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Ma&lstrok;gorzata &Lstrok;ysek-G&lstrok;adysi&nacute;ska,1 Magdalena Starz,2 Agnieszka Borowiec-S&eogon;k,2 Iwona Sufin,2 Anna Wieczorek,1 Magdalena Chrapek,3 Dorota Zar&eogon;bska-Michaluk,4 Patrycja Sufin,2 Stanis&lstrok;aw G&lstrok;uszek,4 Wioletta Adamus-Bia&lstrok;ek4 1Institute of Biology, Jan Kochanowski University, Kielce, Poland; 2Meduniv Sp. z o.o, Kielce, Poland; 3Department of Mathematics, Jan Kochanowski University, Kielce, Poland; 4Institute of Medical Sciences, Jan Kochanowski University, Kielce, PolandCorrespondence: Wioletta Adamus-Bia&lstrok;ek, Jan Kochanowski University, Institute of Medical Sciences, IX Wieków Kielc 19a, Kielce, 25-516, Poland, Tel +48 788 860 604, Email wioletta.adamus-bialek@ujk.edu.plPurpose: The COVID-19 pandemic has been going on for almost three years, and so far, many preventive and therapeutic strategies have been developed. The issue of subsequent booster vaccinations is currently being discussed. We aimed to analyze how the third dose of vaccination against COVID-19 correlates with the dynamics of IgG anti-SARS-CoV-2 spike protein antibody levels in a group of healthy people.Patients and Methods: The prospective study included 93 participants before and after a second booster of COVID-19 vaccination, from whom 4 blood samples were collected at intervals. The levels of IgG anti-SARS-CoV-2 in serum were identified using the chemiluminescent immunoassay specific for the receptor-binding domain (RBD) of the S1 protein. The analysis of the results was performed using appropriate statistical methods, considering p < 0.05 as a statistically significant value.Results: The IgG levels were significantly higher and less diverse after the same follow-up time from the second booster vaccination compared to the first booster. The antibody levels were positively correlated with female, healthcare workers, the elderly and participants with a negative COVID-19 history. Furthermore, the increase in IgG antibodies after the second booster vaccination correlated inversely with the baseline level of antibodies before the vaccination. The latest results showed that antibody levels dropped 1.5-fold after approx. 10 months from the second booster vaccination but still remained at a protective level.Conclusion: Booster vaccinations seem to better stimulate immune memory, and in the case of borderline IgG level induces the greatest increase in antibodies. It is worth considering the individual parameters of patients and measuring antibodies before vaccination.Keywords: pandemic, virus, infection, immune response

The aim of the study was a profound insight into the antibiotic resistance development in uropathogenic Escherichia coli strains with the use of some mathematical and statistical methods. During the previous study some antibiotics (amoxicillin, ciprofloxacin, gentamycin, and tobramycin) were applied to induce the resistance of E. coli strains, which led to the generation of ∼120 derivative strains with changed antibiotic susceptibility profiles. In this work, quantitative analysis was performed based on the strains defined as values of vectors of susceptibility for all the antibiotics' use. The Pearson correlation coefficient was used to define the correlation dissimilarity (distance) of the strains, which was further applied to hierarchical clustering. Analogously, the antibiotics were presented as vectors of susceptibility values of all the investigated strains. Correlation and cluster analysis were performed for antibiotics. The hclust method from the R system with the Ward method was used as a class agglomeration method. Mathematical analysis revealed two types of statistically relevant interactions-between antibiotics and derivative strains, as well as between the effect of individual antibiotics on the bacterial strains. These observed correlations can play a potential role for modeling uropathogenic E. coli (UPEC) resistant changes, based on the particular antibiotic used to initiate resistance development, or a model helping to predict drug resistance interactions in various UPEC strains. The obtained results can lead to development of much more sophisticated mathematical models, which, in turn, can be a potentially useful tool as a drug resistance trend predictor, both for clinicians and epidemiologists.

