New Thiosemicarbazide Derivatives with Multidirectional Biological Action.

Over the years, several new medicinal substances have been introduced for the treatment of diseases caused by bacteria and parasites. Unfortunately, due to the production of numerous defense mechanisms by microorganisms and parasites, they still pose a serious threat to humanity around the world. Therefore, laboratories all over the world are still working on finding new, effective methods of pharmacotherapy. This research work aimed to synthesize new compounds derived from 3-trifluoromethylbenzoic acid hydrazide and to determine their biological activity. The first stage of the research was to obtain seven new compounds, including six linear compounds and one derivative of 1,2,4-triazole. The PASS software was used to estimate the potential probabilities of biological activity of the newly obtained derivatives. Next, studies were carried out to determine the nematocidal potential of the compounds with the use of nematodes of the genus Rhabditis sp. and antibacterial activity using the ACCT standard strains. To determine the lack of cytotoxicity, tests were performed on two cell lines. Additionally, an antioxidant activity test was performed due to the importance of scavenging free radicals in infections with pathogenic microorganisms. The conducted research proved the anthelmintic and antibacterial potential of the newly obtained compounds. The most effective were two compounds with a 3-chlorophenyl substituent, both linear and cyclic derivatives. They demonstrated higher efficacy than the drugs used in treatment.

Antioxidant and antimicrobial properties of an extract rich in proteins obtained from Trametes versicolor.

Introduction: Bioactive proteins and peptides generated from fruit, vegetables, meat or fish have great potential as functional food or substitutes for antibiotics. In recent years it has also been demonstrated that the fungus kingdom could be a source of these compounds. The study investigated the bioactivity of an extract of the lignicolous fungus Trametes versicolor and its hydrolysate. Material and Methods: The fungus was collected in a mixed forest in October, extracted and hydrolysed. To inspect the protein and peptide profiles before and after hydrolysis, matrix-assisted laser desorption/ionisation-time-of-flight mass spectrometric analysis was performed. To evaluate the antioxidant properties of the preparations, 2,2-diphenyl-1-picrylhydrazyl (DPPH•) and 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS•+) radical scavenging assays were used. The activity of the fungus extract and hydrolysate against Aeromonas veronii, Bacillus cereus, Enterococcus faecalis, Enterococcus faecium, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella Typhimurium, Staphylococcus aureus, Staphylococcus epidermidis, Streptococcus agalactiae, Streptococcus dysgalactiae, and Streptococcus uberis was determined by the minimum inhibitory concentration and minimum bactericidal concentration values. Results: The extract and its hydrolysate showed almost 100% ABTS•+ and DPPH• radical scavenging with a low half maximal inhibitory concentration. The water extract and hydrolysate of T. versicolor exhibited antimicrobial activity against two S. aureus strains, E. coli, P. aeruginosa and Salmonella Typhimurium. Conclusion: These results provide compelling evidence that the analysed fungus extract and its hydrolysate hold promise with their antibacterial and antioxidant properties.

Prevalence of Culturable Bacteria and Yeasts in the Nasopharynx Microbiota during the Physiological Course of Pregnancy.

The aim of the study was to compare the prevalence of the nasopharyngeal carriage of culturable microorganisms in the microbiota of asymptomatic women with a physiological pregnancy (PW) and nonpregnant women (NPW). Nasopharyngeal swabs were collected from 53 PW and 30 NPW to detect bacterial and fungal colonization. Isolates were identified using the culture method and the MALDI-TOF MS technique. The nasopharyngeal microbiota (NPM) partially differed between PW and NPW. These differences in the frequency of nasopharyngeal colonization between the PW and NPW groups were not statistically significant (p > 0.05); all cases were colonized by bacteria and only two cases in the PW group were colonized by yeasts, namely, Rhodotorula spp. High levels of staphylococcal colonization, including predominantly coagulase-negative staphylococci and S. aureus in the nasopharyngeal sample, were present in both groups. The reduced number of Gram-negative rods colonized in the cases studied was seen in samples from the NPW group, particularly with Enterobacterales, and anaerobic Cutibacterium spp. were isolated only in the PW group (p < 0.05). Moreover, a higher carriage rate of Enterobacter aerogenes colonization was statistically significant (p < 0.05) and correlated with the NPW group. Pregnancy may disturb the composition of the NPM represented by commensals and opportunistic bacteria and promote yeast colonization as compared to nonpregnant women.

