Karol Fijałkowski

Identification and methicillin resistance of coagulase-negative staphylococci isolated from nasal cavity of healthy horses

The aim of this study was an analysis of the staphylococcal flora of the nasal cavity of 42 healthy horses from 4 farms, along with species identification of CoNS isolates and determination of resistance to 18 antimicrobial agents, particularly phenotypic and genotypic methicillin resistance. From the 81 swabs, 87 staphylococci were isolated. All isolates possessed the gap gene but the coa gene was not detected in any of these isolates. Using PCR-RFLP of the gap gene, 82.8% of CoNS were identified: S. equorum (14.9%), S. warneri (14.9%), S. sciuri (12.6%), S. vitulinus (12.6%), S. xylosus (11.5%), S. felis (5.7%), S. haemolyticus (3.4%), S. simulans (3.4%), S. capitis (1.1%), S. chromogenes (1.1%), and S. cohnii subsp. urealyticus (1.1%). To our knowledge, this was the first isolation of S. felis from a horse. The species identity of the remaining Staphylococcus spp. isolates (17.2%) could not be determined from the gap gene PCR-RFLP analysis and 16S rRNA gene sequencing data. Based on 16S–23S intergenic transcribed spacer PCR, 11 different ITS-PCR profiles were identified for the 87 analyzed isolates. Results of API Staph were consistent with molecular identification of 17 (19.5%) isolates. Resistance was detected to only 1 or 2 of the 18 antimicrobial agents tested in the 17.2% CoNS isolates, including 6.9% MRCoNS. The mecA gene was detected in each of the 5 (5.7%) phenotypically cefoxitin-resistant isolates and in 12 (13.8%) isolates susceptible to cefoxitin. In total, from 12 horses (28.6%), 17 (19.5%) MRCoNS were isolated. The highest percentage of MRCoNS was noted among S. sciuri isolates (100%).

Effects of 50 Hz rotating magnetic field on the viability of Escherichia coli and Staphylococcus aureus

Abstract This study presents results of research on the influence of rotating magnetic field (RMF) of the induction of 30 mT and the frequency of 50 Hz on the growth dynamics and cell metabolic activity of E. coli and S. aureus, depending on the exposure time. The studies showed that the RMF caused an increase in the growth and cell metabolic activity of all the analyzed bacterial strains, especially in the time interval t = 30 to 150 min. However, it was also found that the optical density and cell metabolic activity after exposition to RMF were significantly higher in S. aureus cultures. In turn, the study of growth dynamics, revealed a rapid and a significant decrease in these values from t = 90 min) in the case of E. coli samples. The obtained results prove that RMF (B = 30 mT, f = 50 Hz) has a stimulatory effect on the growth and metabolic activity of E. coli and S. aureus. Furthermore, taking into account the time of exposure, stronger influence of RMF on the viability was observed in S. aureus cultures, which may indicate that this effect depends on the shape of the exposed cells.

Modification of bacterial cellulose through exposure to the rotating magnetic field

The aim of the study was to assess the influence of rotating magnetic field (RMF) on production rate and quality parameters of bacterial cellulose synthetized by Glucanacetobacter xylinus. Bacterial cultures were exposed to RMF (frequency f=50Hz, magnetic induction B=34mT) for 72h at 28°C. The study revealed that cellulose obtained under RMF influence displayed higher water absorption, lower density and less interassociated microfibrils comparing to unexposed control. The application of RMF significantly increased the amount of obtained wet cellulose pellicles but decreased the weight and thickness of dry cellulose. Summarizing, the exposure of cellulose-synthesizing G. xylinus to RMF alters cellulose biogenesis and may offer a new biotechnological tool to control this process. As RMF-modified cellulose displays better absorbing properties comparing to non-modified cellulose, our finding, if developed, may find application in the production of dressings for highly exudative wounds.

Effects of rotating magnetic field exposure on the functional parameters of different species of bacteria.