Background: Due to the decrease in the percentage of perinatal mortality, which is one of the Millennium Development Goals, the number of children with a central coordination disorder (CCD) has increased, present in up to 40% of premature babies. Neurodevelopmental disorders detected in the diagnostic process require early interventions that will eliminate or overcome existing dysfunctions. These treatments often cause discomfort in the infant, which induces insecurity and activation of basic defense mechanisms. The aim of the work is to assess changes in cortisol concentration in infants treated with the Vojta method. Methods and findings: The study included 35 children with CCD aged between three and nine months. The participants had no comorbidities that could have affected the obtained results. The activities were planned to occur in three stages: 1. Collection of a saliva sample directly before the physiotherapy appointment. 2. Collection of saliva immediately after rehabilitation. 3. Collection of saliva 20 min after the end of rehabilitation. The physiotherapeutic intervention included the assessment of seven reactions of the body position in space according to Vojta and the conduct of a therapeutic session consisting of the first phase of rotation and creeping reflex according to Vojta. The concentration of free cortisol in saliva was assessed with LC-MS/MS. In the first measurement, none of the children presented an excess of the normative concentration of cortisol. The cortisol measurement performed directly after rehabilitation showed above-normative values in three children. In the third measurement, all of the children presented a decreased concentration of free cortisol. The analysis (paired two-tailed t-test, p < 0.05) showed statistically significant differences between particular stages of the measurements. The analysis of the scores obtained in the second measurement showed the concentration of scores in the area of “normal” at a level of 0.83 (normal concentration) and the area “above normal” at the level of 0.005 (very weak concentration). Based on the analysis of significance of the obtained scores, it was found that the result was not accidental, and the Vojta method used in the treatment of children with CCD was suitable. Conclusions: Here, for the first time, we presented how Vojta therapy was correlated with cortisol levels among children with a central coordination disorder.

Objectives It has previously been reported in a European case-control study with patients from Germany and France that CEL-HYB1, a hybrid allele of the carboxyl ester lipase (CEL) gene and its pseudogene CELP, increases susceptibility to chronic pancreatitis (CP). Here, we aimed to replicate this finding in Polish pediatric patients with CP. Method The distribution of the CEL-HYB1 allele in a CP pediatric cohort (n = 147, median age at CP onset 7.6 years) with no history of alcohol/smoking abuse was compared with ethnically matched healthy controls (n = 500, median age 46 years). Screening was performed using long-range PCR followed by agarose gel-electrophoresis. Results We observed no significant difference in the carrier frequency of the CEL-HYB1 allele between CP patients (7/147, 4.8%) and controls (12/500, 2.4%; P = 0.16). Conclusions This study found no statistically significant association between CEL-HYB1 and chronic pancreatitis in a cohort of Polish pediatric CP patients.

The associations between serum vaspin levels and metabolic or coronary artery disease (CAD) and polycystic ovary syndrome (PCOS) is under the scope of current researchers. Therefore, this adipokine can be considered as a biomarker of metabolic syndrome (MetS). The aim of the study was to analyze the associations between the vaspin rs2236242 polymorphism and physical activity in relation to MetS and its components. The analysis involved the genetic material and clinical data of 108 individuals with MetS and 110 controls. Vaspin rs2236242 polymorphism was detected using the tetra-primer amplification-refractory mutation system polymerase chain reaction (T-ARMS PCR) method. The TA genotype of vaspin rs2236242 was associated with a greater risk of MetS and its components compared with the TT genotype. The analysis of interactions between genotype and walking time revealed that a walking time longer than 60 min./day significantly decreased the risk of MetS in the TA carriers (p = 0.007). The obtained results suggest that any unfavorable effect of the TA genotype of the vaspin rs2236242 polymorphism can be essentially reduced, or even reversed, in a case of individuals walking longer than 60 min. a day. The analysis of the interaction between vaspin rs2236242 polymorphism and walking showed that a walking time of longer than 1 hour a day significantly reduced the risk of MetS, elevated blood pressure and triglycerides concentration.

