Molecular Investigation of the Fatal Bloodstream Candida orthopsilosis Infection Case following Gastrectomy.

Candida orthopsilosis represents a closely related cryptic genospecies of Candida parapsilosis complex-misidentified in routine diagnostic assays. This is emerging in settings where central venous catheters, invasive medical interventions, and echinocandin treatments are most likely to be used. A 59-year-old, non-neutropenic male patient, was admitted to an intensive care unit (ICU) due to respiratory distress syndrome, following a partial gastrectomy. As a result of duodenal stump leakage, re-laparotomy was required, abdominal drains were provided and central line catheters were exchanged. Multiple isolates of Candida orthopsilosis drawn from consecutive blood cultures were identified, despite ongoing echinocandin therapy and confirmed in vitro echinocandins susceptibility of the isolated strain. Species identification was verified via ITS region sequencing. Herein, we report the well-documented-per clinical data and relevant laboratory diagnosis-first case of a bloodstream infection caused by Candida orthopsilosis in Poland.

TFF3 as a Diagnostic Biomarker in Kidney Transplant Patients.

Intestinal trefoil factor 3 (TFF3) is a protein secreted by many cell types, and its serum and urine levels vary in patients with kidney disease. Therefore, the present study aimed to determine the diagnostic value of TFF3 in allogeneic kidney transplant patients included in the one-year follow-up. To analyze the influence of the diagnostic method used, we studied the type of biological material and the time elapsed since renal transplantation on the parameter's value. The study also aimed to investigate the relationship between TFF3 levels and creatinine and estimated glomerular filtration rate (eGFR) values in the serum and urine of the patients studied. The study used blood and urine samples from adult patients (n = 19) 24-48 h, 6 months, and 12 months after kidney transplantation. We collected one-time blood and urine from healthy subjects (n = 5) without renal disease. We applied immunoenzymatic ELISA and xMap Luminex flow fluorimetry to determine TFF3 in serum and urine. There was a significant difference in TFF3 levels in the serum of patients collected on the first one or two days after kidney transplantation compared to the control group (determined by ELISA and Luminex) and six months and one year after kidney transplantation (ELISA). We observed a correlation between creatinine concentration and urinary TFF3 concentration (ELISA and Luminex) and a negative association between eGFR and urinary (ELISA) and serum (Luminex) TFF3 concentration in patients on the first and second days after kidney transplantation. We noted significant correlations between eGFR and TFF3 levels in the serum and urine of patients determined by the two methods six months and one year after transplantation. In women, we observed that urinary TFF3 concentration increased significantly with increasing creatinine and that with increasing eGFR, urinary TFF3 concentration determined by two methods decreased significantly. In the present study, the choice of diagnostic method for the determination of TFF3 in serum and urine significantly affected the concentration of this biomarker. The values of this parameter determined by ELISA were higher than those assessed using the Luminex assay. Based on the presented results, we can conclude that TFF3 has great potential to monitor renal transplant patients. Determination of this protein in parallel with creatinine and eGFR levels in serum and urine may provide helpful diagnostic information.

Clonal Diversity, Antimicrobial Susceptibility and Presence of Genes Encoding Virulence Factors in Staphylococcus aureus Strains Isolated from Cut Wound Infections.

The aim of the study was to evaluate the clonal relatedness and antimicrobial susceptibility in 52 Staphylococcus aureus strains isolated from cut wound infections in non-related community patients and to determine the presence of selected virulence genes. To analyse the clonal relatedness of investigated strains, pulsed-field gel electrophoresis (PFGE) of macrorestricted DNA fragments was conducted. Antimicrobial susceptibility testing was performed using the AST-P644 card in the VITEK 2 Compact system. All strains were tested for the presence of selected virulence genes using Single and Multiplex PCR. All isolates were classified into 15 PFGE genotypes and seven unique patterns. The vast majority of investigated S. aureus strains were susceptible to all tested antimicrobial agents. Among examined S. aureus strains, 24 combinations of virulence factors were identified. 62.5% of S. aureus strains contained various egc types, alone or together with other staphylococcal enterotoxin genes. A high percentage (86.5%) of isolates harboured superantigen genes. The most frequent enterotoxin gene identified was encoding for sep. All S. aureus strains were classified as agr-positive, and the most frequent agr gene was agr-1. Our results indicate that all examined strains isolated from cut wound infections demonstrated high clonal diversity, diversified gene distribution and good susceptibility to antimicrobial agents.