Staphylococcal Resistance Patterns, blaZ and SCCmec Cassette Genes in the Nasopharyngeal Microbiota of Pregnant Women.

Antimicrobial resistance in Staphylococcus spp. colonising the nasopharynx can create risk factors of therapeutic treatment failure or prophylaxis in pregnant women. Resistance is mostly encoded on plasmids (e.g., blaZ gene for penicillinase synthesis) or chromosomes (e.g., mecA and mecC for methicillin resistance). The mecA gene is part of the chromosomal mec gene cassette (SCCmec), which is also located on the plasmid. The disc diffusion method for the selected drugs (beta-lactams, fluoroquinolones, streptogramins, aminoglicosides, macrolides, oxasolidinones, tetracyclines and other groups) was used. PCR for blaZ, mecA and mecC genes and SCCmec cassette detection and typing were performed. S. aureus (54.4%) and S. epidermidis (27.9%) were the most prevalent and showed the highest diversity of resistance profiles. The blaZ, mecA and mecC genes were reported in 95.6%, 20.6% and 1.5% of isolates, respectively. The highest resistance was found to beta-lactams, commonly used during pregnancy. Resistance to a variety of antimicrobials, including benzylpenicillin resistance in blaZ-positive isolates, and the existence of a very high diversity of SCCmec cassette structures in all staphylococci selected from the nasopharyngeal microbiota of pregnant women were observed for the first time. Knowledge of the prevalence of antimicrobial-resistant staphylococci in the nasopharynx of pregnant women may be important for the appropriate treatment or prophylaxis of this group of patients.

Synthesis and Biological Evaluation of New Schiff Bases Derived from 4-Amino-5-(3-fluorophenyl)-1,2,4-triazole-3-thione.

The treatment of infectious diseases is a challenging issue faced by the medical community. The emergence of drug-resistant strains of bacteria and fungi is a major concern. Researchers and medical professionals are working to develop new and innovative treatments for infectious diseases. Schiff bases are one a promising class of compounds. In this work, new derivatives were obtained of the 4-amino-5-(3-fluorophenyl)-2,4-dihydro-3H-1,2,4-triazole-3-thione reaction, with corresponding benzaldehydes with various substituents at position 4. The antibacterial and antifungal activities of all synthesized compounds were tested. Several new substances have shown moderate antifungal activity against Candida spp. The highest activity directed against C. albicans was shown by compound RO4, with a 4-methoxyphenyl moiety and an MIC value of 62.5 µg/mL. In order to check the toxicity of the synthesized compounds, their effect on cell lines was examined. Additionally, we tried to elucidate the mechanism of the antibacterial and antifungal activity of the tested compounds using molecular docking to topoisomerase IV, D-Alanyl-D-Alanine Ligase, and dihydrofolate reductase.

Plasmid-Mediated Fluoroquinolone Resistance Genes in Quinolone-Susceptible Aeromonas spp. Phenotypes Isolated From Recreational Surface Freshwater Reservoir.

Aeromonas spp. are recognized as opportunistic pathogens causing diseases. Infections in humans can result mainly in gastrointestinal and wound diseases with or without progression to septicemia. Although Aeromonas spp. are not known uropathogens and they rarely cause urinary tract infection, we hypothesize that the presence of these bacteria in the water and the contact during, e.g., recreational and bathing activity can create the conditions for the colonization of the human body and may result to diseases in various locations, including the urinary tract. Our study presents the occurrence of aeromonad fluoroquinolone-susceptible phenotypes with the presence of plasmid-mediated fluoroquinolone resistance (PMQR) genes in a natural freshwater reservoir occasionally used for recreational activities. Sixty-nine isolates collected during the bathing period were identified by mass spectrometry and screened for the presence of fluoroquinolone-resistant phenotypes and genotypes. Fluoroquinolone susceptibility was determined as minimal inhibitory concentration values. PMQR qnr genes were detected by PCR. Isolates comprising eight species, namely, mainly Aeromonas veronii (50.7% isolates) and Aeromonas media (24.6% isolates) and rarely Aeromonas eucrenophila, Aeromonas caviae, Aeromonas bestiarum, Aeromonas ichthiosmia, and Aeromonas hydrophila, were selected. All isolates were phenotypically susceptible either to ciprofloxacin or levofloxacin. Unexpectedly, at least one to three of the PMQR genes were detected in 42.0% of the fluoroquinolone-susceptible Aeromonas spp. phenotypes. Mainly the qnrS (34.8% isolates) and qnrA (14.5% isolates) determinants were detected. In conclusion, the freshwater reservoir occasionally used for bathing was tainted with aeromonads, with a high occurrence of opportunistic pathogens such as A. veronii and A. media. MALDI-TOF MS is a powerful technique for aeromonad identification. Our data reveals the mismatch phenomenon between fluoroquinolone-susceptible aeromonad phenotypes and the presence of plasmid-mediated qnr resistance genes. It suggests that phenotypically susceptible bacteria might be a potential source for the storage and transmission of these genes. The exposure during, e.g., a recreational activity may create the potential risk for causing infections, both diagnostically and therapeutically difficult, after expressing the resistance genes and quinolone-resistant strain selection.

