Carbapenem-Resistant Klebsiella pneumoniae in COVID-19 Era-Challenges and Solutions.

The COVID-19 era brought about new medical challenges, which, together with nosocomial bacterial infections, resulted in an enormous burden for the healthcare system. One of the most alarming nosocomial threats was carbapenem-resistant Klebsiella pneumoniae (CRKP). Monitoring CRKP incidence and antimicrobial resistance globally and locally is vitally important. In a retrospective study, the incidence of CRKP in the pre-COVID-19 period (2017-2019) and the COVID-19 pandemic (2020-2022) was investigated in the Central Military Hospital in Ruzomberok, Slovak Republic. The relative incidence of CRKP significantly increased during the COVID-19 period-by 4.8 times, from 0.18 to 0.76%. At the same time, 47% of CRKP-positive patients also had COVID-19. Twenty-six KPC and sixty-nine NDM-producing isolates were identified. CRKPs isolated in the year 2022 were submitted to whole genome sequencing, and their susceptibility was tested to cefiderocol, ceftazidime-avibactam, imipenem-relebactam and meropenem-vaborbactam, with excellent results. KPC-producing isolates were also highly susceptible to colistin (92%). The NDM isolates revealed lower susceptibility rates, including only 57% colistin susceptibility. ST-307 prevailed in KPC and ST-11 in NDM isolates. Despite the excellent activity of new antimicrobials, rational antibiotic policy must be thoroughly followed, supported by complementary treatments and strict anti-epidemic precautions.

Moving toward Extensively Drug-Resistant: Four-Year Antimicrobial Resistance Trends of Acinetobacter baumannii from the Largest Department of Internal Medicine in Slovakia.

A. baumannii imposes a great burden on medical systems worldwide. Surveillance of trends of antibiotic resistance provides a great deal of information needed for antimicrobial stewardship programmes nationwide. Clinical data from long-term, continuous surveillance on trends of antibiotic resistance of A. baumannii in Slovakia is missing. One hundred and forty-nine samples of A. baumannii were isolated over a period of four years. A panel of 19 antibiotics from seven antibiotic categories were tested for the bacterium's susceptibility. Resistance results were evaluated, and the significance of patterns was estimated using simple linear regression analysis. All isolates were more than 85% resistant to at least 13 out of the 19 tested antibiotics. A significant rise in resistance was recorded for aminoglycosides and imipenem from 2019 to 2022. Colistin and ampicillin-sulbactam have been the only antibiotics maintaining more than 80% efficacy on the bacterium to date. A significant rise in extensively drug-resistant (XDR) strains among carbapenem-resistant (CR) isolates has been recorded. Multidrug-resistance (MDR) among all A. baumannii isolates and XDR among CR strains of the bacterium have risen significantly in the last four years.

High Emergence of Multidrug-Resistant Sequence Type 131 Subclade C2 among Extended-Spectrum ß-Lactamase (ESBL)-Producing Escherichia coli Isolated from the University Hospital Bratislava, Slovakia.

The expansion of sequence type 131 (ST131) extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli (E. coli) represents major worldwide challenges. E. coli strains originating from healthcare facilities (labeled No. 1 and No. 2) of the University Hospital Bratislava (UHB) were analyzed for ST131 emergence, including its (sub)lineages and clonal relatedness. Antimicrobial resistance was determined in most strains. Of a total of 354 E. coli strains, 263 (74.3%) belonged to ST131; of these, 177 (67.3%) were from No. 1. Generally, among 260 ST131 E. coli, clades A/B were confirmed in 20 (7.7%), while clade C was noted in 240 (92.3%) strains; within them, subclades were detected as follows: C0 (17; 7.1%), C1 (3; 1.2%), and C2 (220; 91.7%). Among fifteen randomly selected E. coli strains that were investigated for ST and clonal relatedness, seven STs were identified: eight (53.3%) ST131, two (13.3%) ST73, and one each (6.7%) of ST10, ST12, ST14, ST1193, and ST1196. From No. 1, two ST131 in the first internal clinic and one ST131 from No. 2 in the aftercare department were highly clonally related, suggesting possible epidemiological association. Antimicrobial resistance was as follows: ciprofloxacin 93.8%, ceftazidime 78.4%, meropenem 0%, fosfomycin 2.9% and nitrofurantoin 1.4%. Prevention of ESBL-producing E. coli dissemination, especially for ST131 clade C2, is inevitably necessary for reducing drug resistance and decreasing healthcare-associated infections.

