Assessing the Interpretation of Molecular Test Results in the Diagnosis of Bloodstream Infections.

A retrospective study at the 4th Military Clinical Hospital in Wroclaw, Poland, assessed PCR testing alongside blood cultures to guide antimicrobial therapy decisions in hospitalized patients, to determine how much time the results of the molecular tests preceded conventional methods. Among 118 patients, Staphylococcus aureus (37%) and Escherichia coli (21%) were the most common bloodstream infection agents. Blood cultures utilized the BacT/ALERT 3D system, and molecular diagnostics were conducted using the FilmArray platform with the BIOFIRE BCID2 panel. Methicillin susceptibility was observed in 66% of S. aureus strains, while 26% of Gram-negative bacilli exhibited an ESBL phenotype. Therapeutic decisions based on molecular test results were often incorrect for S. aureus infections, particularly MSSA (64.5%), but generally accurate for Gram-negative bacilli. The median times from positive blood culture to BCID2 and pathogen identification/susceptibility were 10 h and 52 h, respectively. Molecular diagnostics facilitated faster initiation of appropriate antibiotic therapy, highlighting the need to educate medical staff on proper interpretation. Consulting within an antimicrobial stewardship program (ASP) could enhance the benefits of implementing molecular methods in bloodstream infection diagnostics.

Evaluation of the in vitro susceptibility of clinical isolates of NDM-producing Klebsiella pneumoniae to new antibiotics included in a treatment regimen for infections.

Background: Due to the growing resistance to routinely used antibiotics, the search for new antibiotics or their combinations with effective inhibitors against multidrug-resistant microorganisms is ongoing. In our study, we assessed the in vitro drug susceptibility of Klebsiella pneumoniae strains producing New Delhi metallo-ß-lactamases (NDM) to antibiotics included in the Infectious Diseases Society of America (IDSA) and European Society of Clinical Microbiology and Infectious Diseases (ESCMID) recommendations. Methods: A total of 60 strains of NDM-producing K. pneumoniae were obtained from different patients hospitalized at the 4th Military Hospital in Wroclaw between 2019 and 2022 and subjected to drug susceptibility to selected antibiotics, including the effects of drug combinations. Results: Among the tested antibiotics, the highest sensitivity (100%) was observed for cefiderocol, eravacycline (interpreted according to the European Committee on Antimicrobial Susceptibility Testing [EUCAST]), and tigecycline. Sensitivity to intravenous fosfomycin varied depending on the method used. Using the "strip stacking" method, determining cumulative sensitivity to ceftazidime/avibactam and aztreonam demonstrated 100% in vitro sensitivity to this combination among the tested strains. Conclusion: The in vitro susceptibility assessment demonstrated that, the best therapeutic option for treating infections caused by carbapenemase-producing strains seems to be a combination of ceftazidime/avibactam with aztreonam. Due to the safety of using both drugs, cost effectiveness, and the broadest indications for use among the tested antibiotics, this therapy should be the first-line treatment for carbapenemase-producing Enterobacterales infections. Nevertheless, a comprehensive evaluation of the efficacy of treating infections caused by NDM-producing K. pneumoniae strains should include not only in vitro susceptibility assessment but also an analysis of clinical cases.

Impact of the COVID-19 Pandemic on the Epidemiology of Bloodstream Infections in Hospitalized Patients-Experience from a 4th Military Clinical Hospital in Poland.

Bloodstream infections (BSIs) are associated with high mortality and inappropriate or delayed antimicrobial therapy. The purpose of this study was to investigate the impact of the COVID-19 pandemic on the epidemiology of BSIs in hospitalized patients. The research aimed to compare the incidence of BSIs and blood culture results in patients hospitalized before and during the COVID-19 pandemic. METHODS: Retrospective and prospective data were collected from blood cultures obtained from 4289 patients hospitalized between June 2018 and July 2022. Two groups of patients were distinguished: those with BSIs admitted during the pre-COVID-19 period and those admitted during the COVID-19 surge. Demographic and clinical data, blood cytology, and biochemistry results were analyzed, and the usefulness of PCT was assessed in patients with COVID-19. RESULTS: The study showed a significant increase in the incidence of BSIs during the pandemic compared to the pre-COVID-19 period. Positive blood cultures were obtained in 20% of patients hospitalized during the pandemic (vs. 16% in the pre-COVID-19 period). The incidence of BSIs increased from 1.13 to 2.05 cases per 1000 patient days during COVID-19, and blood culture contamination was more frequently observed. The mortality rate was higher for patients hospitalized during the COVID-19 pandemic. An increased frequency of MDRO isolation was observed in the COVID-19 period. CONCLUSIONS: The incidence of BSIs increased and the mortality rate was higher in the COVID-19 period compared to the pre-COVID-19 period. The study showed limited usefulness of procalcitonin in patients with COVID-19, likely due to the administered immunosuppressive therapy.

