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Made available in DSpace on 2018-12-11T17:34:33Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-10-01 A strain of an achlorophyllic alga, named PR24T, was isolated from cow milk samples from the state of Minas Gerais, Brazil. Based on 18S rDNA, 28S rRNA, D1/D2 region of the LSU rDNA and SSU rRNA gene sequence similarities, this strain was found to be a member of the genus Prototheca and closely related to Prototheca blaschkeae SAG2064T. However, the novel strain could easily be distinguished from recognized Prototheca species by internal transcribed spacer, species-specific PCR, single-strand conformation polymorphism-PCR analysis and phenotypic characteristics. The inability to grow in Sabouraud broth at pH 4.0 and the different cellular fatty acid composition clearly distinguished PR24T from the reference strain of P. blaschkeae. The combination of genotypic and phenotypic data indicates that strain PR24T represents a subspecies of P. blaschkeae, for which the name Prototheca blaschkeae subsp. brasiliensis subsp. nov. is proposed. The respective type strain is PR24T (=DSM 103592T=IHEM 26958T). Institute of Sciences of Food Production Italian National Research Council (CNR ISPA), Via Celoria 2 Institute of Agricultural Biology and Biotechnology Italian National Research Council (CNR IBBA), Via Einstein Council for Agricultural Research and Economics – Research Centre for Animal Production and Aquaculture (CREA-ZA), Via Lombardo 11 Department of Veterinary Hygiene and Public Health Universidade Estadual Paulista (UNESP) Department of Veterinary Medicine Federal University of Lavras Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia Romagna (IZSLER), Via Einstein Department of Veterinary Hygiene and Public Health Universidade Estadual Paulista (UNESP)

Dairy cow mastitis associated with microalgae of the genus Prototheca has been reported worldwide. This alga is extremely resistant to most antimicrobials commonly used in mastitis therapy. In milk processing, different thermal treatments are generally efficient at inactivating and eliminating microorganisms. Until recently, no reports on Prototheca blaschkeae susceptibility to heat treatment have been described. Thus, considering the potential zoonotic risk that Prototheca may represent, the objective of this study was to test the susceptibility of P. blaschkeae field isolates retrieved from bovine mastitis to different temperature/time ratios that are generally used in the milk processing industry: 62°C/15 min and 30 min; 70°C/20 s, 15 min, and 30 min; 75°C/20 s; 90°C/1 s; and 100°C/1 s. The results showed a growth reduction of all isolates after the heat treatments, but only at 100°C was a total growth inhibition observed.

We describe the first known case of an algaemia by Prototheca blaschkeae in a dairy cow, which occurred after a chronic episode of mastitis caused by this pathogen. The organism was isolated from milk, joint fluid and blood samples, and microbiologic and molecular methods were performed to obtain a definitive identification of the algae. The affected cow was culled only after confirmation of a systemic infection by Prototheca.

One hundred sixty-one Prototheca spp. strains isolated from composite milk and barn-surrounding environmental samples (bedding, feces, drinking, or washing water, surface swabs) of 24 Italian dairy herds were characterized by genotype-specific PCR analysis. Overall, 97.2% of strains isolated from composite milk samples were characterized as Prototheca zopfii genotype 2, confirming its role as the main mastitis pathogen, whereas Prototheca blaschkeae was only sporadically isolated (2.8%). Regarding environmental sampling, 84.9% of isolates belonged to P. zopfii genotype 2, 13.2% to P. blaschkeae, and 1.9% to P. zopfii genotype 1. The data herein contradict previous hypotheses about the supposed exclusive role of P. zopfii genotype 2 as the causative agent of protothecal mastitis and, on the contrary, confirm the hypothesis that such pathology could be caused by P. blaschkeae in a few instances.