Autovaccines - potent tool against chronic vulvovaginal candidiasis.

OBJECTIVE: Our aim was to determine the effect of immunomodulatory therapy in women with chronic and recurrent vulvovaginal candidiasis (RVVC). BACKGROUND: We present recent highlights in the research into vaginal microbiome and consequences of chronic inflammation such as vulvovaginal candidiasis (VVC). VVC is a widespread vaginal infection primarily caused by Candida albicans. Experience of more than three episodes per year is defined as RVVC. METHODS: The strains were isolated from women suffering from the above infections as for the period of 2017 to 2021 and subsequently used in immunomodulatory treatment. The preparation and administration of autovaccination therapy was performed using standard methodology and procedures cited in the manuscript. RESULTS: In total, autovaccines were produced for 73 patients of whom 30 (41 %) were successfully cured by this treatment, 29 (40 %) experienced a partially successful treatment, and in the remaining 14 (19 %), the autovaccination therapy was ineffective. CONCLUSION: We provide current knowledge about alternative (autovaccine) treatment options for female patients with VVC and RVVC diseases and our experience with the outcomes after autovaccine administration that currently has a promising therapeutic potential (Tab. 2, Ref. 18). Text in PDF www.elis.sk Keywords: autovaccines, chronic infections, vulvovaginal candidiasis, recurrent, Candida albicans.

Next-Generation Sequencing of Carbapenem-Resistant Klebsiella pneumoniae Strains Isolated from Patients Hospitalized in the University Hospital Facilities.

Carbapenem-resistant (CR) Klebsiella pneumoniae represents an urgent worldwide threat. We focused on CR K. pneumoniae in selected facilities of the University Hospital Bratislava (UHB) to investigate sequence types (STs), clonal relatedness, and antimicrobial resistance. Suspected carbapenem-nonsusceptible K. pneumoniae strains were obtained from hospitalized patients. Further examination included carbapenemase confirmation, cgMLST, and quantitative susceptibility testing. Of the total 41 CR K. pneumoniae strains, 26 (63.4%) were determined as ST11 in hospital No. 1; of these ST11, 22 (84.6%) were found in the first internal clinic. Six (14.6%) ST258 and three (7.3%) ST584 occurred in hospital No. 2; the most, i.e., four (66.7%), ST258 were detected in the geriatric clinic. In hospital No. 3, we found two (4.8%) ST584 and one (2.4%) ST258. Of the ST11 set, 24 (92.3%) produced NDM-1. Furthermore, seven (87.5) ST258 and five (100%) ST584 strains generated KPC-2. Antimicrobial resistance was as follows: ertapenem 97.6%, meropenem 63.4%, tigecycline 7.3%, eravacycline 7.3%, and colistin 2.5%. We revealed a presumably epidemiological association of ST11 with transmission, particularly in the first internal clinic of hospital No. 1, while ST258 and ST584 were related to interhospital dissemination between medical facilities No. 2 and No. 3. It is essential to prevent the circulation of these pathogens within and between healthcare facilities.

Susceptibility of Staphylococcus aureus strains to commercial therapeutic phage preparations.

OBJECTIVES: The aim of this study was to determine the susceptibility of Staphylococcus aureus strains to commercial phage preparations. The strains were isolated from clinical patients as well as from nasal mucosa of healthy carriers. BACKGROUND: The elevating number of antibiotic-resistant S. aureus strains present a therapeutic challenge, especially in high-risk patients. One of the promising ways to solve this problem is phage therapy. METHODS: Susceptibility of 111 carrier strains of S. aureus (4 strains were methicillin-resistant; MRSA) and 81 clinical isolates from bloodstream or skin and soft tissue infections (28 were MRSA) to four commercial phage preparations was assessed in vitro by spot assay. The clonality of S. aureus strains was determined by spa typing. RESULTS: Spa typing revealed 97 distinct spa types. A proportion of 73-80 % of the tested S. aureus strains were revealed to have in vitro phage susceptibility, depending on the clonal affiliation of the strains and phage preparation tested. The susceptibility to phage preparations was significantly higher in MRSA strains (p < 0.001). CONCLUSIONS: In vitro results indicate a promising therapeutic potential of the tested commercial anti-staphylococcal phage preparations. They could be applied to a broad spectrum of bacterial clones, and have an excellent activity especially against MRSA strains (Tab. 2, Fig. 2, Ref. 43).

