airport.
Unions began a 12-hour strike and plan more stoppages until 3 May when the state-controlled airline's management is expected to finalise its restructuring plan.
Labour leaders and political parties also appeared to seize the occasion to increase popular discontent against the centre-right government of Prime Minister Silvio Berlusconi ahead of European parliamentary elections in June.
The airline registered pre-tax losses of €510.6 million last year, compared with €260.5 million the previous year, while turnover fell by 9% in the same period to €4.321 billion.
PR man ignored
In February, Berlusconi's government blocked one plan that called for 1500 lay-offs and the outsourcing of 1200 more jobs.
The government also forced out Alitalia's chief executive and replaced him with Marco Zanichelli, a labour relations expert.
Nearly 400 flights were grounded
due to 12 hour strike
However, Zanichelli was drowned out by protesters on Wednesday when he attempted to address Alitalia workers at Rome's Fiumicino airport.
Fiat's car plants in Italy were also at a standstill again as members of FIOM, the metalworkers union, continued to set up blockades around factories.
The union also claimed that a four-hour nationwide strike in support of Fiat workers was widely followed.
Trade unions have sought a review of wage and scheduling conditions at Fiat's plant in Melfi, southern Italy.
Fiat met union leaders last night in Rome and said it was willing to review all labour conditions so long as the blockades were removed.