The Effect of Salvia tomentosa Miller Extracts, Rich in Rosmarinic, Salvianolic and Lithospermic Acids, on Bacteria Causing Opportunistic Infections.

Methanolic-aqueous extracts of Salvia tomentosa Miller roots, aerial parts, and inflorescences were examined for their content of polyphenolic derivatives and the antimicrobial and cytotoxic effect. In the polyphenolic-rich profile, rosmarinic, salvianolic, and lithospermic acids along with various derivatives were predominant. A total of twenty phenolic compounds were identified using the UPLC/DAD/qTOF-MS technique. These were caffeic acid, rosmarinic acid derivatives, lithospermic acid derivatives, salvianolic acids B, F, and K derivatives, as well as sagerinic acid, although rosmarinic acid (426-525 mg/100 g of dry weight-D.W.) and salvianolic acid B (83-346.5 mg/100 g D.W.) were significantly predominant in the metabolic profile. Strong antibacterial activity of S. tomentosa extracts was observed against Staphylococcus epidermidis (MIC/MBC = 0.625 mg/mL) and Bacillus cereus (MIC = 0.312-1.25 mg/mL). The extracts showed low cytotoxicity towards the reference murine fibroblasts L929 and strong cytotoxicity to human AGS gastric adenocarcinoma epithelial cells in the MTT reduction assay. The observed cytotoxic effect in cancer cells was strongest for the roots of 2-year-old plant extracts.

Decreased susceptibility to vancomycin and other mechanisms of resistance to antibiotics in Staphylococcus epidermidis as a therapeutic problem in hospital treatment.

Multidrug-resistant coagulase-negative staphylococci represent a real therapeutic challenge. The aim of the study was to emphasize the importance of heteroresistance to vancomycin presence in methicillin-resistant strains of S. epidermidis. The research comprised 65 strains of S. epidermidis. Heteroresistance to vancomycin was detected with the use of the agar screening method with Brain Heart Infusion and a population profile analysis (PAP test). In addition, types of cassettes and genes responsible for resistance to antibiotics for 22 multidrug resistant strains were determined. Our investigations showed that 56 of 65 S. epidermidis strains were phenotypically resistant to methicillin. The tested strains were mostly resistant to erythromycin, gentamicin, clindamycin, and ciprofloxacin. Six strains showed decreased susceptibility to vancomycin and their heterogeneous resistance profiles were confirmed with the PAP test. All tested multi-resistant strains exhibited the mecA gene. More than half of them possessed type IV cassettes. ant(4')-Ia and aac(6')/aph(2''), ermC and tetK genes were most commonly found. The described phenomenon of heteroresistance to vancomycin in multidrug resistant bacteria of the Staphylococcus genus effectively inhibits a therapeutic effect of treatment with this antibiotic. That is why it is so important to search for markers that will enable to identify heteroresistance to vancomycin strains under laboratory conditions.

Phenotypic and Genotypic Characterization of Antimicrobial Resistance in Streptococci Isolated from Human and Animal Clinical Specimens.

Recently, the phenomenon of infection of humans as hosts by animal pathogens has been increasing. Streptococcus is an example of a genus in which bacteria overcome the species barrier. Therefore, monitoring infections caused by new species of human pathogens is critical to their spread. Seventy-five isolates belonging to streptococcal species that have recently been reported as a cause of human infections with varying frequency, were tested. The aim of the study was to determine the drug resistance profiles of the tested strains, the occurrence of resistance genes and genes encoding the most important streptococcal virulence factors. All tested isolates retained sensitivity to ß-lactam antibiotics. Resistance to tetracyclines occurred in 56% of the tested strains. We have detected the MLSB type resistance (cross-resistance to macrolide, lincosamide, and streptogramin B) in 20% of the tested strains. 99% of the strains had tetracycline resistance genes. The erm class genes encoding MLSB resistance were present in 47% of strains. Among the strains with MLSB resistance, 92% had the streptokinase gene, 58% the streptolysin O gene and 33% the streptolysin S gene. The most extensive resistance concerned isolates that accumulated the most traits and genes, both resistance genes and virulence genes, increasing their pathogenic potential. Among the tested strains, the gene encoding streptokinase was the most common. The results of the prove that bacteria of the species S. uberis, S. dysgalactiae and S. gallolyticus are characterized by a high pathogenic potential and can pose a significant threat in case of infection of the human body.