First report of nosocomial outbreak of vancomycin-resistant Enterococcus faecium infection among COVID-19 patients hospitalized in a non-intensive care unit ward in Slovakia.

BACKGROUND: Enterococcus species account for most of the human enterococcal HAI and multidrug-resistant infections and have become a major threat to modern public health. We examine the rise in the number of vancomycin resistant E. faecium blood stream and urinary tract infections in a COVID-19 department during an epidemiologic outbreak investigation to detect and eliminate nosocomial clusters of the bacteria. METHODS:  Strain identification was performed by classical isolation and biochemical and cultivation methods. Antibiotic testing results were interpreted according to European committee on antimicrobial susceptibility testing (EUCAST) guidelines. Six isolated samples underwent the whole genome sequencing (WGS) during the outbreak investigation. Isolate relatedness was determined using the core genome multi-locus sequence typing. RESULTS:  WGS revealed two genotypically distinct VRE clusters, one of which had genetically closely related patients and environmental isolates. The cluster was terminated by enhanced infection control strategies. CONCLUSIONS:  This study provides the first description of an outbreak caused by vanB-ST117 and vanA-ST17 E. faecium strains among COVID-19 patients in Slovakia. This study can help to raise the awareness about the need for strict adherence to infection control measures and the implementation of rational antimicrobial stewardship as a routine part of COVID-19 management (Tab. 3, Fig. 3, Ref. 27). Text in PDF www.elis.sk Keywords: vancomycin-resistant Enterococcus faecium, antibiotic resistant, COVID-19, SARS-CoV-2, bacterial outbreak, healthcare-associated infection.

Phascinating Phages.

Treatment of infections caused by bacteria has become more complex due to the increasing number of bacterial strains that are resistant to conventional antimicrobial therapy. A highly promising alternative appears to be bacteriophage (phage) therapy, in which natural predators of bacteria, bacteriophages, play a role. Although these viruses were first discovered in 1917, the development of phage therapy was impacted by the discovery of antibiotics, which spread more quickly and effectively in medical practice. Despite this, phage therapy has a long history in Eastern Europe; however, Western countries are currently striving to reintroduce phage therapy as a tool in the fight against diseases caused by drug-resistant bacteria. This review describes phage biology, bacterial and phage competition mechanisms, and the benefits and drawbacks of phage therapy. The results of various laboratory experiments, and clinical cases where phage therapy was administered, are described.

Complex Analysis of Vanillin and Syringic Acid as Natural Antimicrobial Agents against Staphylococcus epidermidis Biofilms.

The presence of Staphylococcus epidermidis biofilms on medical devices is a major cause of nosocomial diseases and infections. Extensive research is directed at inhibiting the formation and maturation of such biofilms. Natural plant-derived phenolic compounds have promising antimicrobial effects against drug-resistant bacteria. The anti-biofilm activity of two selected phenolic compounds (vanillin and syringic acid) was tested against three biofilm-forming methicillin-resistant S. epidermidis strains with different genotypes. Resazurin assay combining crystal violet staining and confocal microscopy was used for biofilm and extracellular polymer substance (EPS) inhibition tests. Effects on EPS compounds such as proteins, extracellular DNA, and polysaccharides were also examined. Combined with quantitative real-time PCR of selected agr quorum-sensing systems and biofilm genetic determinants, our complex analysis of vanillin and syringic acid showed similar biofilm and EPS inhibition effects on S. epidermidis strains, reducing biofilm formation up to 80% and EPS up to 55%, depending on the genotype of the tested strain. Natural antimicrobial agents are thus potentially useful inhibitors of biofilms.

Characterization of Anti-Bacterial Effect of the Two New Phages against Uropathogenic Escherichia coli.