Proteus mirabilis is the third most common etiological factor of urinary tract infection. It produces urease, which contributes to the formation of a crystalline biofilm, considered to be one of the most important virulence factors of P. mirabilis strains, along with their ability to swarm on a solid surface. The aim of this study was to analyze the pathogenic properties of two selected groups of clinical P. mirabilis isolates, antimicrobial susceptible and multidrug resistant (MDR), collected from hospitals in different regions in Poland. The strains were examined based on virulence gene profiles, urease and hemolysin production, biofilm formation, and swarming properties. Additionally, the strains were characterized based on the Dienes test and antibiotic susceptibility patterns. It turned out that the MDR strains exhibited kinship more often than the susceptible ones. The strains which were able to form a stronger biofilm had broader antimicrobial resistance profiles. It was also found that the strongest swarming motility correlated with susceptibility to most antibiotics. The correlations described in this work encourage further investigation of the mechanisms of pathogenicity of P. mirabilis.

Fenugreek seeds are widely used in Asia and other places of the world for their nutritive and medicinal properties. In Asia, fenugreek seeds are also recommended for geriatric populations. Here, we evaluated for the first time the effect of fenugreek seed feed supplementation on the liver antioxidant defense systems in aging mice. The study was conducted on 12-months aged mice which were given fenugreek seed dietary supplement. We evaluated the activities of various antioxidant defense enzymes such as superoxide dismutase (SOD), glutathione reductase (GR), and glutathione peroxidase (GPx), and also estimated the phenolics and free radical scavenging properties in mice liver upon fenugreek supplementation. The estimation of SOD, GPx, and GR activities in aged mice liver revealed a significant (p < 0.01) difference among all the liver enzymes. Overall, this study reveals that fenugreek seed dietary supplementation has a positive effect on the activities of the hepatic antioxidant defense enzymes in the aged mice.

Antibiotic therapy and its consequences in bacterial and human aspects are widely investigated. Despite this, the emergence of new multidrug resistant bacteria is still a current problem. The scope of our work included the observation of changes among uropathogenic Escherichia coli strains after the treatment with a subinhibitory concentration of different antibiotics. The sensitive strains with or without virulence factors were incubated with amoxicillin, ciprofloxacin, gentamycin, or tobramycin. After each passage, the E. coli derivatives were compared to their wild types based on their susceptibility profiles, virulence genes, biofilm formations and the fingerprint profiles of PCR products amplified with using the (N)(6)(CGG)(4) primer. It turned out that antibiotics caused significant changes in the repertoire of bacterial virulence and biofilm formation, corresponding to acquired cross-resistance. The genomic changes among the studied bacteria were reflected in the changed profiles of the CGG-PCR products. In conclusion, the inappropriate application of antibiotics may cause a rapid rise of Multidrug Resistant (MDR) strains and give bacteria a chance to modulate their own pathogenicity. This phenomenon has been easily observed among uropathogenic E. coli strains and it is one of the main reasons for recurrent infections of the urinary tract.

The aim of the study was a profound insight into the antibiotic resistance development in uropathogenic

Fenugreek seeds are widely used in Asia and other places of the world for their nutritive and medicinal properties. In Asia, fenugreek seeds are also recommended for geriatric populations. Here, we evaluated for the first time the effect of fenugreek seed feed supplementation on the liver antioxidant defense systems in aging mice. The study was conducted on 12-months aged mice which were given fenugreek seed dietary supplement. We evaluated the activities of various antioxidant defense enzymes such as superoxide dismutase (SOD), glutathione reductase (GR), and glutathione peroxidase (GPx), and also estimated the phenolics and free radical scavenging properties in mice liver upon fenugreek supplementation. The estimation of SOD, GPx, and GR activities in aged mice liver revealed a significant (p &lt, 0.01) difference among all the liver enzymes. Overall, this study reveals that fenugreek seed dietary supplementation has a positive effect on the activities of the hepatic antioxidant defense enzymes in the aged mice.

The clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins are components of the adaptive immunity system, protecting against foreign DNA, which are present in many bacteria species. Recent years have brought extensive research on this system however, not all of its biological properties have been discovered so far. It was recently discovered that CRISPR-Cas can regulate the formation of biofilm and is closely associated with the DNA repair system in bacterial cells. It is also likely that some of the spacer sequences are complementary to short sequences in the bacterial genome, which may have an influence on regulation of bacterial genes, e.g. virulence factors. Besides, phages can synthesize anti-CRISPR genes, which could be of use in the future for the purpose of development of an alternative therapy against multi-drug resistant bacterial strains. Here we present an elementary characteristic of CRISPR-Cas system, including the structure and the brief mechanism of action, systematic classification and its importance for medicine and biotechnology issues. We would like to stress the huge potential of CRISPR-Cas by discussing the selected but varied aspects. 1. Introduction. 2. Structure, operation and differences. 3. Bacterial typing. 4. Correlation with bacterial pathogenicity. 5. Potential tool for medicine. 5.1. CRISPR-tool for genome editing. 5.2. Instances of CRISPR-tool strategies in medicine. 6. Phage response. 7. Conclusions

Reactive oxygen intermediates, such as hydrogen peroxide, are toxic molecules produced by immune cells in response to bacterial invasion into the host. Bacteria try to protect themselves against the immune system through specific properties such as biofilm formation. This phenomenon occurs also during urinary tract infections. Cellulose is an important factor of Escherichia coli biofilm and contributes to building a protective shield around bacterial cells upon the host immune response. In this study, we aimed to analyze the effect of hydrogen peroxide on the production of this biofilm component. To achieve this goal, 25 clinical E. coli strains isolated from patients with urinary tract infections were used. These bacterial strains were characterized based on their growth characteristics, their ability to form biofilm and their capacity to produce cellulose upon exposure to sub-lethal concentrations of hydrogen peroxide growth, and the biofilm formation of these strains was analyzed. Our results revealed that the analyzed uropathogenic E. coli strains slightly, but significantly, reduced growth and biofilm production upon hydrogen peroxide treatment. However, when separating these strains regarding their ability to produce cellulose, we found that general biofilm production was reduced but cellulose expression was induced upon peroxide treatment. This finding contributes to a better understanding of how bacterial biofilm formation is triggered and provides interesting insights into how uropathogenic E. coli protect themselves in an inhospitable environment.

Pseudomonas aeruginosa infection is problematic in patients with cystic fibrosis (CF). P. aeruginosa secretes a diversity of pigments, such as pyocyanin and pyoverdine. The aim of this study was to evaluate the effects of complexes of nickel(II) ([Ni(iaa)2(H2O)2]·H2O (iaa = imidazole-4-acetate anion), [Ni(1-allim)6](NO3)2 (1-allim = 1-allylimidazole) and NiCl2 on pyocyanin and pyoverdine production by 23 strains of P. aeruginosa isolated from cystic fibrosis under growth conditions specific for the CF respiratory system. The antibacterial effects and biophysical properties of the tested substances were measured by spectrofluorometric techniques, as well as by laser interferometry, confocal and atomic force microscopy. The cytotoxic properties of all compounds were measured by Annexin/IP assay against A549 cells. All tested compounds have no effect on pyocyanin production and decrease the pyoverdine secretion in about 40% of tested P. aeruginosa strains at non-cytotoxic range of concentrations. Imidazole-4-acetate anion and 1-allylimidazole have good diffusion properties in the mature P. aeruginosa PAO1 biofilm. In conclusion, the tested nickel(II) complexes do not have clinical implications in P. aeruginosa eradication in cystic fibrosis. The diffusion properties of 1-allylimidazole and imidazole-4-acetate and their lack of effect on A549 cells suggest that they might be considered for chemical synthesis with other transition metals.