Reliability of E-Tests and the Phoenix Automated Method in Assessing Susceptibility to IV Fosfomycin-Comparative Studies Relative to the Reference Method.

The agar dilution method (ADM) recommended for IV fosfomycin (IV FOS) is complex and labor-intensive. Keeping in mind the reality of everyday laboratory work, we have evaluated the agreement of IV FOS susceptibility results obtained using the E-test and the Phoenix system with the results obtained using the ADM. MATERIALS AND METHODS: The tests were performed on 860 strains. To evaluate susceptibility to IV FOS, BioMerieux E-tests (bioMerieux, Warsaw, Poland), BD Phoenix panels (BD Phoenix, Sparks, MD, USA), and the ADM were used. Clinical interpretation was performed in accordance with EUCAST Guidance (v12.0, 2021). The significance of the E-test and the Phoenix was analyzed in relation to the ADM by defining categorical agreement (CA), major error (ME), and very major error (VME). Essential agreement (EA) has also been defined for the E-test. A method was considered reliable, in accordance with ISO 20776-2:2007, when CA and EA were above 89.9% and VME was <3%. RESULTS: A categorical agreement of >98.9% was demonstrated between the E-test and the ADM for overall strains and for Echerichia coli, ESBL-producing Enterobacterales, and Staphylococcus aureus, while between the Phoenix and the ADM, a CA of >98.9% was shown only for Escherichia coli, Staphylococcus aureus, and Proteus spp. A very major error rate of <3% was obtained only for Staphylococcus aureus and MBL-producing Pseudomonas evaluated by both the E-test and the Phoenix. An essential agreement of >98.9% between the E-test and the ADM has not been demonstrated for any of the tested groups of strains. The Phoenix yielded more VMEs than the E-test (50 and 46, respectively). The highest VME rate was demonstrated using the Phoenix method for Enterobacter spp. (53.83%). CONCLUSIONS: Both the E-test and the Phoenix have turned out to be reliable in assessing IV FOS susceptibility only for Staphylococcus aureus (CA > 89.9% and VME < 3%). For the remaining tested groups of strains and genera, the simultaneous high CA rate and low VME rate required by ISO were not achieved. Both methods fared particularly badly in detecting strains resistant to IV.

Assessment of the Susceptibility of Clinical Gram-Negative and Gram-Positive Bacterial Strains to Fosfomycin and Significance of This Antibiotic in Infection Treatment.

Multidrug resistance of bacteria has prompted intensive development work on new medicines, but also the search for effective options among the oldest antibiotics. Although intravenous fosfomycin (IVFOS) seems to be an interesting proposal, the recommended agar dilution method for susceptibility determination poses a major problem in routine diagnostic testing. As a consequence, there is a lack of comprehensive data on the frequency of isolation of susceptible or resistant strains. This fact triggered the disposition of EUCAST concerning the revision of IVFOS breakpoints (BPs), including withdrawal of BPs for Enterobacterales (excluding E. coli) and coagulase-negative staphylococci. Therefore, the aim of this study was to assess the activity of fosfomycin against numerous clinical strains using recommended methods. Materials and methods: A total of 997 bacterial strains were tested from the following genera: Enterobacterales, Pseudomonas spp., Staphylococcus spp., Acinetobacter spp., and Enterococcus spp., for which there are currently no BPs. The strains were isolated from various clinical materials from patients hospitalized in five hospitals. During the investigation, the recommended agar dilution method was used. Susceptibility to other antibiotics and resistance mechanisms were determined using an automatic method (Phoenix) the disk diffusion method, and E-tests. MIC values of fosfomycin were estimated for all strains and for susceptible and multidrug-resistant (MDR) strains individually. Results: Except for Acinetobacter and Enterococcus, 83% of the strains were susceptible to IVFOS, including the largest percentage of S. aureus and E. coli. Klebsiella spp. turned out to be the least susceptible strains (66%). The highest proportion of susceptibility to fosfomycin was found among strains that were sensitive to other antibiotics (80.9%), and the lowest was found among Gram-negative carbapenemase-producing bacteria (55.6%) and ESBL+ bacteria (61.6%). The MIC evaluation revealed the lowest MIC50 and MIC90 values for S. aureus (0.5 mg/L and 1 mg/L, respectively) and E. coli (4 mg/L and 32 mg/L, respectively). The highest values of MIC50 were found for Acinetobacter spp. (256 mg/L), while the highest values of MIC90 were found for Acinetobacter spp. and Klebsiella spp. (256 mg/L and 512 mg/L, respectively). Conclusions: IVFOS appears to be suitable for the treatment of many infections, including the empirical treatment of polymicrobial infections and those caused by MDR strains, since the sensitivity of the studied strains to this antibiotic in different groups ranged from 66% to as much as 99%. Sensitivity to fosfomycin was also demonstrated by 60% of carbapenem-resistant strains; therefore, IVFOS is one of the few therapeutic options that can be effective against the most resistant Gram-negative rods. In light of the general consultation posted by EUCAST, obtaining data such as IVFOS MIC value distributions may be vital for the decision of implementing fosfomycin into breakpoint tables.

