Structure-activity relationship and cytotoxicity of the new thiosemicarbazide derivatives and their Cu(II) complexes against prostate and melanoma cancer cells.

In this study, eighteen new ligands (B1-B18) containing a thiosemicarbazide core were synthesized and characterized in terms of physicochemical properties, molecular docking and in vitro biological activity. The structures of eleven ligands were investigated using X-Ray diffraction and Hirschfeld Surface analysis. To study the structure-activity relationship, the organic ligands contained pyridin-2-ylmethyl, pyridin-3-ylmethyl or pyridin-4-ylmethyl moieties and various substituents. Their pharmakokinetic profiles and molecular docking results suggest high potential as new drug candidates. The complexing ability of the selected organic ligands was also evaluated, yielding five new Cu(II) complexes (Cu(B1)Cl2, Cu(B4)Cl2, Cu(B10)Cl2, Cu(B17)Cl2, Cu(B18)Cl2). The obtained results suggest the formation of the polymeric structures. All organic ligands and Cu(II) complexes were tested for anticancer activity against prostate and melanoma cancer cells (PC-3, DU-145, LNCaP, A375, G-361, SK-MEL-28) and normal fibroblasts (BJ), as well as antimicrobial activity against six selected bateria strains. Among B1-B18 compounds, B3, B5, B9, B10, B12 and B14 exhibited cytotoxic activity. The studied Cu(II) complexes were in general more active, with Cu(B1)Cl2 exhibiting antincancer activity agains all three prostate cancer cells and Cu(B10)Cl2 reaching the IC50 value equal to 88 µM against G-361 melanoma cells. Several compounds also exhibited antimicrobial activity against gram-positive and gram-negative bacteria. It was found that the type of specific substituents, especially the presence of -chloro and -dichloro substituents had a greated impact on the cytotoxicity than the position of the nitrogen atom in the pyridylacetyl moiety.

Synergistic Antifungal Interactions between Antibiotic Amphotericin B and Selected 1,3,4-thiadiazole Derivatives, Determined by Microbiological, Cytochemical, and Molecular Spectroscopic Studies.

In recent years, drug-resistant and multidrug-resistant fungal strains have been more frequently isolated in clinical practice. This phenomenon is responsible for difficulties in the treatment of infections. Therefore, the development of new antifungal drugs is an extremely important challenge. Combinations of selected 1,3,4-thiadiazole derivatives with amphotericin B showing strong synergic antifungal interactions are promising candidates for such formulas. In the study, microbiological, cytochemical, and molecular spectroscopy methods were used to investigate the antifungal synergy mechanisms associated with the aforementioned combinations. The present results indicate that two derivatives, i.e., C1 and NTBD, demonstrate strong synergistic interactions with AmB against some Candida species. The ATR-FTIR analysis showed that yeasts treated with the C1 + AmB and NTBD + AmB compositions, compared with those treated with single compounds, exhibited more pronounced abnormalities in the biomolecular content, suggesting that the main mechanism of the synergistic antifungal activity of the compounds is related to a disturbance in cell wall integrity. The analysis of the electron absorption and fluorescence spectra revealed that the biophysical mechanism underlying the observed synergy is associated with disaggregation of AmB molecules induced by the 1,3,4-thiadiazole derivatives. Such observations suggest the possibility of the successful application of thiadiazole derivatives combined with AmB in the therapy of fungal infections.

Variations in Blood Platelet Proteome and Transcriptome Revealed Altered Expression of Transgelin-2 in Acute Coronary Syndrome Patients.

Proteomic analyses based on mass spectrometry provide a powerful tool for the simultaneous identification of proteins and their signatures. Disorders detection at the molecular level delivers an immense impact for a better understanding of the pathogenesis and etiology of various diseases. Acute coronary syndrome (ACS) refers to a group of heart diseases generally associated with rupture of an atherosclerotic plaque and partial or complete thrombotic obstruction of the blood flow in the infarct-related coronary artery. The essential role in the pathogenesis of ACS is related to the abnormal, pathological activation of blood platelets. The multifactorial and complex character of ACS indicates the need to explain the molecular mechanisms responsible for thrombosis. In our study, we performed screening and comparative analysis of platelet proteome from ACS patients and healthy donors. Two-dimensional fluorescence difference gel electrophoresis and nanoscale liquid chromatography coupled to tandem mass spectrometry showed altered expressions of six proteins (i.e., vinculin, transgelin-2, fibrinogen ß and γ chains, apolipoprotein a1, and tubulin ß), with the overlapping increased expression at the mRNA level for transgelin-2. Dysregulation in protein expression identified in our study may be associated with an increased risk of thrombotic events, correlated with a higher aggregability of blood platelets and induced shape change, thus explaining the phenomenon of the hyperreactivity of blood platelets in ACS.

