Enterococcus hirae belongs in the Enterococcus faecium group within the genus Enterococcus. This species occurs naturally in the environment, commensally in the alimentary tracts of animals, and pathologically for example in humans with urinary infections. Some strains of E. hirae possess virulence factors, including biofilm formation. Biofilm growth protects bacteria against host de- fences; biofilm can be a source of persistent infection. Testing bacterial strains for their ability to form biofilm might therefore facilitate their treatment or prevention. This study focuses on bio- film formation by E. hirae strains derived from various animals. This kind of testing has never been done before. A total of 64 identified E. hirae from laying hens, ducks, pheasants, ostriches, rabbits, horses and a goat were tested by means of three methods; using Congo red agar, the tube method and microtiter plate agar. The majority of strains were found to form biofilm. 62.5% of strains were biofilm-forming, four categorized as highly positive (OD570 ≥1); most strains were low-grade biofilm positive (0.1 ≤ OD 570 < 1). Related to poultry, 55 E. hirae strains were tested nd found to produce biofilm; 24 strains did not form biofilm, 31 strains were biofilm-forming; 27 strains showed low-grade biofilm formation, and four strains were highly biofilm-forming. Four strains from hens and ostriches reached the highest OD570 values, more than 0.500. Rabbit-derived E. hirae strains as well as strains isolated from horses and the goat were low-grade bio- film-forming. Microtiter plate assay proved to be the best tool for testing the in vitro biofilm for- mation capacity of E. hirae strains from different species of animals.

Microbiological studies were carried out of atmospheric air sampled on the area and in the surroundings of a mechanical and biological wastewater treatment plant (WTP) treating municipal sewage. The capacity of the wastewater treatment plant, which also received some wastewater from the dairy industry, was ca 3· 103 m3d-1. Counts ofheterotrophic psychrophilic, psychrotrophic and mesophilic bacteria as well as some physiological groups of microorganisms which belong to Enterobacteriaceae family, Staphylococcus and Enterococcus genera, Pseudomonas fluorescens and P. aeruginosa species, hemolysing bacteria and actinomycetes were analyzed. Air samples were collected in summer, autumn, winter and spring seasons simultaneously by the sedimentation and impact methods at 6 sites located on the area of the WTP and at 5 sites situated in its surroundings. The background was established depending on the direction of wind, always on the windward side in relation to the location of the WTP. In addition, temperature and air humidity as well as wind speed and direction at each sampling sites were observed. Statistically significant differences were found in studied groups of microorganisms counts between air samples collected in different seasons of the year (with the exception of psychrophilic bacteria and by the two different methods (with the exception of psychrophilic bacteria) and microorganisms which belong to Enterobacteriaceae family). The highest mean counts of the microorganisms were usually determined in air samples collected by the sedimentation method, especially during the autumn (with the exception of actinomycetes, which are the most numerous in spring), the lowest ones in winter and/or in summer. No statistically significant differences were observed in counts of the analyzed groups of microorganisms in air sampled at particular sites (with the exception of Enterobacteriaceae bacteria isolated on Chromocult medium). However, higher counts of these microorganisms were typically found in the air sampled in the area of the WTP, particularly near the grit chamber, phosphorus removal tank, nitrification and denitrification chambers and secondary settling tank. According to the Polish Standards used for evaluation of atmospheric air pollution, the air sampled in the area of wastewater treatment plant and in its surroundings was classified as only slightly and sporadically strongly polluted. It was mainly in the spring and autumn seasons that the air was strongly polluted with psychrophilic and mesophilic bacteria. No increased emission of the analyzed groups of microorganisms, including faecal bacteria was determined in the air samples collected outside the WT

Faecal Enterococcus hirae from domestic ducks were studied for their bioactivity to select bioactive strain for more detailed study with its probable use in poultry and also to bring novelty in basic research. After defecation, faeces (n=23, faecal mixture of 40 ducks) were sampled from domestic ducks in eastern Slovakia; birds were aged from eight to 14 weeks. E. hirae strains were identified using Matrix-assisted laser desorption/ionization time-of flight mass spectrometry with a highly probable species identification score (2.300-3.000) or a secure genus identification/ /probable species identification score (2.000-2.299), confirmed by polymerase chain reaction and phenotypization in accordance with the properties for the type strain E. hirae ATCC 9790. Strains were hemolysis negative (γ-hemolysis), and did not have active enzyme stimulating disorders. Enterocin genes were detected in three strains out of seven. Three out of four Enterocin genes were detected in Kč1/b (Ent A, P, L50A); the most frequently detected was the Ent P gene. The strains inhibited indicator strains E. faecalis, listeriae, but also Escherichia coli and Buttiauxiella strains. Lactic-acid producing E. hirae were mostly susceptible to antibiotics. Based on parameter evaluation, E. hirae Kč1/b, Kč6 can be additionally studied to select the type of bioactive substance.