The Utility of Novel Kidney Injury Biomarkers in Early Detection of CSA-AKI.

Cardiac surgery-associated acute kidney injury (CSA-AKI) is one of the most common complications of cardiac surgery procedures. In this study, the authors attempt to provide new data regarding the application of novel kidney injury biomarkers in the early diagnostics of CSA-AKI. 128 adult patients undergoing elective cardiac surgery procedures with the use of cardiopulmonary by-pass (CPB) were enrolled in this study. Novel kidney injury biomarkers were marked in the plasma and urine 6 h after weaning from the CPB. A significant difference in the postoperative biomarkers' concentration between the AKI and no-AKI group was found, regarding plasma IL-8, plasma TNF-α and urine NGAL, normalized for creatinine excretion (NGAL/Cr). These were also independent predictors of CSA-AKI. An independent risk factor for CSA-AKI proved to be preoperative CKD. Plasma IL-8 and TNF-α, as well as urine NGAL/Cr, are independent early indicators of CSA-AKI and pose a promising alternative for creatinine measurements. The cut-off points for these biomarkers proposed in this investigation should be confronted with more data and revised to achieve a suitable diagnostic value.

Regulatory and Enterotoxin Gene Expression and Enterotoxins Production in Staphylococcus aureus FRI913 Cultures Exposed to a Rotating Magnetic Field and trans-Anethole.

The study aimed to examine the influence of a rotating magnetic field (RMF) of two different frequencies (5 and 50 Hz) on the expression of regulatory (agrA, hld, rot) and staphylococcal enterotoxin (SE-sea, sec, sel) genes as well as the production of SEs (SEA, SEC, SEL) by the Staphylococcus aureus FRI913 strain cultured on a medium supplemented with a subinhibitory concentration of trans-anethole (TA). Furthermore, a theoretical model of interactions between the bacterial medium and bacterial cells exposed to RMF was proposed. Gene expression and SEs production were measured using quantitative real-time PCR and ELISA techniques, respectively. Based on the obtained results, it was found that there were no significant differences in the expression of regulatory and SE genes in bacteria simultaneously cultured on a medium supplemented with TA and exposed to RMF at the same time in comparison to the control (unexposed to TA and RMF). In contrast, when the bacteria were cultured on a medium supplemented with TA but were not exposed to RMF or when they were exposed to RMF of 50 Hz (but not to TA), a significant increase in agrA and sea transcripts as compared to the unexposed control was found. Moreover, the decreased level of sec transcripts in bacteria cultured without TA but exposed to RMF of 50 Hz was also revealed. In turn, a significant increase in SEA and decrease in SEC and SEL production was observed in bacteria cultured on a medium supplemented with TA and simultaneously exposed to RMFs. It can be concluded, that depending on SE and regulatory genes expression as well as production of SEs, the effect exerted by the RMF and TA may be positive (i.e., manifests as the increase in SEs and/or regulatory gene expression of SEs production) or negative (i.e., manifests as the reduction in both aforementioned features) or none.

Antibacterial Activity of Selected Essential Oil Compounds Alone and in Combination with ß-Lactam Antibiotics Against MRSA Strains.