Opportunistic Pathogens of Recreational Waters with Emphasis on Antimicrobial Resistance-A Possible Subject of Human Health Concern.

Infections caused by exposure to opportunistic pathogens can cause serious health problems during recreational water use. The problem of diseases caused by microbes transmitted by water is a major public health challenge, especially in developing countries with economic problems and poor hygiene conditions. Moreover, the quality of water in natural reservoirs is often at a very low level in terms of microbiological water purity, which means that their use for recreational purposes, but also as a source of drinking water, may have serious health consequences. Recreational waters pose a threat to human health. Therefore, the quality of recreational waters is closely monitored in many jurisdictions. In this review, we summarize key information on the most common pathogens that can be water-based or waterborne. The issue of antimicrobial resistance among opportunistic pathogens remains equally important. It is important not only to fight pathogens, but also to take action to reduce chemical stressors (especially antibiotics) in the aquatic environment, and to understand the various mechanisms of the spread of antibiotic-resistant genes.

Antimicrobial resistance and genetic diversity of Enterococcus faecalis from yolk sac infections in broiler chicks.

Despite restrictions on the use of antibiotics in poultry, the percentage of multidrug resistant bacteria, isolated from both adult birds and chicks, remains high. These bacteria can spread between countries via hatching eggs or chicks. Antibiotic resistant bacteria can also pose a threat to hatchery and farm workers or to consumers of poultry. The aim of the study was to perform a phenotypic and genotypic analysis of the drug resistance of E. faecalis isolates from yolk sac infections in broiler chicks from Poland and the Netherlands and to determine their genetic diversity. The tests revealed resistance to antibiotics from category D, that is, tetracycline (69.7%); category C - lincomycin (98.7%), erythromycin (51.3%), aminoglycosides (high-level streptomycin and kanamycin resistance - 10.5% and 3.95%, respectively), and chloramphenicol (7.9%); and category B - ciprofloxacin (25% with resistance or intermediate resistance). No resistance to penicillin, ampicillin, high-level gentamicin, tigecycline, or linezolid was noted. Various combinations of the erm(B), tet(M), tet(L), tet(O), ant(6)-Ia, aph(3')-IIIa, ant(4')-Ia, cat, and msr(A/B) genes were detected in all isolates (irrespective of the drug-resistance phenotype). Among isolates that carried the tet(M) and/or the tet(L) gene, 28% also had the Int-Tn gene, in contrast with isolates possessing tet(O). There were 28 sequence types and 43 PFGE restriction patterns. About 60% of isolates were of sequences types ST59, ST16, ST116, ST282, ST36, and ST82. Nine new sequence types were shown (ST836-ST844). In conclusion, broiler chicks can be a source of drug-resistant sequence types of E. faecalis that are potentially hazardous for people and animals. Restrictive programs for antibiotic use in broiler breeding flocks should be developed to decrease drug resistance in day-old chicks and reduce economic losses during rearing.

Phenotypic and genotypic characterization of Enterococcus spp. from yolk sac infections in broiler chicks with a focus on virulence factors.