Vancomycin heteroresistance among methicillin-resistant clinical isolates S. haemolyticus, S. hominis, S. simulans, and S. warneri.

Besides being an essential part of the skin microbiome, coagulase-negative staphylococci are the etiological factors of serious infections. The aim of the study was to evaluate the heteroresistance to vancomycin and the potential antimicrobial efficacy of teicoplanin and daptomycin against the multiresistant strains of S. haemolyticus, S. hominis, S. warneri, and S. simulans. The study covered 80 clinical coagulase-negative staphylococci. Teicoplanin, vancomycin, and daptomycin MICs for the tested strains were determined according to EUCAST recommendation. The vanA and vanB genes were searched. The brain heart infusion screen agar method detected vancomycin heteroresistance. The population analysis profile method and analysis of autolytic activity were applied for the strains growing on BHI containing 4 mg/L vancomycin. Seven S. haemolyticus, two S. hominis, and two S. warneri strains presented a heterogeneous resistance to vancomycin. Their subpopulations were able to grow on a medium containing 4-12 mg/L of vancomycin. Monitoring heteroresistance to peptide antibiotics, which are often the last resort in staphylococcal infections, is essential due to the severe crisis in antibiotic therapy and the lack of alternatives to treat infections with multiresistant strains. Our work highlights the selection of resistant strains and the need for more careful use of peptide antibiotics.

Pathogenic potential and antimicrobial resistance of Staphylococcus pseudintermedius isolated from human and animals.

Crossing of interspecies barriers by microorganisms is observed. In recent years, Staphylococcus pseudintermedius-a species formerly thought to be animal-has also been isolated from human clinical materials. Many virulence factors are responsible for the colonization, which is the first step an infection, of the new host organism. We analyzed the factors influencing this colonization as well as susceptibility to antibiotics in fourteen S. pseudintermedius strains isolated from clinical cases from humans and animals. The occurrence of genes responsible for binding elastin, fibronectin, and fibrinogen and some phenotypic features, although different between strains, is comparable in both groups. However, the animal isolates had more genes coding for virulence factors. All isolates tested had the exfoliating toxin gene and the leukotoxin determining genes, but only the human strains had enterotoxin genes. The assessment of antibiotic resistance of strains of both groups indicates their broad resistance to antibiotics commonly used in veterinary medicine. Antibiotic resistance was more common among animal isolates. The multilocus sequence typing analysis of the studied strains was performed. The results indicated a large diversity of the S. pseudintermedius population in both studied groups of strains. Equipped with important virulence factors, they showed the ability to infect animals and humans. The clonal differentiation of the methicillin-susceptible strains and the multidrug resistance of the strains of both studied groups should be emphasized. The considerable genetic diversity of strains from a limited geographical area indicates the processes of change taking place within this species. Thus, careful observation of the ongoing process of variation is necessary, as they may lead to the selection of S. pseudintermedius, which will pose a significant threat to humans.

Design, Synthesis, Anti-Varicella-Zoster and Antimicrobial Activity of (Isoxazolidin-3-yl)Phosphonate Conjugates of N1-Functionalised Quinazoline-2,4-Diones.

Dipolar cycloaddition of the N-substituted C-(diethoxyphosphonyl)nitrones with N3-allyl-N1-benzylquinazoline-2,4-diones produced mixtures of diastereoisomeric 3-(diethoxyphosphonyl)isoxazolidines with a N1-benzylquinazoline-2,4-dione unit at C5. The obtained compounds were assessed for antiviral and antibacterial activities. Several compounds showed moderate inhibitory activities against VZV with EC50 values in the range of 12.63-58.48 µM. A mixture of isoxazolidines cis-20c/trans-20c (6:94) was found to be the most active against B. cereus PCM 1948, showing an MIC value 0.625 mg/mL, and also was not mutagenic up to this concentration.