This study aimed to determine the effect of selected essential oil compounds (EOCs) on the antibacterial activity of ß-lactam antibiotics (ßLAs) against methicillin-resistant Staphylococcus aureus (MRSA) strains. The following parameters were studied: antibiotic susceptibility testing, detection of mecA gene and evaluation of genotypic relativity of isolates using molecular techniques, analysis of chemical composition applying Fourier-transform infrared (FTIR) spectroscopy, and determination of antibacterial activity of EOCs alone and in combination with ßLAs against MRSA strains using microdilution and checkerboard methods. It was found that all isolates expressed MRSA and resistance phenotypes for macrolides, lincosamides, and streptogramins B. All isolates harbored the mecA gene and belonged to three distinct genotypes. Eight of the 10 EOCs showed efficient antimicrobial activity against the MRSA reference strain. The analysis of interaction between EOCs and ßLAs against the MRSA reference strain revealed a synergistic and additive effect of the following combinations: methicillin (Met)-linalyl acetate (LinAc), penicillin G (Pen)-1,8-cineole (Cin), and Pen-LinAc. Analysis of EOC-ßLA interactions showed a synergistic and additive effect in the following combinations: Met-LinAc (against low- and high-level ßLAs resistance strains), Pen-Cin, and Pen-LinAc (against low-level ßLAs resistance strains). It was also confirmed that changes in phosphodiester, -OH, -CH2 and -CH3 groups may change the interactions with ßLAs. Moreover, the presence of two CH3O- moieties in the Met molecule could also play a key role in the synergistic and additive mechanism of LinAc action with Met against MRSA strains. Direct therapy using a Met-LinAc combination may become an alternative treatment method for staphylococcal infections caused by MRSA. However, this unconventional therapy must be preceded by numerous cytotoxicity tests.

Innate Immune Response against Staphylococcus aureus Preincubated with Subinhibitory Concentration of trans-Anethole.

The study aimed to analyze morphological and functional changes of Staphylococcus aureus cells due to trans-anethole (a terpenoid and the major constituent of fennel, anise, or star anise essential oils) exposition, and their consequences for human neutrophils phagocytic activity as well as IL-8 production (recognized as the major chemoattractant). The investigation included the evaluation of changes occurring in S. aureus cultures, i.e., staphyloxanthin production, antioxidant activities, cell size distribution, and cells composition as a result of incubation with trans-anethole. It was found that the presence of trans-anethole in the culture medium reduced the level of staphyloxanthin production, as well as decreased antioxidant activities. Furthermore, trans-anethole-treated cells were characterized by larger size and a tendency to diffuse in comparison to the non-treated cells. Several cell components, such as phospholipids and peptidoglycan, were found remarkably elevated in the cultures treated with trans-anethole. As a result of the aforementioned cellular changes, the bacteria were phagocytized by neutrophils more efficiently (ingestion and parameters associated with killing activity were at a higher level as compared to the control system). Additionally, IL-8 production was at a higher level for trans-anethole modified bacteria. Our results suggest that trans-anethole represents a promising measure in combating severe staphylococcal infections, which has an important translational potential for clinical applications.

Small Intestinal Bacterial Overgrowth (SIBO) and Twelve Groups of Related Diseases-Current State of Knowledge.

The human gut microbiota creates a complex microbial ecosystem, characterized by its high population density, wide diversity, and complex interactions. Any imbalance of the intestinal microbiome, whether qualitative or quantitative, may have serious consequences for human health, including small intestinal bacterial overgrowth (SIBO). SIBO is defined as an increase in the number of bacteria (103-105 CFU/mL), an alteration in the bacterial composition, or both in the small intestine. The PubMed, Science Direct, Web of Science, EMBASE, and Medline databases were searched for studies on SIBO and related diseases. These diseases were divided into 12 groups: (1) gastrointestinal disorders; (2) autoimmune disease; (3) cardiovascular system disease; (4) metabolic disease; (5) endocrine disorders; (6) nephrological disorders; (7) dermatological diseases; (8) neurological diseases (9); developmental disorders; (10) mental disorders; (11) genetic diseases; and (12) gastrointestinal cancer. The purpose of this comprehensive review is to present the current state of knowledge on the relationships between SIBO and these 12 disease groups, taking into account risk factors and the causal context. This review fills the evidence gap on SIBO and presents a biological-medical approach to the problem, clearly showing the groups and diseases having a proven relationship with SIBO, as well as indicating groups within which research should continue to be expanded.