Abstract The aim of the present study was to determine the effect of the rotating magnetic field (RMF) on the growth, cell metabolic activity and biofilm formation by S. aureus, E. coli, A. baumannii, P. aeruginosa, S. marcescens, S. mutans, C. sakazakii, K. oxytoca and S. xylosus. Bacteria were exposed to the RMF (RMF magnetic induction B = 25–34 mT, RMF frequency f = 5–50 Hz, time of exposure t = 60 min, temperature of incubation 37 °C). The persistence of the effect of exposure (B = 34 mT, f = 50 Hz, t = 60 min) on bacteria after further incubation (t = 300 min) was also studied. The work showed that exposure to RMF stimulated the investigated parameters of S. aureus, E. coli, S. marcescens, S. mutans, C. sakazakii, K. oxytoca and S. xylosus, however inhibited cell metabolic activity and biofilm formation by A. baumannii and P. aeruginosa. The results obtained in this study proved, that the RMF, depending on its magnetic induction and frequency can modulate functional parameters of different species of bacteria.

Staphylococci isolated from ready-to-eat meat - Identification, antibiotic resistance and toxin gene profile.

The aim of this study was to analyse the staphylococci isolated from ready-to-eat meat products, including pork ham, chicken cold cuts, pork sausage, salami and pork luncheon meat, sliced in the store to the consumers specifications, along with species identification and determination of antibiotic resistance. Genes encoding staphylococcal enterotoxins, staphylococcal enterotoxin-like proteins, exfoliative toxins, and toxic shock syndrome toxin 1 were also investigated. From the 41 samples, 75 different staphylococcal isolates were obtained. Based on PCR-RFLP analysis of the gap gene using AluI and HpyCH4V restriction enzymes, the isolates were identified as Staphylococcus equorum (28%), S. vitulinus (16%), S. carnosus (14%), S. succinus (11%), S. xylosus (11%), S. saprophyticus (9%), S. warneri (9%), S. haemolyticus (1%) and S. pasteuri (1%). The incidence and number of resistances to antimicrobials was found to be species but not source of isolation dependent. All S. xylosus, S. saprophyticus, S. haemolyticus and S. pasteuri isolates showed antibiotic resistance. A lower percentage of resistance was recorded for S. warneri (71%) and S. vitulinus (58%), followed by S. equorum (57%), S. carnosus (50%) and S. succinus (50%). The most frequent resistance was observed to fusidic acid (43%). The mecA gene was amplified in 4% of the staphylococci. However, phenotypic resistance to methicillin was not confirmed in any of these isolates. On the other hand, the mecA gene was not detected in any of 9% of the isolates resistant to cefoxitin. It was also found that among 75 isolates, 60 (80%) harbored from 1 to 10 out of 21 analyzed superantigenic toxin genes. The most prevalent genes were: sei (36% isolates) among enterotoxins, seln (32% isolates) among enterotoxin-like proteins and eta encoding exfoliative toxin A (37% isolates). The findings of this study further extend previous observations that, when present in food, not only S. aureus but also other species of staphylococci could be of public health significance.

Comparative Analysis of Superantigen Genes in Staphylococcus xylosus and Staphylococcus aureus Isolates Collected from a Single Mammary Quarter of Cows with Mastitis

The purpose of this study was to analyze and compare genes encoding superantigens (SAgs) in Staphylococcus xylosus and Staphylococcus aureus isolates collected simultaneously from milk of the same cows with clinical mastitis. Genes encoding staphylococcal enterotoxins and enterotoxin-like proteins (sea-selu), toxic shock syndrome toxin 1 (tst-1) and exfoliative toxins (eta and etd) were investigated. It was found that among 30 isolates of S. xylosus, 16 (53.3%) harbored from 1 to 10 SAg genes. In total, in 16 SAg positive S. xylosus, 11 different enterotoxin genes were detected: sec, sed, seg, seh, sei, selm, seln, selo, selp, ser, selu and one etd gene encoding exfoliative toxin D. The most prevalent genes were ser, selu, and selo. Among all the positive isolates of S. xylosus, a total of 14 different SAg gene combinations were detected. One combination was repeated in 3 isolates, whereas the rest were detected only once. However, in the case of S. aureus all the 30 isolates harbored the same combination of SAg genes: seg, sei, selm, seln, selo and on the basis of PFGE analysis all belonged to the same clonal type. Also noteworthy was the observation that SAg genes detected in S. aureus have also been found in S. xylosus. The findings of this study further extend previous observations that SAg genes are present not only in S. aureus but also in coagulase-negative staphylococci, including S. xylosus. Therefore, taking into account that the SAg genes are encoded on mobile genetic elements it is possible that these genes can be transferred between different species of coexisting staphylococci.