Antibacterial properties of Allium sativum L. against the most emerging multidrug-resistant bacteria and its synergy with antibiotics.

Garlic has long been known as the most effective plant species in treatment of bacterial infections. Considering the vast potential of garlic as a source of antimicrobial drugs, this study is aimed to evaluate the antibacterial activity of Allium sativum extracts and their interactions with selected antibiotics against drug-sensitive and multidrug-resistant isolates of emerging bacterial pathogens that are frequently found in healthcare settings. As shown by the in vitro data obtained in this study, the whole Allium sativum extract inhibited the growth of a broad range of bacteria, including multidrug-resistant strains with bactericidal or bacteriostatic effects. Depending on the organism, the susceptibility to fresh garlic extract was comparable to the conventional antibiotic gentamycin. Since the combinations of fresh garlic extract with gentamycin and ciprofloxacin inhibited both the drug sensitive and MDR bacteria, in most cases showing a synergistic or insignificant relationship, the potential use of such combinations may be beneficial, especially in inhibiting drug-resistant pathogens. The study results indicate the possibility of using garlic as e.g. a supplement used during antibiotic therapy, which may increase the effectiveness of gentamicin and ciprofloxacin.

Evaluation of the role of staphylococci in the pathomechanism of conjunctivitis.

PURPOSE: Determination of the association between ica genes and phenotypic biofilm formation in staphylococcal isolates involved in conjunctivitis, their antibiotic resistance as well as detection of selected virulence characteristics: adhesion to epithelial cells and in vitro cytotoxicity. METHODS: The study included 26 Staphylococcus aureus (SA) and 26 Staphylococcus epidermidis (SE) isolates. The presence of icaAD genes and ica operon was determined by the PCR assay. Phenotypic biofilm formation was verified using the microtitre plate assay. Antibiotic resistance was performed using the disc diffusion method. Staphylococcal ability to attach to host cells was assessed by flow cytometry. Cytotoxicity on epithelial cells was evaluated by LDH assay. RESULTS: The ica genes were detected in 26.9% of SE and in 42.3% of SA isolates. Only 15.3% of isolates (SE) were positive for both the icaAD and the ica operon. Phenotypically, 19.2% of SE isolates were strong biofilm producers, among which three were both icaAD- and ica operon-positive. About 26.9% of SA isolates were strong biofilm producers. Methicillin resistance (MR) was detected in 34.6% of SE and 26.9% of SA isolates. About 75% of MR isolates were multidrug resistant. SA isolates adhered to host cells more extensively than SE. SA isolates released higher level of LDH than SE. CONCLUSIONS: Adherence abilities were commonly observed in staphylococci associated with conjunctivitis. However, low prevalence of isolates positive for a complete and functional ica locus and low prevalence of strong biofilm producers was detected. SA adhered to a greater extent to eukaryotic cells than SE and were more cytotoxic.

The first outbreak of methicillin-resistant Staphylococcus aureus in dairy cattle in Poland with evidence of on-farm and intrahousehold transmission.