Influence of Agronomic Practice on Total Phenols, Carotenoids, Chlorophylls Content, and Biological Activities in Dry Herbs Water Macerates.

Oregano (Origanum vulgare L.) and thyme (Thymus vulgaris L.) have long been known for their organoleptic properties. Both plants are widely used in cuisine worldwide in fresh and dried form and as a pharmaceutical raw material. The study aimed to assess if the type of cultivation influenced chosen chemical parameters (total polyphenols by Folin-Ciocalteu method; carotenoids and chlorophyll content by Lichtenthaler method), antimicrobial activity (with chosen reference microbial strains) and shaped cytotoxicity (with L929 mouse fibroblasts cell line) in water macerates of dry oregano and thyme. Polyphenols content and antimicrobial activity were higher in water macerates obtained from conventional cultivation (independently from herb species), unlike the pigments in a higher amount in macerates from organic herbs cultivation. Among all tested macerates stronger antimicrobial properties (effective in inhibiting the growth of Pseudomonas aeruginosa, Bacillus cereus and Salmonella enteritidis) and higher cytotoxicity (abilities to diminish the growth of L929 fibroblasts cytotoxicity) characterized the conventionally cultivated thyme macerate.

Pathogenicity and drug resistance of animal streptococci responsible for human infections.

Bacteria of the genus Streptococcus, earlier considered typically animal, currently have also been causing infections in humans. It is necessary to make clinicians aware of the emergence of new species that may cause the development of human diseases. There is an increasing frequency of isolation of streptococci such as S. suis, S. dysgalactiae, S. iniae and S. equi from people. Isolation of Streptococcus bovis/Streptococcus equinus complex bacteria has also been reported. The streptococcal species described in this review are gaining new properties and virulence factors by which they can thrive in new environments. It shows the potential of these bacteria to changes in the genome and the settlement of new hosts. Information is presented on clinical cases that concern streptococcus species belonging to the groups Bovis, Pyogenic and Suis. We also present the antibiotic resistance profiles of these bacteria. The emerging resistance to ß-lactams has been reported. In this review, the classification, clinical characteristics and antibiotic resistance of groups and species of streptococci considered as animal pathogens are summarized.

Transformed Shoots of Dracocephalum forrestii W.W. Smith from Different Bioreactor Systems as a Rich Source of Natural Phenolic Compounds.

Transformed shoots of the Tibetan medicinal plant Dracocephalum forrestii were cultured in temporary immersion bioreactors (RITA and Plantform) and in nutrient sprinkle bioreactor (NSB) for 3 weeks in MS (Murashige and Skoog) liquid medium with 0.5 mg/L BPA (N-benzyl-9-(2-tetrahydropyranyl)-adenine) and 0.2 mg/L IAA (indole-3-acetic acid). The greatest biomass growth index (GI = 52.06 fresh weight (FW) and 55.67 dry weight (DW)) was observed for shoots in the RITA bioreactor, while the highest multiplication rate was found in the NSB (838 shoots per bioreactor). The levels of three phenolic acids and five flavonoid derivatives in the shoot hydromethanolic extract were evaluated using UHPLC (ultra-high performance liquid chromatography). The predominant metabolite was rosmarinic acid (RA)-the highest RA level (18.35 mg/g DW) and total evaluated phenol content (24.15 mg/g DW) were observed in shoots grown in NSB. The NSB culture, i.e., the most productive one, was evaluated for its antioxidant activity on the basis of reduction of ferric ions (ferric reducing antioxidant power, FRAP) and two scavenging radical (O2•- and DPPH, 1,1-diphenyl-2-picrylhydrazyl radical) assays; its antibacterial, antifungal, and antiproliative potential against L929 cells was also tested (3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium bromide (MTT) test). The plant material revealed moderate antioxidant and antimicrobial activities and demonstrated high safety in the MTT test-no cytotoxicity at concentrations up to 50 mg/mL was found, and less than a 20% decrease in L929 cell viability was observed at this concentration.