Higher perfusion pressure and pump flow during cardiopulmonary bypass are beneficial for kidney function-a single-centre prospective study.

Background: Kidneys play an essential role in the circulatory system, regulating blood pressure and intravascular volume. They are also set on maintaining an adequate filtration pressure in the glomerulus. During the CPB, a decrease in systemic blood pressure and hemoglobin concentration may lead to renal ischemia and subsequent acute kidney injury. Methods: One hundred nine adult patients were prospectively enrolled in this study. The intervention in this study was increasing the flow of the CPB pump to reach the target MAP of > 90 mmHg during the procedure. The control group had a standard pump flow of 2.4 L/min/m2. Results: Standard pump flow of 2.4 L/min/m2 resulted in mean MAP < 90 mmHg during the CPB in most patients in the control group. Maintaining a higher MAP during CPB in this study population did not affect CSA-AKI incidence. However, it increased the intraoperative and postoperative diuresis and decreased renin release associated with CPB. Higher MAP during the CPB did not increase the incidence of cerebrovascular complications after the operation; patients in the highest MAP group had the lowest incidence of postoperative delirium, but the result did not obtain statistical significance. Conclusion: Maintaining MAP > 90 mmHg during the CPB positively impacts intraoperative and postoperative kidney function. It significantly reduces renal hypoperfusion during the procedure compared to MAP < 70 mmHg. MAP > 90 mmHg is safe for the central nervous system, and preliminary results suggest that it may have a beneficial impact on the incidence of postoperative delirium.

Endogenous or exogenous origin of vaginal candidiasis in Polish women?

Vaginal candidiasis is a common problem of clinical practice. Many studies have been conducted to explain its origin but only a few have included Polish women. The aim of the study was to determine the prevalence and similarity of oral, anal and vaginal Candida albicans strains isolated from Polish women with vaginal candidiasis. The study involved 20 from 37 recruited women. Swab samples were collected from their vagina, anus, and oral cavity at two-month intervals. All the women were treated with nystatin. Yeast were recovered and identified by the germ-tube test, API /Vitek system, typed by API ZYM and RAPD-PCR. Chi-square test was used to analyze the data. A total of 170 Candida albicans isolates were recovered from 180 samples collected 3 times from 3 sites of 20 women. Positive yeast vaginal cultures were found in all patients before administration of nystatin. Vaginal yeast recovery rate was decreased statistically significant in both follow-up visits (p= 0.001; p= 0.003). The same and different genotypes/biotypes were found concomitantly in a few body sites and/ or repeatedly at time interval from the same body site. The results support the concept of dynamic exchange of yeast within one woman and endogenous or exogenous origin of vaginal candidiasis.

Proteomic study on the lymphocytes from pregnant Wistar rat females treated with immunosuppressive regimen.

Kidney transplantation remains the therapeutic option for patients with end-stage kidney disease. Current immunosuppressive regimens are efficient in combating acute kidney rejection. However, insights into chronic kidney allograft injury remains limited. Simultaneously, pregnancy is more common after kidney transplantation than during dialysis treatment. Due to ethical issues, comprehensive studies on the impact of immunosuppressive regimens on pregnancy are challenging. The study aimed to investigate the proteomic status of lymphocytes obtained from pregnant female rats under immunosuppressive treatment. The experiment involved a group of 10 female, pregnant Wistar rats, five of which were treated with tacrolimus, mofetil mycophenolate, and glucocorticosteroids; five were used as control. The lymphocytes were obtained and analyzed with mass spectrometry. Measurements were processed by a database search in the ProteinPilot software with a cutoff of 1% false discovery rate. The outcomes were verified statistically by a t-test (p value < 0.05) regarding proteins up- and downregulation. A total of 2082 proteins were identified in all experiments. Eight hundred five proteins were quantified in an absolute manner in a data-independent acquisition-total protein approach analysis. Ninety-five proteins were recognized as present at different concentrations in analyzed groups and were annotated to intracellular pathways. The proteins involved in nonsense-mediated decay and L13a-mediated translational silencing of ceruloplasmin expression were recognized as downregulated. The set of proteins clinically identified as acute phase proteins was upregulated. Despite the blockade of adaptive cellular immunity, the lymphocytes in the analyzed group reveal sustained proinflammatory status with decreased ability to regulate translation. This potentially affects pregnancy and immunity.