Urinary tract infections (UTIs) are among the events that most frequently need medical intervention. Uropathogenic Escherichia coli are frequently their causative agents and the infections are sometimes complicated by the presence of polyresistant nosocomial strains. Phage therapy is a tool that has good prospects for the treatment of these infections. In the present study, we isolated and characterized two bacteriophages with broad host specificity against a panel of local uropathogenic E. coli strains and combined them into a phage cocktail. According to genome sequencing, these phages were closely related and belonged to the Tequatrovirus genus. The newly isolated phages showed very good activity on a panel of local clinical E. coli strains from urinary tract infections. In the form of a two-phage cocktail, they were active on E. coli strains belonging to phylogroups B2 and D, with relatively lower activity in B1 and no response in phylogroup A. Our study is a preliminary step toward the establishment of a national phage bank containing local, well-characterized phages with therapeutic potential for patients in Slovakia.

Chronic hepatitis C virus infection in the Czech Republic and Slovakia: an analysis of patient and virus characteristics.

OBJECTIVES: The MOSAIC study gathered data on chronic hepatitis C virus (HCV) infection and its treatment in various countries worldwide. Here we summarise patient and HCV characteristics in the Czech Republic and Slovakia. METHODS: MOSAIC was an observational study that included patients with chronic HCV infection untreated at the time of enrolment. Study collected and descriptively analysed patient demographics, disease stage and viral characteristics. Data were collected between February 2014 to October 2014. RESULTS: Among 220 patients enrolled, 51.4% were treatment-naïve. The most prevalent HCV genotype was G1 (78.4%), followed by G3 (19.7%). Higher prevalence of G1 was found in treatment-experienced patients (94.3%) compared to treatment-naïve (63.4%). Most participants (67.7%) presented viral RNA load of ≥ 800,000 IU/mL. Liver cirrhosis was reported in 24.5% of patients. Higher HCV RNA load and duration of HCV infection correlated with the degree of liver fibrosis. Anti-HCV interferon-based treatments were initiated in 88.2% of participants. CONCLUSIONS: The study confirmed significant changes in the HCV genotypes partition with G3 genotype rapidly increasing in both countries, with possible impact on the WHO eradication initiative and treatment selection.

Glucose Metabolism Changes in Patients with Chronic Hepatitis C Treated with Direct Acting Antivirals.

Background and Aims: Chronic hepatitis C is a systemic disease and type 2 diabetes mellitus (T2DM) belongs to more common extrahepatic. The aim of this study was to (i) explore the prevalence of impaired fasting glucose (IFG) and T2DM in patients with chronic hepatitis C, (ii) explore the effect of direct acting antivirals (DAA) treatment on the glycemia, and (iii) explore the factors that modulate the effect of DAA treatment on glycemia in patients with chronic hepatitis C. Methods: We performed a longitudinal retrospective observational study focused on the patients undergoing DAA treatment of chronic hepatitis C. Data about glycemia, history of diabetes, hepatitis C virus, treatment, and liver status, including elastography, were obtained at baseline (before treatment start), at the end of treatment and 12 weeks after the end of treatment. Patients were treated with various regimens of direct acting antivirals. Results: We included 370 patients; 45.9% had F4 fibrosis. At baseline, the prevalence of T2DM increased with the degree of fibrosis (F0-F2 14.4%, F3 21.3%, and F4 31.8%, p=0.004). Fasting glycemia also increased with the degree of fibrosis (F0-F2 5.75±0.18 F3 5.84±0.17, and F4 6.69±0.2 mmol/L, p=0.001). We saw significant decrease of glycemia after treatment in all patients, but patients without T2DM or IFG from 6.21±0.12 to 6.08±0.15 mmol/L (p=0.002). The decrease was also visible in treatment experienced patients and patients with Child-Pugh A cirrhosis. Conclusion: We confirmed that the prevalence of either T2DM or IFG increases in chronic hepatitis C patients with the degree of fibrosis. The predictive factors for T2DM were, besides F4, fibrosis also higher age and BMI. Significant decrease of fasting glycemia after the DAA treatment was observed in the whole cohort and in subgroups of patients with T2DM, IFG, cirrhotic, and treatment experienced patients.

Association of interleukin-10 gene promoter polymorphisms with susceptibility to acute pyelonephritis in children.