Staphylococcus aureus is a widely recognized pathogen responsible for many serious diseases in both humans and animals. It is also one of the major causative agents of bovine mastitis. Methicillin-resistant S. aureus (MRSA), although relatively rare in this pathology, has been increasingly reported in livestock animals, mainly in pigs, but also cattle, sheep, and poultry. The recent emergence of livestock-associated (LA-)MRSA is cause for an immediate public health concern due to the risk of zoonotic transmission to humans, and is of particular concern for people who work in animal husbandry or have prolonged contact with livestock animals. This study reports on the first LA-MRSA outbreak in dairy cattle and the first probable case of MRSA transmission between humans and cows in Poland. A single dairy farm located in Eastern Poland was monitored on a regular basis for the occurrence of mastitis. Over a 1-yr study period, 717 quarter-milk samples from 583 cows were collected and examined microbiologically. A total of 5 MRSA isolates from as many cows with subclinical mastitis were cultured. They all belonged to the same outbreak, given a 2-mo time window in which they were identified. During the outbreak, 24 oral and nasal swabs were voluntarily taken from 6 people: a milker, a veterinarian, and 4 members of the veterinarian's family. Eight swabs from a milker, veterinarian, and 2 family members yielded positive MRSA cultures. All MRSA isolates were genotyped with a combination of multiple-locus variable number tandem repeat analysis, multilocus sequence typing, and staphylococcal protein A gene (spa) typing. Eleven bovine (n = 5; 5 cases) and human (n = 6; 4 cases) isolates showed an identical drug-susceptibility profile and were indistinguishable upon multiple-locus variable number tandem repeat analysis (pattern A), multilocus sequence typing (ST398) and spa (t034) typing. The results of this study provide the evidence of transmission of MRSA between humans and cows, and between humans in the family setting. This work, despite being a preliminary investigation, underscores the risk of intra- and interspecies transmission of LA-MRSA and urges enhancement of the existing biosecurity measures aimed at preventing MRSA (and other milk pathogens) spread at both the farm- and household levels.

Tetracycline resistance in Moraxella catarrhalis clinical strains isolated in Poland.

Moraxella catarrhalis is considered an important, exclusively human respiratory tract pathogen which, along with Streptococcus pneumoniae and Haemophilus influenzae, is classified as one of the most frequent bacterial etiological factors causing upper respiratory tract infections. In this manuscript, we report the existence of five tetracycline-resistant M. catarrhalis strains with confirmed presence of tetracycline resistance tetB gene. The strains were isolated from children under the age of three with signs of upper respiratory tract infections. Our research also investigated the occurrence of virulence genes in these strains and involved the analysis of drug resistance to five antibiotic groups. It is the first description of clinical strains with confirmed presence of drug resistance tetB genes isolated in Europe.

The role of the PI3K/mTOR signaling pathway in Staphylococcus epidermidis small colony variants intracellular survival.

The objective of this study was to analyze how Staphylococcus epidermidis SCV and WT strains manipulate the PI3K/Akt/mTOR signaling pathway. Six S. epidermidis strains with normal phenotype (WT) and six S. epidermidis strains with SCV phenotype were isolated in parallel from six patients with the prosthetic hip joint infections. THP-1 activated cells were incubated with or without PI3K inhibitor-wortmannin or with mTOR inhibitor-rapamycin. Next, macrophages were exposed to S. epidermidis WT and SCV strains. After 4 h incubation, bacterial survival inside macrophages as well as PI3K-mTOR activation was analyzed. SCV strains of S. epidermidis increased the level of Akt phosphorylation, compared to uninfected macrophages and to their parental WT forms. Wild type variants of S. epidermidis phosphorylated Akt at similar or lower levels as control uninfected cells. Next, the induction of mTOR target, phosphorylated ribosomal protein S6, was measured in bacteria-infected macrophages. The level of phosphorylation was significantly reduced when the cells were exposed to WT strains of S. epidermidis. In contrast, the SCV strains activated S6 protein mostly at a level comparable to the control cells. Rapamycin inhibited mTOR activation as the number of p-S6 positive cells decreased in the tested cases. To conclude, the SCV strains activate the PI3K-Akt signaling pathway in opposite to WT strains. This fact however did not influence the increase in the number of live SCV bacteria as compared to the WT strains. Knowing that the PI3K-Akt pathway is involved in proinflammatory cytokines suppression, SCVs seem to use this pathway to reduce the inflammatory response during the infection.

A Novel Approach to Study the Effect of Ciprofloxacin on Biofilms of Corynebacterium spp. Using Confocal Laser Scanning Microscopy.