Immunomodulatory effects of trans-anethole-treated Staphylococcus aureus Newman strain.

In our former studies based on a human whole-blood model infected with trans-anethole (TA)-treated Staphylococcus aureus Newman strain, we have observed that selected parameters/mechanisms of innate and acquired immune response were more enhanced in comparison to samples infected with non-treated bacteria. Due to this observation, the current study aimed to evaluate the concentration of selected proteins involved in both types of responses (IL-1α, IL-1ß, IL-2, IL-6, IL-12, IL-17, TNF-α, IFN-γ, G-CSF, C5a, CCL1-CCL5, CXCL1, CXCL2, CXCL9-CXCL11, MMP-8, TLR2, and PGLYRP1) in healthy participants' plasma after blood stimulation of TA-treated S. aureus Newman strain. Determination of analyzed protein concentration was conducted using Luminex and ELISA assays. Based on the results, it has been proven that the immunomodulatory potential of TA-treated S. aureus Newman strain on increasing IL-1ß, IL-6, TNF-α, IL-12, G-CSF, C5a, CCL2-CCL4, CXCL1, CXCL2, MMP-8 and PGLYRP1 levels in plasma. Moreover, it has been also demonstrated an association between TNF-α and CCL4 in a blood model infected with TA-treated cells. More research is warranted to find more underlying mechanisms involved in the effects of TA-treated S. aureus Newman in human blood, mainly whether the observed "immunity boost" can be regulated after bacteria elimination. Therefore, the potential of TA should be further explored to understand under which conditions it might help treat or prevent infections caused by S. aureus.

Digital PCR (dPCR) Quantification of miR-155-5p as a Potential Candidate for a Tissue Biomarker of Inflammation in Rabbits Infected with Lagovirus europaeus/Rabbit Hemorrhagic Disease Virus (RHDV).

