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Summary The Prototheca algae have recently emerged as an important cause of bovine mastitis globally. Here, we present results of a first large‐scale, cross‐country survey on the prevalence of Prototheca spp. in dairy cows, and their environment in Poland. A total of 1211 samples were collected and microbiologically analysed. Included within this number were milk ( n = 638), body swabs ( n = 374) and environmental samples ( n = 199), originating from 400 dairy cows and their surroundings, on 16 dairy farms, based in all major provinces of the country. Prototheca spp. were the third, after Streptococcus and Staphylococcus spp., most common mastitis pathogens. The overall prevalence of protothecal mastitis was 8.3% (33/400), with the majority (75.8%) of cases having a subclinical course, and all but one attributable to P. zopfii genotype 2. Prototheca spp. were cultured from body swabs of both healthy and mastitic cows, yet the isolation rate among the latter was conspicuously lower (12.3% vs. 17.8%). Forty‐two (21.2%) environmental samples yielded growth of Prototheca spp. However, no clear association between Prototheca mastitis in dairy cows and the algal isolation from the herd environment was found. Nor was there any association between the environmental recovery of the algae and farm management practices.

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Achlorophyllous unicellular microalgae of the genus (, ) are the only known plants that cause infections in both humans and animals, collectively referred to as protothecosis. Human protothecosis, most commonly manifested as cutaneous, articular, and disseminated disease, is primarily caused by, followed by and, sporadically, by and In veterinary medicine, however, is a major pathogen responsible for bovine mastitis, which is a predominant form of protothecal disease in animals. Historically, identification of spp. has relied upon phenotypic criteria; these were later replaced by molecular typing schemes, including DNA sequencing. However, the molecular markers interrogated so far, mostly located in the ribosomal DNA (rDNA) cluster, do not provide sufficient discriminatory power to distinguish among all spp. currently recognized. Our study is the first attempt to develop a fast, reliable, and specific molecular method allowing identification of all spp. We propose the mitochondrial gene as a new and robust marker for diagnostics and phylogenetic studies of the algae. The gene displayed important advantages over the rDNA markers. Not only did the gene have the highest discriminatory capacity for resolving all species, but it also performed best in terms of technical feasibility, understood as ease of amplification, sequencing, and multiple alignment analysis. Based on the species-specific polymorphisms in the partial gene, we developed a fast and straightforward PCR-restriction fragment length polymorphism (RFLP) assay for identification and differentiation of all species described so far. The newly proposed method is advocated to be a new gold standard in diagnostics of protothecal infections in human and animal populations.

Currently, mutations in three genes, namely rrs, rpsL, and gidB, encoding 16S rRNA, ribosomal protein S12, and 16S rRNA-specific methyltransferase, respectively, are considered to be involved in conferring resistance to streptomycin (STR) in Mycobacterium tuberculosis. The aim of this study was to investigate the spectrum and frequency of these mutations in M. tuberculosis clinical isolates, both resistant and susceptible to STR. Sixty-four M. tuberculosis isolates recovered from as many TB patients from Poland in 2004 were included in the study. Within the sample were 50 multidrug-resistant (32 STR-resistant and 18 STR-susceptible) and 14 pan-susceptible isolates. Preliminary testing for STR resistance was performed with the 1% proportion method. The MICs of STR were determined by the Etest method. Mutation profiling was carried out by amplifying and sequencing the entire rrs, rpsL, and gidB genes. Non-synonymous mutations in either rrs or rpsL gene were detected in 23 (71.9%) of the STR-resistant and none of the STR-susceptible isolates. Mutations in the gidB gene were distributed among 12 (37.5%) STR-resistant and 13 (40.6%) STR-susceptible isolates. Four (12.5%) STR-resistant isolates were wild-type at all three loci examined. None of the rrs, rpsL or gidB mutations could be linked to low, intermediate or high level of STR resistance. In accordance with previous findings, the gidB 47T→G (L16R) mutation was associated with the Latin American-Mediterranean genotype family, whereas 276A→C (E92D) and 615A→G (A205A) mutations of the gidB gene were associated with the Beijing lineage. The study underlines the usefulness of rrs and rpsL mutations as molecular markers for STR resistance yet not indicative of its level. The gidB polymorphisms can serve as phylogenetic markers.