Bacterial infections of yolk sacs contribute to increased mortality of chicks, chronic infections during their rearing, or increased selection in the flock, which in turn leads to high economic losses in poultry production worldwide. The aim of this study was a phenotypic and genotypic characterization of enterococci isolated from yolk sac infections (YSI) of broiler chickens from Poland and the Netherlands. Biochemical, matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF) MS, and rpoA gene sequencing identification was performed. Moreover, phenotypic and genotypic characterization of virulence factors and analysis of the clonal relationship of isolates by MALDI-TOF MS and enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) were performed. The biochemical test identified 70 isolates as Enterococcus faecalis and 6 as Enterococcus mundtii. The results of MALDI-TOF MS were 100% concordant with those obtained by rpoA gene sequencing, and all 76 isolates were identified as E. faecalis. Differences were noted in the ß-glucuronidase, ß-glucosidase, α-galactosidase, phosphatase, melibiose, lactose, and raffinose tests that is going about the results of biochemical identification. None of the isolates were beta-hemolytic on blood agar in aerobic conditions, but all but one were gelatinase positive. Among biofilm-forming isolates (30/76; 39.5%), as many as 66.7% (20/30) were Polish E. faecalis strains. Most of the isolates carried virulence genes, that is gelE, ace, asa1, efaAfs, fsrA, fsrB, fsrC, cob, cpd, and ccf, but none had the hyl gene. Some isolates harbored cyl operon genes. One Polish strain (ST16) had all of the tested cyl genes and the esp gene, considered clinically important, and showed the highest biofilm-forming ability. Nearly 50% of the isolates showed close genetic relatedness in ERIC typing. In contrast with MALDI-TOF MS cluster analysis, ERIC-PCR results did not show a relationship with the origin of the strains. Using MALDI-TOF MS, 7 peaks were found in Polish and Dutch isolates, which may type them as species-specific biomarkers in E. faecalis from YSI.

Prevalence of susceptibility patterns of opportunistic bacteria in line with CLSI or EUCAST among Haemophilus parainfluenzae isolated from respiratory microbiota.

The application of CLSI and EUCAST guidelines led to many discrepancies. Various doubts have already appeared in preliminary stages of microbiological diagnostics of Haemophilus spp. A total of 87 H. parainfluenzae isolates were obtained from throat or nasopharyngeal swabs from adults 18 to 70 years old, both healthy volunteers and patients with chronic diseases between 2013 to 2015 in eastern Poland. Haemophilus spp. were identified by colony morphology, Gram-staining, API NH and MALDI-TOF MS technique. Both susceptibility to various antimicrobials and phenotypes of Haemophilus spp. resistance to beta-lactams were determined. Statistically significant association between applied guidelines and drug resistance patterns were observed to as follows: ampicillin, cefuroxime, cefotaxime, amoxicillin-clavulanate, azithromycin, tetracycline and trimethoprim-sulfamethoxazole. Resistance phenotypes according to CLSI vs. EUCAST were as follows: 3.4% vs. 8.0% for BLNAR and 6.9% vs. 19.5% for BLPACR isolates. In conclusion, this is the first study that reports comparative analysis of drug susceptibility interpretation using CLSI and EUCAST of haemophili rods from human respiratory microbiota in Poland. In case of susceptible, increased exposure (formerly intermediate) category of susceptibility within H. parainfluenzae isolates we have observed EUCAST as more restrictive than CLSI. Moreover, BLNAI and BLPAI phenotype isolates have been observed, as well as BLPBR using only CLSI or EUCAST guidelines, respectively.

Antibacterial Activity of Fluorobenzoylthiosemicarbazides and Their Cyclic Analogues with 1,2,4-Triazole Scaffold.

The development of drug-resistant bacteria is currently one of the major challenges in medicine. Therefore, the discovery of novel lead structures for the design of antibacterial drugs is urgently needed. In this structure-activity relationship study, a library of ortho-, meta-, and para-fluorobenzoylthiosemicarbazides, and their cyclic analogues with 1,2,4-triazole scaffold, was created and tested for antibacterial activity against Gram-positive bacteria strains. While all tested 1,2,4-triazoles were devoid of potent activity, the antibacterial response of the thiosemicarbazides was highly dependent on substitution pattern at the N4 aryl position. The optimum activity for these compounds was found for trifluoromethyl derivatives such as 15a, 15b, and 16b, which were active against both the reference strains panel, and pathogenic methicillin-sensitive and methicillin-resistant Staphylococcus aureus clinical isolates at minimal inhibitory concentrations (MICs) ranging from 7.82 to 31.25 µg/mL. Based on the binding affinities obtained from docking, the conclusion can be reached that fluorobenzoylthiosemicarbazides can be considered as potential allosteric d-alanyl-d-alanine ligase inhibitors.