MicroRNAs (miRNAs, miRs) are a group of small, 17-25 nucleotide, non-coding RNA sequences that, in their mature form, regulate gene expression at the post-transcriptional level. They participate in many physiological and pathological processes in both humans and animals. One such process is viral infection, in which miR-155 participates in innate and adaptive immune responses to a broad range of inflammatory mediators. Recently, the study of microRNA has become an interesting field of research as a potential candidate for biomarkers for various processes and disease. To use miRNAs as potential biomarkers of inflammation in viral diseases of animals and humans, it is necessary to improve their detection and quantification. In a previous study, using reverse transcription real-time quantitative PCR (RT-qPCR), we showed that the expression of ocu-miR-155-5p in liver tissue was significantly higher in rabbits infected with Lagovirus europaeus/Rabbit Hemorrhagic Disease Virus (RHDV) compared to healthy rabbits. The results indicated a role for ocu-miR-155-5p in Lagovirus europaeus/RHDV infection and reflected hepatitis and the impairment/dysfunction of this organ during RHD. MiR-155-5p was, therefore, hypothesized as a potential candidate for a tissue biomarker of inflammation and examined in tissues in Lagovirus europaeus/RHDV infection by dPCR. The objective of the study is the absolute quantification of ocu-miR-155-5p in four tissues (liver, lung, kidney, and spleen) of rabbits infected with Lagovirus europaeus/RHDV by digital PCR, a robust technique for the precise and direct quantification of small amounts of nucleic acids, including miRNAs, without standard curves and external references. The average copy number/µL (copies/µL) of ocu-miRNA-155-5p in rabbits infected with Lagovirus europaeus GI.1a/Rossi in the liver tissue was 12.26 ± 0.14, that in the lung tissue was 48.90 ± 9.23, that in the kidney tissue was 16.92 ± 2.89, and that in the spleen was 25.10 ± 0.90. In contrast, in the tissues of healthy control rabbits, the average number of copies/µL of ocu-miRNA-155-5p was 5.07 ± 1.10 for the liver, 23.52 ± 2.77 for lungs, 8.10 ± 0.86 for kidneys, and 42.12 ± 3.68 for the spleen. The increased expression of ocu-miRNA-155-5p in infected rabbits was demonstrated in the liver (a fold-change of 2.4, p-value = 0.0003), lung (a fold-change of 2.1, p-value = 0.03), and kidneys (a fold-change of 2.1, p-value = 0.01), with a decrease in the spleen (a fold-change of 0.6, p-value = 0.002). In the study of Lagovirus europaeus/RHDV infection and in the context of viral infections, this is the first report that shows the potential use of dPCR for the sensitive and absolute quantification of microRNA-155-5p in tissues during viral infection. We think miR-155-5p may be a potential candidate for a tissue biomarker of inflammation with Lagovirus europaeus/RHDV infection. Our report presents a new path in discovering potential candidates for the tissue biomarkers of inflammation.

Macroelement and Microelement Levels in the Urine in Experimental Acanthamoebiasis.

Free-living amoebas can impact the excretion of macroelements and microelements in urine. The aim of the present study was to examine the concentrations of macroelements, including calcium (Ca), phosphorus (P), sodium (Na), potassium (K), and magnesium (Mg), as well as microelements such as manganese (Mn), zinc (Zn), copper (Cu), iron (Fe), and chromium (Cr), in the urine during acanthamoebiasis while considering the host's immunological status. This is the first study to show an increase in urinary excretion of Ca, Mn, Cu, Fe, Na, and Cr, along with a decreased excretion of K, in immunocompetent mice 16 days post Acanthamoeba sp. infection. In the final phase of infection (24 dpi), there was a further decrease in urinary K excretion and a lower level of P in Acanthamoeba sp. infected immunocompetent hosts. During acanthamoebiasis in immunosuppressed hosts, increased excretion of Zn, Fe, and Cr was observed at the beginning of the infection, and increased Na excretion only at 16 days post Acanthamoeba sp. infection. Additionally, host immunosuppression affected the concentration of Fe, Cr, Zn, Cu, Mn, and Ca in urine.

Staphyloxanthin inhibitory potential of trans-anethole: A preliminary study.

The reduction of staphyloxanthin (STX) production in Staphylococcus aureus under trans-anethole (TA) influence was proven in former studies. However, no tests concerning the impact of TA on a biosynthetic pathway of this carotenoid pigment have been published so far. Thus, for the first time, the present preliminary study evaluated the influence of TA on the expression level of genes (crtOPQMN operon and aldH) encoding STX pathway enzymes. Additional attention was paid to the identification of STX and its intermediates. Gene expression and identification of extracted compounds were conducted using quantitative real-time PCR and HPLC-MS techniques, respectively. The analyzes showed no difference in crtM, crtN, crtO, crtP, crtQ, and aldH gene expression between bacterial samples isolated from the non-stimulated (control) medium and the stimulated one with TA. Compared to the control group that showed the presence of all metabolic intermediates and STX, the TA-treated bacteria were characterized by a lack or a significant reduction of the majority of compounds, except 4,4'-diaponeurosporenoate, the content of which was elevated in the TA-treated sample. Moreover, in silico molecular docking analysis revealed that TA is capable to create relatively strong interactions with both 4,4'-diapophytoene synthase and 4,4'-diapophytoene desaturase. The preliminary findings indicate that the previously observed TA effect reducing the number of S. aureus colonies pigmentation is probably not associated with the expression levels of genes encoding STX pathway enzymes. It has been proven that adding TA to the medium can interfere with the formation of STX at different levels of its biosynthetic pathway.