The Medium Composition Impacts Staphylococcus aureus Biofilm Formation and Susceptibility to Antibiotics Applied in the Treatment of Bone Infections.

The biofilm-associated infections of bones are life-threatening diseases, requiring application of dedicated antibiotics in order to counteract the tissue damage and spread of microorganisms. The in vitro analyses on biofilm formation and susceptibility to antibiotics are frequently carried out using methods that do not reflect conditions at the site of infection. To evaluate the influence of nutrient accessibility on Staphylococcus aureus biofilm development in vitro, a cohesive set of analyses in three different compositional media was performed. Next, the efficacy of four antibiotics used in bone infection treatment, including gentamycin, ciprofloxacin, levofloxacin, and vancomycin, against staphylococcal biofilm, was also assessed. The results show a significant reduction in the ability of biofilm to grow in a medium containing elements occurring in the serum, which also translated into the diversified changes in the efficacy of used antibiotics, compared to the setting in which conventional media were applied. The differences indicate the need for implementation of adequate in vitro models that closely mimic the infection site. The results of the present research may be considered an essential step toward the development of in vitro analyses aiming to accurately indicate the most suitable antibiotic to be applied against biofilm-related infections of bones.

Modifications of bacterial cellulose in wound care.

Wound infection may occur in acute and chronic wounds, wounds resulting from surgery or traffic accidents, and burns. Regardless of the extent and cause of the wound, prompt treatment is essential in reducing the patient's pain and limiting the spread of contamination. Improper wound care and associated chronic diseases may hinder the therapeutic success. Bacterial cellulose (BC) is highly biocompatible and has no cytotoxic effect on cells engaged in wound healing, such as fibroblasts and keratinocytes. Its high hydration level guarantees the maintenance of a moist wound environment. High mechanical strength, flexibility and resistance to damage make BC a promising material for dressings. Unfortunately, it does not display an inhibitory effect on bacterial growth. Introducing antimicrobial agents into the structure of BC has been a subject of many studies. This paper aims to present the latest reports on the possibility of the absorption of bacteriostatic and bactericidal agents in BC, such as metal particles, essential oils, antibiotics, antiseptics, and wound irrigation solutions. Moreover, the modifications in BC culture and post-production treatments in order to improve its physical properties are discussed.

The Antimicrobial and Antibiofilm In Vitro Activity of Liquid and Vapour Phases of Selected Essential Oils against Staphylococcus aureus.

The high resistance of staphylococcal biofilm against antibiotics and developing resistance against antiseptics induces a search for novel antimicrobial compounds. Due to acknowledged and/or alleged antimicrobial activity of EOs, their application seems to be a promising direction to follow. Nevertheless, the high complexity of EOs composition and differences in laboratory protocols of the antimicrobial activity assessment hinders the exact estimation of EOs effectiveness. To overcome these disadvantages, in the present work we analysed the effectiveness of volatile and liquid forms of seven EOs (derived from thyme, tea tree, basil, rosemary, eucalyptus, lavender, and menthol mint) against 16 staphylococcal biofilm-forming strains using cohesive set of in vitro techniques, including gas chromatography-mass spectrometry, inverted Petri dish, modified disk-diffusion assay, microdilution techniques, antibiofilm dressing activity measurement, AntiBioVol protocol, fluorescence/confocal microscopy, and dynamic light scattering. Depending on the requirements of the technique, EOs were applied in emulsified or non-emulsified form. The obtained results revealed that application of different in vitro techniques allows us to get a comprehensive set of data and to gain insight into the analysed phenomena. In the course of our investigation, liquid and volatile fractions of thyme EO displayed the highest antibiofilm activity. Liquid fractions of rosemary oil were the second most active against S. aureus. Vapour phases of tea tree and lavender oils exhibited the weakest anti-staphylococcal activity. The size of emulsified droplets was the lowest for T-EO and the highest for L-EO. Bearing in mind the limitations of the in vitro study, results from presented analysis may be of pivotal meaning for the potential application of thymol as a antimicrobial agent used to fight against staphylococcal biofilm-based infections.