Non-diphtherial corynebacteria are Gram-positive rods that cause opportunistic infections, what is supported by their ability to produce biofilm on artificial surfaces. In this study, the characteristic of the biofilm produced on vascular and urological catheters was determined using a confocal microscopy for the most frequently involved in infections diphtheroid species. They were represented by the reference strains of Corynebacterium striatum ATCC 6940 and C. amycolatum ATCC 700207. The effect of ciprofloxacin on the biofilm produced by the antibiotic-susceptible C. striatum strain was evaluated using three concentrations of the antimicrobial agent (2 ×, 4 ×, and 6 × the MIC - the Minimum Inhibitory Concentration). The basis for the interpretation of results was the statistical analysis of maximum points readings from the surface comprising a total of 245 areas of the biofilm image under the confocal microscope. It was observed that ciprofloxacin at a concentration equal to 4 × MIC paradoxically caused an enlargement of areas with live bacteria within the biofilm. Biofilm destruction required the application of ciprofloxacin at a concentration higher than 6 × MIC. This suggests that the use of relatively low doses of antimicrobial agents may increase the number of live bacteria within the biofilm, and further facilitate their detachment from the biofilm's structure thus leading to the spread of bacteria into the bloodstream or to the neighboring tissues. The method of biofilm analysis presented here provides the original and novel approach to the investigation of the diphtheroid biofilms and their interaction with antimicrobial agents.Non-diphtherial corynebacteria are Gram-positive rods that cause opportunistic infections, what is supported by their ability to produce biofilm on artificial surfaces. In this study, the characteristic of the biofilm produced on vascular and urological catheters was determined using a confocal microscopy for the most frequently involved in infections diphtheroid species. They were represented by the reference strains of Corynebacterium striatum ATCC 6940 and C. amycolatum ATCC 700207. The effect of ciprofloxacin on the biofilm produced by the antibiotic-susceptible C. striatum strain was evaluated using three concentrations of the antimicrobial agent (2 ×, 4 ×, and 6 × the MIC ­ the Minimum Inhibitory Concentration). The basis for the interpretation of results was the statistical analysis of maximum points readings from the surface comprising a total of 245 areas of the biofilm image under the confocal microscope. It was observed that ciprofloxacin at a concentration equal to 4 × MIC paradoxically caused an enlargement of areas with live bacteria within the biofilm. Biofilm destruction required the application of ciprofloxacin at a concentration higher than 6 × MIC. This suggests that the use of relatively low doses of antimicrobial agents may increase the number of live bacteria within the biofilm, and further facilitate their detachment from the biofilm's structure thus leading to the spread of bacteria into the bloodstream or to the neighboring tissues. The method of biofilm analysis presented here provides the original and novel approach to the investigation of the diphtheroid biofilms and their interaction with antimicrobial agents.

Synergistic Effect of Ceftazidime-Avibactam with Aztreonam on Carbapenemase-Positive Klebsiella pneumoniae MBL+, NDM.

Background: The difficulties in attaining effective antibiotic therapy arising from the multidrug resistance of Gram-negative bacilli compel the exploration of new possibilities for synergistic interactions among existing antibiotics. Research Design and Methods: An analysis was conducted to assess the efficacy of two antibiotic therapy regimens in the treatment of infections caused by Klebsiella pneumoniae strains producing carbapenemases (MBL). Two patient groups were considered: Group A - individuals in whom the treatment of infection involved the application of ceftazidime-avibactam in combination with aztreonam. Group B comprised patients subjected to an alternative antibiotic therapy regimen. Results: In the group subjected to the treatment regimen involving ceftazidime-avibactam and aztreonam, as compared to alternative antibiotic combinations, a statistically lower mortality rate during the course of treatment and a faster clinical response to the administered therapy were evident. Conclusion: The results obtained may be applicable to routine in vitro assays performed and serve as valuable guidance for the potential utilization of the positive effect of antibiotic therapy through the synergy between ceftazidime-avibactam and aztreonam. The selection of antibiotics employed in the therapy of invasive infections caused by K. pneumoniae influences the ultimate treatment outcome.

A rare case of invasive Enterococcus cecorum infection and related diagnostic difficulties.

Key Clinical Message: This report presents a rare case of invasive infection caused by Enterococcus cecorum. There are no specific guidelines regarding antibiotic therapy for this infection. Based on this case, it can be concluded that linezolid demonstrates in vivo activity against Enterococcus cecorum and can be successfully used in therapy. Abstract: Enterococcus cecorum is an extremely rare pathogen in humans. Since 1984, when the microorganism was first described, only a dozen cases of invasive infections in humans have been reported in the literature. The diagnostic pathway may involve difficulties in correctly identifying this microorganism. Based on the case described, it can be thought that Enterococcus cecorum is a more challenging bacterium than the much more common Enterococcus faecium or Enterococcus faecalis. The described case underscores the importance of medical vigilance in clinical practice. It seems that due to increasingly advanced techniques in molecular biology, we will more frequently detect pathogens that were previously encountered only sporadically. Since not every center has access to modern and advanced microbiological diagnostic methods, publications that practically combine classical microbiological diagnostic methods with those less accessible but more modern are exceptionally valuable. In the case described, it is also worth noting that classical methods still play a significant and crucial role in conducting microbiological diagnostics. In the era of rapid diagnostic tool development, it is important to emphasize the necessity of combining different methods rather than replacing one with another.