Prevalence of Resistance to ß-Lactam Antibiotics and bla Genes Among Commensal Haemophilus parainfluenzae Isolates from Respiratory Microbiota in Poland.

(1) Background: Beta-lactams are the most frequently used antimicrobials, and are the first-line drugs in many infectious diseases, e.g., pneumonia, otitis media. Due to this fact, various bacteria have developed resistance to this group of drugs. (2) Methods: Eighty-seven Haemophilus parainfluenzae isolates were obtained from adults 18-70 years old in eastern Poland. The presence of 10 bla genes and 2 substitutions in ftsI reported as the most frequent in H. parainfluenzae were analyzed. (3) Results: Among 57 beta-lactam-resistant isolates, 63.2% encoded bla genes; blaTEM-1 predominated (54.4%), followed by blaOXA (19.3%), blaDHA (12.3%), blaSHV (10.5%), blaGES (7.0%), blaCMY (5.3%), blaVEB (1.8%) and blaROB-1 (1.8%). Lys-526 was the most common substitution in ftsI gene. The resistance genotypes were as follows: gBLNAS (17.5%), low-gBLNAR I (1.8%), low-gBLNAR II (1.8%), gBLNAR II (15.8%), gBLPAS (15.8%), gBLPAR (19.3%), gBLPBS I (8.8%) and gBLPBS II (1.8%); (4) Conclusions: This has been the first study to report on the high diversity of bla genes in H. parainfluenzae isolates in Poland. High sensitivity and specificity of benzylpenicillin test, as well as PCR of bla genes were shown, indicating that these methods may be useful as tools for the rapid screening of beta-lactamase prevalence and resistance to beta-lactams among H. parainfluenzae isolated from respiratory microbiota.

Biofilm formation capacity and presence of virulence factors among commensal Enterococcus spp. from wild birds.

Enterococci are opportunistic pathogens that can form biofilms during infections and many virulence determinants are involved in this process. Although the virulence factors are often analysed in Enterococcus spp. from humans and food animals, little is known about gut enterococcal isolates from wild birds. Therefore, the determination of virulence factors among enterococci isolated from wild birds may provide new information about a possible source of infection for humans and animals or vice versa via the environment. We analysed different phenotypic and genotypic traits in enterococci from wild birds related to potential virulence in humans and animals and to evaluate biofilm formation and its relationship to virulence genes. The E. faecalis isolates were characterised by greater frequency of biofilm formation in BHI than E. faecium. There was a correlation between hydrophobicity and biofilm formation in BHI broth in E. faecalis. None of the isolates was haemolytic. The presence of some adhesion and gelatinase genes was detected in biofilm-positive isolates. The enterococcal pathogenic factors (esp, hyl, and cyl operon genes) did not seem to be necessary or sufficient for production of biofilm by analysed bacteria. Enterococcus species isolated from wild birds should be considered as a possible source of some virulence determinants.

Influence of Thiazolidine-2,4-Dione Derivatives with Azolidine or Thiosemicarbazone Moieties on Haemophilus spp. Planktonic or Biofilm-Forming Cells.

Biofilm, naturally formed by microorganisms as integrated surface-bound communities, is one of the reasons for the development of antimicrobial resistance. Haemophilus spp. are common and representative opportunistic Gram-negative rods forming from the upper respiratory tract microbiota. The aim of this paper was to evaluate the influence of thiazolidine-2,4-dionebased azolidine and chlorophenylthiosemicarbazone hybrids against both planktonic and biofilm-forming Haemophilus spp. cells. The in vitro activity against planktonic and biofilm-forming cells of the tested compounds were evaluated by using the broth microdilution method. These activities were detected against reference and clinical strains of Haemophilus spp. on the basis of MICs (minimal inhibitory concentrations) and MBICs (minimal biofilm inhibitory concentrations). In addition, anti-adhesive properties of these compounds were examined. The target compounds showed potential activity against planktonic cells with MIC = 62.5⁻500 mg/L and biofilm-forming cells with MBIC = 62.5⁻1000 mg/L. The observed anti-adhesive properties of the tested compounds were reversible during long-term incubation in a lower concentration of compounds.

Topinambur - new possibilities for use in a supplementation diet.

