Preliminary Results, Perspectives and Proposal for a Screening Method of in vitro Susceptibility of Prototheca spp. to Antimicrotubular Agents

Microorganisms belonging to the genus Prototheca are achlorophyllousmicroalgae, occasionally behaving as environmental pathogens that cause severemastitis in milk cows, as well as localized or systemic infections in humans and animals.Among the different species belonging to the genus, Prototheca zopfii genotype2 (recently reclassified as P. bovis) and P. blaschkeae are most commonly associatedwith bovine mastitis. To date, no pharmacological treatment is available to cureprotothecal mastitis, and infected animals must be quarantined to avoid spreadingthe infection. The few antibiotic and antifungal drugs effective in vitro against Protothecagive poor results in vivo. This failure is likely due to the lack of specificity ofsuch drugs. As microalgae are more closely related to plants than to bacteria orfungi, an alternative possibility is to test molecules with herbicidal properties, in particular,antimicrotubular herbicides, for which plant rather than animal tubulin is theselective target. Once a suitable test protocol was set up, a panel of 11 antimicrotubularagents belonging to different chemical classes and selective for plant tubulinwere tested for the ability to inhibit growth of Prototheca cells in vitro. Two dinitroanilines,dinitramine and chloralin, showed strong inhibitory effects on P. blaschkeaeat low micromolar concentrations, with half-maximal inhibitory concentrations (IC50)of 4.5 and 3 M, respectively, while both P. zopfii genotype 1 (now reclassified as P.ciferrii) and P. bovis showed susceptibility to dinitramine only, to different degrees.Suitable screening protocols for antimitotic agents are suggested.