Antibiotic resistance and detection of the most common mechanism of resistance (MLSB) of opportunistic Corynebacterium.

BACKGROUND: Determination of antibiotic resistance of opportunistic Corynebacterium colonizing the nose that cause infections and evaluation of the applicability of a simple method for detecting the most common constitutive-type resistance to macrolides, lincosamides and streptogramin B (MLSB). METHODS: 70 isolates colonizing the nose and 70 clinical isolates of various infection sites were used and identified using APICoryne and 16S rRNA. Minimal inhibitory concentrations (MICs) were determined (Etest) for 12 antibiotics. MLSB was defined based on MIC, a simple method using two disks (erythromycin/clindamycin) and detection of the gene erm X (PCR). RESULTS: There was a high percentage--in both groups at the same level--of strains with MLSB (88.5% colonizing the nose and 87.1% causing infections). Detection with the phenotypic method MLSB was confirmed genetically (erm X) in all cases. In both groups, a high percentage of resistance was found to trimethoprim/sulfamethoxazole (in both groups 71.4%), chloramphenicol (nose 44.2%/infections 37.1%), tetracycline (28 and 45.7%) and ß-lactam antibiotics (18.5 and up to 32.8%). CONCLUSION: Differences in antibiotic resistance were found between strains colonizing the respiratory tract and various infections. Isolates from infections more frequently exhibited multidrug resistance. The possibility of using a simple method was confirmed for MLSB detection, which can be applied to determine drug resistance in routine microbiological diagnostics of infections caused by opportunistic Corynebacterium.

[Problems with identification of beta-hemolytic streptococcus resistant to bacitracin isolated from patients with pharyngitis]. / Problemy z identyfikacja paciorkowców beta-hemolizujacych opornych na bacytracyne izolowanych od chorych z zapaleniem gardla.

INTRODUCTION: The genus Streptococcus comprises a number of species characterized by a differential pathogenic potential. These bacteria can be considered as members of microbial physiological flora but they can also cause mild infections or severe, life threatening conditions. The majority of infections of streptococcal etiology are caused by beta-hemolysing species. The predominant causative agent of bacterial pharyngitis is Streptococcus pyogenes. This species usually doesn't give rise to any identification difficulties due to the introduction the well determined diagnostic schemes. Problems concerning laboratory identification can be, however, associated with other species of beta-hemolysing streptococci isolated from patients with pharyngitis. These streptococci can demonstrate features similar to those of S. pyogenes and share the group antygen A, such as some strains of Streptococcus anginosus and Streptococcus dysgalactiae subsp. equisimilis. The determination of sensitivity to bacitracin, which is a feature typical of S. pyogenes, is the basic test useful for its preliminary identification. Nevertheless, the identification of some strains by this test can give rise to incompatibility. The aim of the study was characterisation of beta-hemolysing streptococci resistant to bacitracin isolated from patients with pharyngitis. The examined bacterial strains caused identification problems by the use of routine diagnostic methods. METHODS: The material included 14 streptococcal strains resistant to bacitracin which were isolated from adult patients suffering from pharyngitis. The bacteria were cultured on media dedicated for the species. The following routine diagnostic tests were used for the bacterial identification: sensitivity to bacitracin (0.04 U/disc), CAMP test, determination of the group antigens A, B, C, D, F and G (Slidex Strepto-Kit), and determination of biochemical features by the API 20 STREP test (bioMèrieux). The sensitivity of streptococcal isolates to antibiotics (penicillin, clindamycin, erythromycin, tetracycline, vancomycin, ofloxacin) and trimethoprim/sulfamethoxazole, was determined by the disc diffusion method on the Mueller-Hinton agar with 5% sheep blood (the inoculum-0.5 McFarland). RESULTS: Among the 14 isolates resistant to bacitracin, 6 isolates of S. pyogenes, 6 isolates of S. constellatus, and 2 isolates of S. dysgalactiae subsp. equisimilis were identified. All isolates were sensitive to penicillin and vancomycin. One isolate ofS. pyogenes demonstrated constitutive MLSB resistance mechanism. Seven isolates were resistant to tetracycline: S. dysgalactiae subsp. equisimilis (3 isolates), S. constellatus (3), and S. pyogenes (1). The number of isolates resistant to trimethoprim/sulfamethoxazole was as follows: S. pyogenes (6) and S. dysgalactiae subsp. equisimilis (1), whereas four isolates were resistant to ofloxacin. CONCLUSIONS: The bacitracin test cannot be used as the only test for the laboratory identification of S. pyogenes even if it is combined with the determination of the Lancefield group antigen due to the existence of bacitracin resistant S. pyogenes strains. Therefore, it is necessary to perform biochemical commercial tests in addition to routine phenotypic tests. Isolation of beta-hemolysing streptococci other than S. pyogenes from patients with pharyngitis confirms their role in the etiology of this infection. Taken into account the significance of determination of sensitivity to bacitracin for the preliminary identification of S. pyogenes, it is necessary to standardize this test, which will make obtaining of the comparability of results possible.