Activity of Liquid and Volatile Fractions of Essential Oils against Biofilm Formed by Selected Reference Strains on Polystyrene and Hydroxyapatite Surfaces.

Biofilms are surface-attached, structured microbial communities displaying higher tolerance to antimicrobial agents in comparison to planktonic cells. An estimated 80% of all infections are thought to be biofilm-related. The drying pipeline of new antibiotics efficient against biofilm-forming pathogens urges the search for alternative routes of treatment. Essential Oils (EOs), extracted from medicinally important plants, are a reservoir of bioactive compounds that may serve as a foothold in investigating novel antibiofilm compounds. The aim of this study was to compare antimicrobial activity of liquid and volatile fractions of tested EOs against biofilm-forming pathogens using different techniques. In this research, we tested five EOs, extracted from Syzygium aromaticum L., Boswelia serrata Roxb., Juniperus virginiana L., Pelargonium graveolens L. and Melaleuca alternifolia Cheel., against planktonic and biofilm forms of five selected reference strains, namely Staphylococcus aureus, Enterococcus faecalis, Klebsiella pneumoniae, Pseudomonas aeruginosa, Escherichia coli, and Candida albicans. To obtain cohesive results, we applied four various methodological approaches: to assess the activity of the liquid fraction of EOs, disc diffusion and the microdilution method were applied; to test EOs' volatile fraction, the AntiBioVol assay and modified Antibiofilm Dressing Activity Measurement (A.D.A.M.) were used. The molecular composition and dynamics of antimicrobial substances released from specific EOs was measured using Gas Chromatography-Mass Spectrometry (GC-MS). The antimicrobial potency of EO's volatile fraction against biofilm formed by tested strains differed from that of the liquid fraction and was related to the molecular weight of volatile compounds. The liquid fraction of CW-EO and volatile fraction of F-EO acted in the strongest manner against biofilm of C. albicans. The addition of 0.5% Tween 20 to liquid phase, enhanced activity of G-EO against E. coli and K. pneumoniae biofilm. EO activity depended on the microbial species it was applied against and the chosen assessment methodology. While all tested EOs have shown a certain level of antimicrobial and antibiofilm effect, our results indicate that the choice of EO to be applied against a specific biofilm-forming pathogen requires careful consideration with regard to the above-listed aspects. Nevertheless, the results presented in this research contribute to the growing body of evidence indicating the beneficial effects of EOs, which may be applied to fight biofilm-forming pathogens.

Corrigendum to "Clinical Trials of Probiotic Strains in Selected Disease Entities".

[This corrects the article DOI: 10.1155/2020/8854119.].

The Minimum Inhibitory Concentration of Antibiotics: Methods, Interpretation, Clinical Relevance.

Inefficiency of medical therapies used in order to cure patients with bacterial infections requires not only to actively look for new therapeutic strategies but also to carefully select antibiotics based on variety of parameters, including microbiological. Minimal inhibitory concentration (MIC) defines in vitro levels of susceptibility or resistance of specific bacterial strains to applied antibiotic. Reliable assessment of MIC has a significant impact on the choice of a therapeutic strategy, which affects efficiency of an infection therapy. In order to obtain credible MIC, many elements must be considered, such as proper method choice, adherence to labeling rules, and competent interpretation of the results. In this paper, two methods have been discussed: dilution and gradient used for MIC estimation. Factors which affect MIC results along with the interpretation guidelines have been described. Furthermore, opportunities to utilize MIC in clinical practice, with pharmacokinetic /pharmacodynamic parameters taken into consideration, have been investigated. Due to problems related to PK determination in individual patients, statistical estimation of the possibility of achievement of the PK/PD index, based on the Monte Carlo, was discussed. In order to provide comprehensive insights, the possible limitations of MIC, which scientists are aware of, have been outlined.