Jerusalem artichoke (Helianthus tuberosus Linne) is a tuberous perennial plant of the Asteraceae family, which originates from North America, and is also known as wild sunflower or topinambur (TPB). It is characterized by good tolerance to frost, drought and poor soil, strong resistance to pests and plant diseases. For ages it was cultivated due to being both an edible tuber and having healing properties. In folk medicine, TPB leaves are used for the treatment of bone fractures and pain. TPB tubers are rich in sugar and have therefore been used for the production of functional food ingredients, such as inulin. Moreover, TPB is one of the potential crops for bioenergy production, such as bioethanol, biobuthanol and biodiesel, and chemicals (lactic acid, butyric acid). A number of bioactive compounds from the above ground parts of this plant have been isolated which have demonstrating antifungal, antioxidant and anticancer activities. In recent years, a number of animal experiments have been carried out to assess the health properties of TPB. Obtained results show that TPB possess a wide spectrum of medical applications, e.g. reduction in the levels of plasma glucose, total cholesterol and triglyceride. Interestingly, TPB has been shown to be a valuable alternative source of prebiotic compounds. This review article presents recent scientific reports on the chemical and biological properties of TPB and its potential use as a prebiotic diet supplement.

Synthesis and antibacterial activity of new (2,4-dioxothiazolidin-5-yl/ylidene)acetic acid derivatives with thiazolidine-2,4-dione, rhodanine and 2-thiohydantoin moieties.

A series of new (2,4-dioxothiazolidin-5-yl/ylidene)acetic acid derivatives with thiazolidine-2,4-dione, rhodanine and 2-thiohydantoin moiety (28-65) were synthesized by the reaction of (2,4-dioxothiazolidin-5-yl/ylidene)acetic acid chlorides with 5-(hydroxybenzylidene) thiazolidine-2,4-dione, rhodanine and 2-thiohydantoin derivatives. Obtained compounds (28-65) were tested on reference strains of Gram-positive bacteria and ones of the Gram-negative bacteria. The antibacterial activity of target compounds was determined by broth microdilution method. These derivatives showed antibacterial activity generally against Gram-positive bacterial strains. Most active compounds possess MIC = 3.91 mg/L. Our results suggest that presence of electron-withdrawing substituent at phenyl ring is favorable while geometry of molecule does not play important role in antibacterial response. It was confirmed the lack of direct influence of substitution pattern at phenyl ring on antibacterial activity of closely related compounds of series 1-3. The antibacterial activity of some compounds was similar or higher than the activity of commonly used reference drugs such as oxacillin and cefuroxime.

Synthesis and Antibacterial Activity of New Thiazolidine-2,4-dione-Based Chlorophenylthiosemicarbazone Hybrids.

Series of new thiazolidine-2,4-dione-based chlorophenylthiosemicarbazone hybrids (17⁻40) were synthesized by the reaction of condensation chlorophenylthiosemicarbazides with formylphenyl 2-(2,4-dioxothiazolidin-5-yl/ylidene)acetates. New compounds were tested on reference strains of Gram-positive and Gram-negative bacteria. The antibacterial activity of target compounds was determined by broth dilution method. Most active compounds possess minimum inhibitory concentration (MIC) = 3.91 mg/L. These compounds were non-toxic at concentrations close to their antibacterial effect. The antibacterial activity of some compounds was similar to or higher than the activity of used reference drugs such as oxacillin and cefuroxime. The structure⁻activity relationships (SARs) analysis collectively suggests that at least two different molecular mechanisms of their antibacterial activity should be expected.

Rubber elastomeric nanocomposites with antimicrobial properties.

In this paper we show an elastomeric nanocomposite that exhibits antibacterial and antifungal activity. It comprises a rubber blend matrix and a nanofiller, which is a modified bentonite clay (Nanobent® ZR2). We have developed innovative technology for the nanofiller incorporation into the rubber matrix. This new approach was successfully implemented in pilot production at the Polish chemical manufacturer Spoiwo (Spoldzielnia Pracy Chemiczno-Wytworczej 'Spoiwo', Radom, Poland). Here we reveal that addition of the functionalised bentonite affects the mechanical and thermal behaviour of elastomers. For example, by adding 1-3% of bentonite nanoparticles we strongly enhanced elongation and tensile stress at break, whereas stiffness remained unchanged. We observed improvement of the thermal properties of the nanocomposites yielded and extension of the temperature usage range (from -29 to 311°C). As a proof of concept we present the antimicrobial effect of elastomeric nanocomposites verified on a wide range of both pathogenic and opportunistic reference bacteria strains, as well as reference strains of yeasts. The proposed method of hydrophilic nanofiller introduction into the rubber elastomer is economically viable and enables fabrication of elastomeric elements with high added value. Their significant antibacterial and antifungal activity makes them desirable in medicine, biomedical engineering, and the food industry.