Dysregulation in the Expression of Platelet Surface Receptors in Acute Coronary Syndrome Patients-Emphasis on P2Y12.

The pathological conditions caused by blood platelet activation constitute a fundamental core in the pathogenesis of Acute Coronary Syndrome (ACS). The hyperactivity of platelets in ACS is well-documented, but there is still little research into the molecular basis of phenotypic changes in platelet functionality. To expand the knowledge of this phenomenon, we analyzed the disturbances in the expression of several key platelet receptors and the aspect of regulating potential abnormalities. Platelet surface receptors are responsible for maintaining the hemostatic balance, platelet interaction with immune cells, and support of the coagulation cascade leading to occlusion of the vessel lumen. Due to their prominent role, platelet receptors constitute a major target in pharmacological treatment. Our work aimed to identify the molecular alteration of platelet surface receptors, which showed augmented mRNA expression of P2Y12, GP1BB, ITGA2B, and ITGB3 and increased protein concentrations of P2Y12 and GP IIb/IIIa in ACS. The upregulation of the P2Y12 level was also confirmed by confocal and cytometric visualization. Furthermore, we evaluated the expression of two microRNAs: miR-223-3p and miR-126-3p, which were suggested to regulate platelet P2Y12 expression. Results of our study present new insight into the molecular background of ACS.

Evaluation of Selected Parameters of the Specific Immune Response against Pseudomonas aeruginosa Strains.

The immune response to Pseudomonas aeruginosa strains could be influenced by differences in antibiotic resistance and virulence. At the present time, it is unclear which type of immune responses enables uncontrolled invasion of opportunistic pathogens. The conditional pathogenicity of Pseudomonas aeruginosa served as an inspiration to begin a study on this bacterium. The aim of this study was to gain insight into selected parameters describing immune responses with regards to the adaptable agents of this pathogen. For the analysis of the specific immune response, the potential of Pseudomonas aeruginosa to stimulate lymphocytes, including Th17 lymphocytes, dendritic cells and other components of the adaptive immune response, was examined. The highest percentage of CD83+CD1a-HLA-DR++ cells was found after stimulation with lysates of strains isolated from the patients with severe systemic infection. We found statistically significant differences in percentages of HLA-DR+ PBMCs and MFI of HLA-DR between groups of Pseudomonas aeruginosa strains isolated from the patients with different clinical courses of infection. Our results suggest that the clinical course and outcomes of Pseudomonas aeruginosa infections are not associated with impairment of the specific immune response.

[Wound infections due to opportunistic corynebacterium species]. / Zakazenia ran wywolane przez maczugowce reprezentujace gatunki bakterii uwazanych za oportunistyczne.