Evaluation of 1,2-Benzothiazine 1,1-dioxide Derivatives In Vitro Activity towards Clinical-Relevant Microorganisms and Fibroblasts.

The global concern related with growing number of bacterial pathogens, resistant to numerous antibiotics, prone scientific environment to search for new antimicrobials. Antiseptics appear to be suitable candidates as adjunctive agents to antibiotics or alternative local treatment option aiming to prevent and treat infections. 1,2-benzothiazines are considered one the most promising of them. In this research twenty 1,2-benzothiazine 1,1-dioxide derivatives were scrutinized with regard to their biological activity. Three of them are new. For evaluation of compounds' activity against microbial pathogens, disk diffusion method and serial microdilution method was applied. To establish the cytotoxicity profile of tested 1,2-benzothiazines 1,1-dioxides derivatives, the cytotoxicity assay using fibroblasts L292 was performed. Antimicrobial activity of all tested compounds against Gram-positive Staphylococcus aureus and Enterococcus faecalis strains was higher than antimicrobial activity of DMSO solvent, which possesses antimicrobial activity itself. Gram-negative P. aeruginosa, E. coli and K. pneumoniae have shown susceptibility only to compounds 3e, 7i and 7l. None of tested compounds was effective against C. albicans. Compound 6g has demonstrated the strongest antimicrobial potency (MIC = 0.00975 mg/mL) among compounds of series 6. Compounds of series 7, namely 7d, 7f, 7g had the lowest minimum inhibitory concentration (MIC). Compound 7f displayed also the lowest cytotoxic effect against fibroblast cell line among series 7 compounds. All tested derivatives displayed lower MIC against Gram-positive bacteria than commercially applied antiseptic, povidone iodine, which MIC value range for tested Gram-positive bacteria was 1.56-6.25 mg/mL.

Clinical Trials of Probiotic Strains in Selected Disease Entities.

Probiotics are live microorganisms which when administered in adequate amounts confer a health benefit on the host. Although their mechanism of action is not clearly explained, it is known that they positively modulate the immune system, which leads to immunity potentiation. A number of studies prove that probiotics strengthen cognitive functions, reduce anxiety, and regulate the lipid metabolism in the human body. Probiotics used in humans are most often of the Lactobacillus and Bifidobacterium species. However, as more research is conducted, new species with beneficial, probiotic properties are being discovered. This paper provides a review of available information about the influence of probiotics on human health. It summarizes the current knowledge on the mechanism of action of probiotics as well as clinical trial results proving their efficacy in allergic, neurodegenerative, and cardiac diseases. This review also discusses the data concerning the safety of probiotics in clinical treatment.

In vitro efficacy of gentamicin released from collagen sponge in eradication of bacterial biofilm preformed on hydroxyapatite surface.

Biofilm-related infections of bones pose a significant therapeutic issue. In this article we present in vitro results of the efficacy of gentamicin released from a collagen sponge carrier against Staphylococcus aureus, Pseudomonas aeruginosa and Klebsiella pneumoniae biofilms preformed on hydroxyapatite surface. The results indicate that high local concentrations of gentamicin released from a sponge eradicate the biofilm formed not only by gentamicin-sensitive strains but, to some extent, also by those that display a resistance pattern in routine diagnostics. The data presented in this paper is of high clinical translational value and may find application in the treatment of bone infections.

The influence of antibiotics and dietary components on gut microbiota.

The gut microbiota acts as a real organ. It exerts important metabolic functions, and regulates the inflammatory response by stimulating the immune system. Gut microbial imbalance (dysbiosis) has been linked to important human diseases and inflammation-related disorders. The symbiotic interactions between resident microorganisms and the gastrointestinal tract significantly contribute to maintaining gut homeostasis. The present review summarizes our knowledge regarding the impact of different antibiotics causing such long-term consequences as decreased microbial diversity, modulation of the Bacteroidetes/Firmicutes ratio, Clostridium difficile overgrowth, and increased expansion of the opportunistic pathogens Salmonella typhimurium, Escherichia spp., and Klebsiella spp. Also, food additives, such as emulsifiers and artificial sweeteners, which are meant to reduce the risk of obesity and diabetes, may actually increase the risk of diseases due to microbial alterations. On the other hand, dietary components such as polyphenols, omega-3 acids or curcumin may positively affect gut microbiota composition.