Secretory virulence factors produced by Staphylococcus aureus isolates obtained from mastitic bovine milk--effect on bovine polymorphonuclear neutrophils.

The aim of the research was to test whether exogenic virulence factors secreted by Staphylococcus aureus isolates are involved in mechanisms that allow the bacteria to modulate and evade phagocytosis by bovine polymorphonuclear neutrophils. The research was based on the comparison of the effects of supernatants, prepared from cultures of 30 S. aureus isolates, on the functional properties of bovine neutrophils in vitro. S. aureus isolates were collected from milk samples from cows with clinical mastitis. Supernatants, which were used to treat leukocytes, were prepared from 18 h S. aureus cultures. Exogenic virulence factors secreted by S. aureus isolates significantly influenced the phagocytosis parameters evaluated. Depending on their leukotoxic or superantigenic properties, supernatants could affect the ingestion process, and also showed an influence on the digestion efficiency and phagocytosis carried out by bovine polymorphonuclear neutrophils in vitro.

The effect of auto-vaccination therapy on the phenotypic variation of one clonal type of Staphylococcus aureus isolated from cows with mastitis.

The aim of this study was to demonstrate the effect of auto-vaccine therapy on selected properties of Staphylococcus aureus strains, isolated from milk of cows with subclinical mastitis. The experiment was based on auto-vaccines which were prepared from S. aureus strains isolated from 16 cows. S. aureus strains isolated from cows on the 7th, 21st and 35th day following auto-vaccination, were analyzed phenotypically and genotypically. The isolated strains represented 17 biotypes all belonging to one clonal type. Increases of new biotypes of S. aureus were detected on the 35th day of therapy. Among 48 re-isolated strains, 18.75% (9/48) revealed single and 12.50% (6/48) multiple phenotypical changes. The present study demonstrated that during auto-vaccine therapy, S. aureus strains can change phenotypically, pointing out the necessity for using precise diagnostic methods, that would make possible a better assessment of the used therapy.

Synthesis and antibacterial activity of Schiff bases and amines derived from alkyl 2-(2-formyl-4-nitrophenoxy)alkanoates

A series of novel Schiff bases and secondary amines were obtained in good yields, as a result of the reductive amination of alkyl 2-(2-formyl-4-nitrophenoxy)alkanoates with both aniline and 4-methoxyaniline under established mild reaction conditions. Sodium triacetoxyborohydride as well as hydrogen in the presence of palladium on carbon were used as efficient reducing agents of the Schiff bases, in both direct and stepwise reductive amination processes. The Schiff bases, amines, and amine hydrochlorides were designed as potential antibacterial agents, and structure–activity relationship could be established following in vitro assays against Gram-positive and Gram-negative bacteria. The minimal inhibitory concentration and zone of inhibition were also determined. In these tests, some of Schiff bases and secondary amine hydrochlorides showed moderate-to-good activity against Gram-positive bacteria, including S. aureus, M. luteus, and S. mutans.

Increased water content in bacterial cellulose synthesized under rotating magnetic fields

ABSTRACT The current study describes properties of bacterial cellulose (BC) obtained from Komagataeibacter xylinus cultures exposed to the rotating magnetic field (RMF) of 50 Hz frequency and magnetic induction of 34 mT for controlled time during 6 days of cultivation. The experiments were carried out in the customized RMF exposure system adapted for biological studies. The obtained BC displayed an altered micro-structure, degree of porosity, and water-related parameters in comparison to the non-treated, control BC samples. The observed effects were correlated to the duration and the time of magnetic exposure during K. xylinus cultivation. The most preferred properties in terms of water-related properties were found for BC obtained in the setting, where RMF generator was switched off for the first 72 h of cultivation and switched on for the next 72 h. The described method of BC synthesis may be of special interest for the production of absorbent, antimicrobial-soaked dressings and carrier supports for the immobilization of microorganisms and proteins.