Changes in the prevalence and biofilm formation of Haemophilus influenzae and Haemophilus parainfluenzae from the respiratory microbiota of patients with sarcoidosis.

BACKGROUND: Healthy condition and chronic diseases may be associated with microbiota composition and its properties. The prevalence of respiratory haemophili with respect to their phenotypes including the ability to biofilm formation in patients with sarcoidosis was assayed. METHODS: Nasopharynx and sputum specimens were taken in 31 patients with sarcoidosis (average age 42.6 ± 13), and nasopharynx specimens were taken in 37 healthy people (average age 44.6 ± 11.6). Haemophili were identified by API-NH microtest and by the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) system. Biofilm was visualised by crystal violet staining and confocal scanning laser microscopy (CSLM). The statistical analysis was performed with Statgraphics Plus for Windows. RESULTS: In total, 30/31 patients with sarcoidosis and 31/37 healthy people were colonized by Haemophilus influenzae (6/30 vs. 1/31) and Haemophilus parainfluenzae (28/30 vs. 31/31) in the nasopharynx. The overall number of nasopharyngeal haemophili isolates was 59 in patients with sarcoidosis and 67 in healthy volunteers (H. influenzae 6/59 vs. 1/67, P = 0.05; H. parainfluenzae 47/59 vs. 65/67, P = 0.0032). Moreover, the decreased number of H. parainfluenzae biofilm-producing isolates was shown in nasopharyngeal samples in patients with sarcoidosis as compared to healthy people (19/31 vs. 57/65, P = 0.006), especially with respect to isolates classified as strong and very strong biofilm-producers (8/31 vs. 39/65, P = 0.002). CONCLUSIONS: The obtained data suggest that the qualitative and quantitative changes within the respiratory microbiota concerning the overall prevalence of H. influenzae together with the decreased number of H. parainfluenzae strains and the decreased rate of H. parainfluenzae biofilm-producing isolates as compared to healthy people may be associated with sarcoidosis.

Inhibitory effect of 1,2,4-triazole-ciprofloxacin hybrids on Haemophilus parainfluenzae and Haemophilus influenzae biofilm formation in vitro under stationary conditions.

Haemophilus parainfluenzae and Haemophilus influenzae, upper respiratory tract microbiota representatives, are able to colonize natural and artificial surfaces as biofilm. The aim of the present study was to assay the effect of ten 1,2,4-triazole-ciprofloxacin hybrids on planktonic or biofilm-forming haemophili cells in vitro under stationary conditions on the basis of MICs (minimal inhibitory concentrations) and MBICs (minimal biofilm inhibitory concentrations). In addition, anti-adhesive properties of these compounds were examined. The reference strains of H. parainfluenzae and H. influenzae were included. The broth microdilution microtiter plate (MTP) method with twofold dilution of the compounds, or ciprofloxacin (reference agent) in 96-well polystyrene microplates, was used. The optical density (OD) reading was made spectrophotometrically at a wavelength of 570 nm (OD570) both to measure bacterial growth and to detect biofilm-forming cells under the same conditions with 0.1% crystal violet. The following values of parameters were estimated for 1,2,4-triazole-ciprofloxacin hybrids - MIC = 0.03-15.63 mg/L, MBIC = 0.03-15.63 mg/L, MBIC/MIC = 0.125-8, depending on the compound, and for ciprofloxacin - MIC = 0.03-0.06 mg/L, MBIC = 0.03-0.12 mg/L, MBIC/MIC = 1-2. The observed strong anti-adhesive properties (95-100% inhibition) of the tested compounds were reversible during long-term incubation at subinhibitory concentrations. Thus, 1,2,4-triazole-ciprofloxacin hybrids may be considered as starting compounds for designing improved agents not only against planktonic but also against biofilm-forming Haemophilus spp. cells.