Enhancement of neutrophil chemotaxis by trans-anethole-treated Staphylococcus aureus strains.

This study aimed to analyze the chemotactic response of differentiated HL-60 neutrophil-like (dHL-60) cells to trans-anethole (TA)-treated Staphylococcus aureus strains. Special attention was paid to evaluate the influence of TA on the chp gene expression level, as well as molecular docking and molecular dynamics (MD) simulation studies on interactions of TA with chemotaxis inhibitory protein of S. aureus (CHIPS). The following parameters were studied: susceptibility to TA using the agar diffusion method, the chp gene detection and its expression under TA influence, and clonal diversity of S. aureus strains using molecular techniques. Furthermore, a chemotactic response of dHL-60 cells to TA-treated S. aureus using Boyden chamber assay was detected and molecular modeling using both the docking methodology and unbiased MD simulations was conducted. It was found that TA showed antibacterial activity against all strains. Three genotypes and one unique pattern were distinguished among the strains. 50% of the isolates were chp-positive. It was observed that TA reduced/inhibited chp gene expression in most S. aureus strains. Enhanced chemotactic response of dHL-60 cells to TA-treated S. aureus strains was also noted. This correlation was similar for both chp-positive and chp-negative strains. Both molecular docking and MD simulations studies confirmed that TA is preferentially bound in the complement component 5a/CHIPS interface interaction region and can interfere with any processes exploiting this binding cavity. It has been proven that dHL-60 cells exhibited a higher chemotactic response to TA-treated S. aureus strains in comparison to non-treated bacteria, regardless of the achieved expression of the chp gene or its lack. Nevertheless, further analyses are required to understand this mechanism better.

Improved RAPD Method for Candida parapsilosis Fingerprinting.

Recently, methods based on the analysis of arbitrarily amplified target sites of genome microorganisms have been extensively applied in microbiological studies, and especially in epidemiological studies. The range of their application is limited by problems with discrimination and reproducibility resulting from a lack of standardized and reliable methods of optimization. The aim of this study was to obtain optimal parameters of the Random Amplified Polymorphic DNA (RAPD) reaction by using an orthogonal array as per the Taguchi and Wu protocol, modified by Cobb and Clark for Candida parapsilosis isolates. High Simpson's index values and low Dice coefficients obtained in this study indicated a high level of interspecies DNA polymorphism between C. parapsilosis strains, and the optimized RAPD method proved useful in the microbiological and epidemiological study.

The Most Promising Biomarkers of Allogeneic Kidney Transplant Rejection.

Clinical transplantology is a constantly evolving field of medicine. Kidney transplantation has become standard clinical practice, and it has a significant impact on reducing mortality and improving the quality of life of patients. Allogenic transplantation induces an immune response, which may lead to the rejection of the transplanted organ. The gold standard for evaluating rejection of the transplanted kidney by the recipient's organism is a biopsy of this organ. However, due to the high invasiveness of this procedure, alternative diagnostic methods are being sought. Therefore, the biomarkers may play an essential predictive role in transplant rejection. A review of the most promising biomarkers for early diagnosis and prognosis prediction of allogenic kidney transplant rejection summarizes novel data on neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (KIM-1), C-X-C motif chemokine 10 (CXCL-10), cystatin C (CysC), osteopontin (OPN), and clusterin (CLU) and analyses the dynamics of changes of the biomarkers mentioned above in kidney diseases and the mechanism of rejection of the transplanted kidney.

Cross-Reactive Results in Serological Tests for Borreliosis in Patients with Active Viral Infections.