Wound infections are often due to endogenous bacterial flora which penetrates into a site of injury. The establishment of the etiologic agent can be problematic, especially when opportunistic bacteria are present, suggesting contamination of clinical material. Among bacteria that can cause such diagnostic problems are opportunistic Corynebacterium spp. and coryneforms colonizing skin. The aim of the study was to analyze the 24 clinical samples collected from wounds of different location, with Gram positive rods isolated in numbers suggesting the cause of infection. Bacterial identification was performed by API Coryne and additional biochemical tests (API ZYM, API NE). It was detected that the commonest species isolated were: C. amycolatum (29.2%), C. striatum (16.7%), C. group G (16.7%) and Brevibacterium spp., C. jeikeium, C. urealyticum, C. group F1. The drug susceptibility testing was performed by E-test method. Among isolated strains, 83.3% were simultaneously resistant to erythromycin and clindamycin. In 75% cases resistance to co-trimoxazole was noted, in 71.7% resistance to chloramphenicol and in 16.7% resistance to beta-lactams were detected. In presented study the high percentage of strains resistant to macrolids and linkosamids (MLSB) was noted. All strains were susceptible to vancomycin and teicoplanin.

Draft whole-genome sequence of Brevibacterium casei strain isolated from a bloodstream infection.

Despite its low virulence potential and a commensal lifestyle as a member of the human skin microbiota, Brevibacterium casei has been increasingly reported as an opportunistic pathogen, especially in immunocompromised patients. Here, we present the draft genome sequence of the S51 strain isolated from a bloodstream infection. To the best of the authors' knowledge, this is the first report of the draft genome sequence of the B. casei strain isolated from the clinical infection. The strain was identified using phenotypic and molecular methods and subsequently sequenced using the next-generation sequencing. The draft whole genome was assembled de novo, automatically annotated by Rapid Annotations using Subsystems Technology (RAST) server and scrutinized to predict the presence of virulence, resistance, and stress response proteins. The genome size of the S51 strain was 3,743,532 bp and an average G+C content was 68.3%. The predicted genes included 48 genes involved in resistance to antibiotics (including vancomycin, fluoroquinolones, and beta-lactams) and toxic compounds (heavy metals), 16 genes involved in invasion and intracellular resistance (Mycobacterium virulence operons), and 94 genes involved in stress response (osmotic, oxidative stress, cold and heat shock). ResFinder has indicated the presence of a beta-lactamase, and a phenotypic analysis showed resistance to penicillin. This whole-genome NGS project for the S51strain has been deposited at EMBL/GenBank under the accession no. QNGF00000000.

Infiltrating Treg and Th17 Cells of the Prostate Hypertrophy Gland Associated with Propionibacterium Acnes Infection.

PURPOSE: The aim of the study was to evaluate prostate cell infiltration by CD4(+)IL-17(+) and Treg cells in BPH and PCa patients depending on P. acnes infection in the prostate gland. PATIENTS AND METHODS: Prostate fragments were collected from 54 patients with PCa and 34 patients with BPH. Rapid ID 32 was used to identify the bacteria. Cells were analyzed by flow cytometry BD FACSCanto II. Statistical analysis was performed using Statistica 7 software (TIBCO Software Inc, USA). RESULTS: P. acnes was detected in 35% of patients with PCa and 41% of individuals with BPH. The infiltration of CD4(+)IL-17(+) and Treg cells was statistically significantly higher (P = 0.001) in patients with BPH and positive for P. acnes. A statistically considerably higher (P = 0.001) infiltration of Treg cells in treated for PCa with P.acnes infection was also demonstrated. CONCLUSION: Prostatitis caused by P. acnes may contribute to the development of BPH and PCa.

Cytokine Levels in the In Vitro Response of T Cells to Planktonic and Biofilm Corynebacterium amycolatum.