Escherichia coli and Proteus mirabilis are important urinary tract pathogens. The constant increase in the antibiotic resistance of clinical bacterial strains has become an important clinical problem. The aim of this study was to compare the antibiotic resistance of 141 clinical (Sweden and Poland) and 42 laboratory (Czech Republic) P. mirabilis strains and 129 clinical (Poland) uropathogenic E. coli strains. The proportion of unique versus diverse patterns in Swedish clinical and laboratory P. mirabilis strain collections was comparable. Notably, a similar proportion of unique versus diverse patterns was observed in Polish clinical P. mirabilis and E. coli strain collections. Mathematical models of the antibiotic resistance of E. coli and P. mirabilis strains based on Kohonen networks and association analysis are presented. In contrast to the three clinical strain collections, which revealed complex associations with the antibiotics tested, laboratory P. mirabilis strains provided simple antibiotic association diagrams. The monitoring of antibiotic resistance patterns of clinical E. coli and P. mirabilis strains plays an important role in the treatment procedures for urinary tract infections and is important in the context of the spreading drug resistance in uropathogenic strain populations. The adaptability and flexibility of the genomes of E. coli and P. mirabilis strains are discussed.

Diseases that are caused by non-tuberculous mycobacteria (NTM) continue to pose difficult clinical problems, and the epidemiological aspect of NTM-caused diseases is of great importance. In the case of Mycobacterium gordonae there is no adequate genotyping scheme. Here we present a potential rapid and reproducible genetic assay that uses trinucleotide repeat sequence-based PCR (TRS-PCR) for genotyping M. gordonae. The proposed method constitutes a useful single-primer PCR screen for genotyping this species. Among 10 TRS-containing primers, after applying (CAC)₄-based PCR to 36 strains of M. gordonae, we found a discriminatory index of 0.975. The accuracy of this analysis was supported by a reasonable reproducibility of 92%. These results were compared with the Enterobacterial Repetitive Intergenic Consensus Sequences (ERIC)-PCR typing scheme which had lower discriminatory index of 0.93 and its reproducibility was only 86.3%.

The ability to form different types of biofilm enables bacteria to survive in a harsh or toxic environment. Different structures of biofilms are related to different surfaces and environment of bacterial growth. The aim of this study was analysis of the biofilm formation of 115 clinical uropathogenic Escherichia coli strains under different growth conditions: surface for biofilm formation, medium composition and time of incubation. The biofilm formation after 24 h, 48 h, 72 h and 96 h was determined spectrophotometrically (A531) after crystal violet staining and it was correlated with bacterial growth (A600). The live and dead cells in biofilm structures was also observed on the glass surface by an epi-fluorescence microscope. Additionally, the presence of rpoS, sdiA and rscA genes was analyzed. The statistical significance was estimated by paired T-test. The observed biofilms were different for each particular strain. The biofilm formation was the highest in the rich medium (LB) after 24 h and its level hasn't changed in time. When biofilm level was compared to bacterial growth (relative biofilm) - it was higher in a minimal medium in comparison to enriched medium. These results suggest that most of the bacterial cells prefer to live in a biofilm community under the difficult environmental conditions. Moreover, biofilm formation on polyurethane surface did not correlate with biofilm formation on glass. It suggests that mechanisms of biofilm formation can be correlated with other bacterial properties. This phenomenon may explain different types of biofilm formation among one species and even one pathotype - uropathogenic Escherichia coli.

One of the important environmental issues is the quality of surface waters in the world. Poland belongs to countries with a low quality of the inland waters. The sanitary condition of the five water reservoirs of south-east Poland was analyzed. Water and sediment samples were incubated on the selective and/or differential media. High concentrations of many common and pathogenic microbial indicators were shown in those samples. Those reservoirs are used by people, especially during summer. Because of the high epidemiological risk, detailed analysis of all inland waters should be performed routinely.

Background & aims Infrared spectroscopy is an increasingly common method for bacterial strains' testing. For the analysis of bacterial IR spectra, advanced mathematical methods such as artificial neural networks must be used. The combination of these two methods has been used previously to analyze taxonomic affiliation of bacteria. The aim of this study was the classification of Escherichia coli strains in terms of susceptibility/resistance to cephalothin on the basis of their infrared spectra. The infrared spectra of 109 uropathogenic E. coli strains were measured. These data are used for classification of E. coli strains by using designed artificial neural networks. Results The most efficient artificial neural networks classify the E. coli sensitive/resistant strains with an error of 5%. Conclusions Bacteria can be classified in terms of their antibiotic susceptibility by using infrared spectroscopy and artificial neural networks.