Staphylococcal species less frequently isolated from human clinical specimens - are they a threat for hospital patients?

BACKGROUND: Coagulase-negative staphylococci belonging to S. haemolyticus, S. hominis subsp. hominis, S. simulans, and S. warneri are often described as etiological factors of infections. Staphylococci are a phylogenetically coherent group; nevertheless, there are differences among the species which may be important to clinicians. METHODS: We investigated selected virulence factors and antibiotic resistance that were phenotypically demonstrated, the presence and expression of genes encoding the virulence factors, and the type of the SCCmec cassette. RESULTS: The differences between the tested species were revealed. A great number of isolates produced a biofilm and many of them contained single icaADBC operon genes. Clear differences between species in the lipolytic activity spectrum could be related to their ability to cause various types of infections. Our studies also revealed the presence of genes encoding virulence factors homologous to S. aureus in the analysed species such as enterotoxin and pvl genes, which were also expressed in single isolates of S. simulans and S. warneri. S. haemolyticus and S. hominis subsp. hominis isolates were resistant to all clinically important antibiotics including ß-lactams. The identified SCCmec cassettes belonged to IV, V, VII, and IX type but most of the detected cassettes were non-typeable. Among the investigated species, S. hominis subsp. hominis isolates accumulated virulence genes typical for S. aureus in the most efficient way and were widely resistant to antibiotics. CONCLUSIONS: Our results clearly indicated significant differences between the tested species, which might be a result of the horizontal gene transfer (HGT) and can lead to the formation and selection of multi-drug resistant strains as well as strains with new virulence features. Such strains can have a new clinical relevance.

Differences in distribution of MLS antibiotics resistance genes in clinical isolates of staphylococci belonging to species: S. epidermidis, S. hominis, S. haemolyticus, S. simulans and S. warneri.

BACKGROUND: Macrolides and lincosamides are two leading types of antibiotics commonly used in therapies. The study examines the differences in resistance to these antibiotics and their molecular bases in S. epidermidis as well as in rarely isolated species of coagulase-negative staphylococci such as S. hominis, S. haemolyticus, S. warneri and S. simulans. The isolates were tested for the presence of the erm(A), erm(B), erm(C), lnu(A), msr(A), msr(B), mph(C), ere(A) and ere(B) genes. Phenotypic resistance to methicillin and mecA presence were also determined. RESULTS: The MLSB resistance mechanism was phenotypically found in isolates of species included in the study. The most prevalent MLSB resistance mechanism was observed in S. hominis, S. haemolyticus and S. epidermidis isolates mainly of the MLSB resistance constitutive type. Macrolide, lincosamide and streptogramin B resistance genes were rarely detected in isolates individually. The erm(B), ere(A) and ere(B) genes were not found in any of the strains. The erm(A) gene was determined only in four strains of S. epidermidis and S. hominis while lnu(A) was seen in eight strains (mainly in S. hominis). The erm(C) gene was present in most of S. epidermidis strains and predominant in S. hominis and S. simulans isolates. The examined species clearly differed between one another in the repertoire of accumulated genes. CONCLUSIONS: The presence of genes encoding the MLSB resistance among CoNS strains demonstrates these genes' widespread prevalence and accumulation in opportunistic pathogens that might become gene reservoir for bacteria with superior pathogenic potential.

New Gene Responsible for Resistance of Clinical Corynebacteria to Macrolide, Lincosamide and Streptogramin B.

The subject of the study was phenotypic marking of the antibiotic susceptibility and MLSB resistance mechanism in Corynebacterium spp. isolated from human skin (18 isolates) and from clinical materials (19 isolates). The strains were tested for the presence of the erm(A), erm(B), erm(C), erm(X), lnu(A), msr(A), msr(B) and mph(C) genes. Clinical isolates showed wide resistance to antibiotics. In 89% clinical isolates and 72% skin microbiota a constitutive type of MLSB resistance was found. In 12 clinical isolates the erm(C) gene was detected-eight of which had erm(X) as well as erm(C), two harboured erm(X), erm(C) and erm(A) and two demonstrated only erm(C).