Unravelling of the interplay between the immune system and non-diphtheria corynebacteria would contribute to understanding their increasing role as medically important microorganisms. We aimed at the analysis of pro- (TNF, IL-1ß, IL-6, IL-8, and IL-12p70) and anti-inflammatory (IL-10) cytokines produced by Jurkat T cells in response to planktonic and biofilm Corynebacterium amycolatum. Two reference strains: C. amycolatum ATCC 700207 (R-CA), Staphylococcus aureus ATCC 25923 (R-SA), and ten clinical strains of C. amycolatum (C-CA) were used in the study. Jurkat T cells were stimulated in vitro by the planktonic-conditioned medium (PCM) and biofilm-conditioned medium (BCM) derived from the relevant cultures of the strains tested. The cytokine concentrations were determined in the cell culture supernatants using the flow cytometry. The levels of the cytokines analyzed were lower after stimulation with the BCM when compared to the PCM derived from the cultures of C-CA; statistical significance (p < 0.05) was observed for IL-1ß, IL-12 p70, and IL-10. Similarly, planktonic R-CA and R-SA stimulated a higher cytokine production than their biofilm counterparts. The highest levels of pro-inflammatory IL-8, IL-1ß, and IL-12p70 were observed after stimulation with planktonic R-SA whereas the strongest stimulation of anti-inflammatory IL-10 was noted for the BCM derived from the mixed culture of both reference species. Our results are indicative of weaker immunostimulatory properties of the biofilm C. amycolatum compared to its planktonic form. It may play a role in the persistence of biofilm-related infections. The extent of the cytokine response can be dependent on the inherent virulence of the infecting microorganism.Unravelling of the interplay between the immune system and non-diphtheria corynebacteria would contribute to understanding their increasing role as medically important microorganisms. We aimed at the analysis of pro- (TNF, IL-1ß, IL-6, IL-8, and IL-12p70) and anti-inflammatory (IL-10) cytokines produced by Jurkat T cells in response to planktonic and biofilm Corynebacterium amycolatum. Two reference strains: C. amycolatum ATCC 700207 (R-CA), Staphylococcus aureus ATCC 25923 (R-SA), and ten clinical strains of C. amycolatum (C-CA) were used in the study. Jurkat T cells were stimulated in vitro by the planktonic-conditioned medium (PCM) and biofilm-conditioned medium (BCM) derived from the relevant cultures of the strains tested. The cytokine concentrations were determined in the cell culture supernatants using the flow cytometry. The levels of the cytokines analyzed were lower after stimulation with the BCM when compared to the PCM derived from the cultures of C-CA; statistical significance (p < 0.05) was observed for IL-1ß, IL-12 p70, and IL-10. Similarly, planktonic R-CA and R-SA stimulated a higher cytokine production than their biofilm counterparts. The highest levels of pro-inflammatory IL-8, IL-1ß, and IL-12p70 were observed after stimulation with planktonic R-SA whereas the strongest stimulation of anti-inflammatory IL-10 was noted for the BCM derived from the mixed culture of both reference species. Our results are indicative of weaker immunostimulatory properties of the biofilm C. amycolatum compared to its planktonic form. It may play a role in the persistence of biofilm-related infections. The extent of the cytokine response can be dependent on the inherent virulence of the infecting microorganism.

Antifungal effects of a 1,3,4-thiadiazole derivative determined by cytochemical and vibrational spectroscopic studies.

Compounds belonging to the group of 5-substituted 4-(1,3,4-thiadiazol-2-yl) benzene-1,3-diols exhibit a broad spectrum of biological activity, including antibacterial, antifungal, and anticancer properties. The mechanism of the antifungal activity of compounds from this group has not been described to date. Among the large group of 5-substituted 4-(1,3,4-thiadiazol-2-yl) benzene-1,3-diol derivatives, the compound 4-(5-methyl-1,3,4-thiadiazole-2-yl) benzene-1,3-diol, abbreviated as C1, was revealed to be one of the most active agents against pathogenic fungi, simultaneously with the lowest toxicity to human cells. The C1 compound is a potent antifungal agent against different Candida species, including isolates resistant to azoles, and molds, with MIC100 values ranging from 8 to 96 µg/ml. The antifungal activity of the C1 compound involves disruption of the cell wall biogenesis, as evidenced by the inability of cells treated with C1 to maintain their characteristic cell shape, increase in size, form giant cells and flocculate. C1-treated cells were also unable to withstand internal turgor pressure causing protoplast material to leak out, exhibited reduced osmotic resistance and formed buds that were not covered with chitin. Disturbances in the chitin septum in the neck region of budding cells was observed, as well as an uneven distribution of chitin and ß(1→3) glucan, and increased sensitivity to substances interacting with wall polymerization. The ATR-FTIR spectral shifts in cell walls extracted from C. albicans cells treated with the C1 compound suggested weakened interactions between the molecules of ß(1→3) glucans and ß(1→6) glucans, which may be the cause of impaired cell wall integrity. Significant spectral changes in the C1-treated cells were also observed in bands characteristic for chitin. The C1 compound did not affect the ergosterol content in Candida cells. Given the low cytotoxicity of the C1 compound to normal human dermal fibroblasts (NHDF), it is possible to use this compound as a therapeutic agent in the treatment of surface and gastrointestinal tract mycoses.