Currently, serological tests for Lyme disease (LD), routinely performed in laboratories following the European Concerted Action on Lyme Borreliosis recommendations as part of two-stage diagnostics, are often difficult to interpret. This concerns both the generation of false positive and negative results, which frequently delay the correct diagnosis and implementation of appropriate treatment. The above problems result from both morphological and antigenic variability characteristics for the life strategy of the spirochete Borrelia burgdorferi sensu lato, a complicated immune response, and imperfections in diagnostic methods. The study aimed to check the reactivity of sera from 69 patients with confirmed infection with Epstein-Barr virus (EBV), cytomegalovirus (CMV) and BK virus (BKV) with Borrelia antigens used in serological tests: indirect immunofluorescence (IIFT), enzyme-linked immunosorbent (ELISA) and immunoblot (IB). In the group of patients infected with EBV, the highest percentage of positive/borderline anti-Borrelia IgM and IgG results was obtained in the following tests: IIFT (51.9% for IgM, 63.0% for IgG), ELISA (22.2% for IgM, 29.6% for IgG) and IB (11.1% for IgM, 7.4% for IgG). In the group of CMV-infected patients, the highest percentage of positive/borderline anti-Borrelia IgM results were obtained in the following tests: IB (23.1%), IIFT (15.4%) and ELISA (7.7%), while in the IgG class in the IIFT (15.4%), IB (11.5%) and ELISA (3.9%) tests. In the group of patients infected with BKV, the highest percentage of positive/borderline anti-Borrelia IgM results was obtained in the following tests: IIFT (25.0%), IB (25.0%) and ELISA (3.9%), and in the IgG class in the tests: IB (50.0%), IIFT (6.2%) and ELISA (6.2%). The native flagellin (p41) and OspC proteins were the most frequently detected Borrelia antigens in all studied groups of patients in both classes of antibodies. Similar to other authors, the study confirmed the fact that serological tests used in the diagnosis of LD have a high potential to generate false positive results in patients with active viral infections, which may be related to cross-reacting antibodies appearing during the most common polyclonal activation of T/B lymphocytes, activated by viral superantigens.

Could the Optiplex Borrelia assay replace the traditional, two-step method of diagnosing Lyme disease?

INTRODUCTION AND OBJECTIVE: Serological assays for Lyme disease (LD) routinely performed in laboratories often give inconclusive results, thereby making correct diagnosis difficult and delaying treatment. The aim of the study was to assess the usefulness of a commercial Optiplex Borrelia (OB) assay in the serological diagnostics of LD. Based on the results obtained in a previous study on the seroreactivity of the sera of patients with LD to Borrelia spp. antigens using enzyme immunoassays (ELISA) and immunoblotting (IB), the same sera were re-analyzed using the OB assay. RESULTS: The assays carried out with the use of OB method showed a statistically significant lower number of positive/borderline results for the presence of IgM antibodies, compared to the ELISA assay. Moreover, statistically lower positive/borderline results were obtained for antibodies in the IgG class with use of the OB method, compared to the IB assay and a two-stage diagnostic protocol (ELISA with IB). The specificity analysis showed that in both the IB and OB assays, anti-OspC IgM and anti-p41 antibodies were detected. Additionally, high positive/borderline values were found in the OB assay for native antigens derived from B. afzelii lysate. The IB assay most frequently detected antibodies against OspC, p39 (BmpA) and VlsE proteins in the IgG class. There were fewer positives/borderlines for anti-p41-I B. afzelii antibodies in the OB assay and a higher number for antigens: VlsE-C6, p18 B. afzelii (DbpA), and p39 B. afzelii (BmpA). CONCLUSIONS: Answering the question whether the OB assay could replace the traditional, two-step method of LD diagnostics, it can be concluded that it could not. It can be used to diagnose LD only as a complementary assay and not as an optimal and dedicated method of Borrelia spp. infection detection.