[Animal Staphylococcus felis with the potential to infect human skin]. / Zwierzecy Staphylococcus felis o potencjale zakazania skóry czlowieka.

INTRODUCTION: Breaking interspecies barrier by microorganisms has become in the recent years an alarming phenomenon that threatens public health worldwide. An important potential interspecies transmission risk factor is close contact animal-human including occupational exposure of pet breeders and veterinarians. MATERIAL AND METHODS: The features of Staphylococcus felis ZMF 13 strain isolated from a swab from a cat's wound connected with potential pathogenicity were investigated. Results: The virulence factors of strain found were hydroxamate siderophores, production of invasins - intracellular proteolytic and lipolytic enzymes and the ability of biofilm production. The ability of bacteriocin-like substance production was also observed. The substance has an antagonistic activity against bacteria belong to physiological flora of the human skin which may be important in breaking the colonization resistance of human organism. Although the strain of S. felis ZMF 13 was methicillin-susceptible it demonstrated the constutive type of MLSB resistance mechanism. The genes ermA, msrB, linA connected with macrolide, lincosamides and streptogramin B resistance were detected. CONCLUSIONS: The current evidence suggest that Staphylococcusfelis has a number of features that can be crucial in its potential interspecies transmission.

Macrolide, lincosamide, and streptogramin B resistance in lipophilic Corynebacteria inhabiting healthy human skin.

Corynebacteria exist as part of human skin microbiota. However, under some circumstances, they can cause opportunistic infections. The subject of the study was to examine the macrolide-lincosamide-streptogramin B (MLSB) antibiotic resistance in 99 lipophilic strains of Corynebacterium genus isolated from the skin of healthy men. Over 70% of the tested strains were resistant to erythromycin and clindamycin. All of which demonstrated a constitutive type of MLSB resistance mechanism. In all strains, there were being investigated the erm(A), erm(B), erm(C), erm(X), lin(A), msr(A), and mph(C) genes that could be responsible for the different types of resistance to marcolides, lincosamides, and streptogramin B. In all strains with the MLSB resistance phenotype, the erm(X) gene was detected. None of the other tested genes were discovered. Strains harboring the erm(X) gene were identified using a phenotypic method based on numerous biological and biochemical tests. Identification of the chosen strains was compared with the results of API Coryne, MALDI-TOF MS, and 16S rDNA sequencing methods. Only 7 out of the 23 investigated resistant strains provided successful results in all the used methods, showing that identification of this group of bacteria is still a great challenge. The MLSB resistance mechanism was common in most frequently isolated from healthy human skin Corynebacterium tuberculostearicum and Corynebacterium jeikeium strains. This represents a threat as these species are also commonly described as etiological factors of opportunistic infections.

Species-specific sensitivity of coagulase-negative Staphylococci to single antibiotics and their combinations.

The activity of beta-lactam antibiotics (oxacillin, cloxacillin, cephalotin), vancomycin, gentamicin and rifampicin applied in vitro individually and in combination against 37 nosocomial methicillin-resistant strains of coagulase-negative staphylococci (CNS) was assessed to demonstrate the heterogeneity of this group of bacteria and estimate the chance of the efficacy of such therapy. The strains belonged to four species: Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus cohnii, Staphylococcus hominis. They originated from a hospital environment and from the skin of medical staff of the intensive care unit of a paediatric ward at a university hospital. All strains were methicillin-resistant, according to CLSI standards, but individual strains differed in MIC(ox) values. Susceptibility to other tested antibiotics was also characteristic for the species. The increased susceptibility to antibiotics in combinations, tested by calculating the fractional inhibitory concentration (FIC) index, concerned 26 out of 37 investigated strains and it was a feature of a particular species. Combinations of vancomycin and cephalotin against S. epidermidis and oxacillin with vancomycin were significant, as well as cephalotin and rifampicin in growth inhibition of multiresistant S. haemolyticus strains.