Interleukin-10 (IL-10) is a potent inhibitor of leukocyte chemotaxis, bacterial killing in phagocytes and synthesis of pro-inflammatory cytokines and chemokines, and recent studies have suggested an important role for this immunoregulatory cytokine in the pathogenesis of urinary tract infections (UTIs). Therefore, the gene encoding IL-10 (IL10) is an attractive candidate for association studies attempting to identify susceptibility genes conferring risk of UTIs. In this case-control study, we aimed to investigate the association of single nucleotide polymorphisms (SNPs) in the promoter region of IL10 with acute pyelonephritis in the Slovak population. Polymerase chain reaction with sequence-specific primers was used to analyse IL10 -1082A/G (rs1800896), -819C/T (rs1800871) and -592C/A (rs1800872) SNPs in 147 children with acute pyelonephritis and 215 healthy controls. Comparison of patients with healthy controls using the logistic regression analysis revealed significantly increased risk of developing recurrent attacks of acute pyelonephritis for -1082 G allele in a dominant genetic model GG (GG + AG vs. AA, P = 0.019, odds ratio (OR) = 2.26). A similar tendency was also found when the recurrent acute pyelonephritis subgroup was compared to episodic pyelonephritis cases (GG + AG vs. AA, P = 0.009, OR = 3.38). In conclusion, our results suggest that IL10 -1082 A/G SNP is a susceptibility factor for development of recurrent attacks of acute pyelonephritis.

Phage therapy of Cronobacter-induced urinary tract infection in mice.

BACKGROUND: Cronobacter spp. is an opportunistic pathogen causing rare but dangerous cases of meningitis, sepsis and urinary tract infection. Phage therapy overcomes antibiotic resistance and represents an alternative approach to standard antimicrobial treatment. There are no published studies on the use of phages against Cronobacter spp. in vivo. The aim of our study was to prove the effects of isolated Cronobacter-specific phages on renal colonization in a model of urinary tract infection in mice. MATERIAL/METHODS: Urinary tract infection was induced by transurethral application of Cronobacter turicensis (1011 CFU/ml). Simultaneously, isolated Cronobacter-specific phages were administered intraperitoneally (1011 PFU/ml). After 24 hours, kidneys and bladder were collected and used for cultivation and analysis of gene expression and oxidative stress markers. RESULTS: Phage therapy reduced the number of Cronobacter colonies in the kidney by 70%. Higher levels of malondialdehyde were reduced by phage therapy without affecting the antioxidant status. The expression of pro-inflammatory cytokines tumor necrosis factor-alpha and monocyte chemoattractant protein-1 increased by the infection and was attenuated by phage therapy. CONCLUSIONS: Phage therapy proved effective in the prevention of ascending renal infection in a murine model of urinary tract infection. Long-term effects and safety of the treatment are currently unknown. Further studies should test phage therapy in other Cronobacter infection models.

Bactofection with toll-like receptor 4 in a murine model of urinary tract infection.

The role of innate immunity in the prevention of urinary tract infection is well-documented. Toll-like receptor 4 (TLR4) is a major determinant of innate immune response. In an animal model of urinary tract infection, bactofection-mediated gene transfer of TLR4 was tested in a preventive approach. Bactofection with TLR4 reduced the colonization with uropathogenic Escherichia coli by 91% in the kidney and by 41% in the bladder. Reduced colonization was associated with lower oxidative stress and expression of monocyte chemoattractant protein-1 and myeloperoxidase in the kidney. Bactofection with TLR4 was successful in the prevention of ascending pyelonephritis. Further studies should focus on long-term effects, the dose response and the potential therapeutic use in models of chronic urinary tract infection.

[Selected mechanisms of resistance to antimicrobial drugs in Pseudomonas aeruginosa and their detection]. / Vybrané mechanizmy rezistencie na antimikrobiálne látky u Pseudomonas aeruginosa a ich detekcia.

By now most clinically significant bacterial species have resistance mechanisms against almost all accessible antibiotics. One of the most significant nosocomial pathogens is Pseudomonas aeruginosa. Its antibiotic resistance directly correlates with higher morbidity, mortality and longer hospital stay. An essential condition for preserving the antibiotic efficacy against this pathogen is the exploration of its resistance mechanisms. To protect itself against the action of antibacterials, P. aeruginosa uses the cellular wall, which prevents the drug from entering the bacterium, as also an active efflux from the cell. It is also capable of modifying the molecule of the antimicrobial. The bacterium can effectively combine all these mechanisms and the result are multiresistant bacterial clones. The paper describes some diagnostic methods that reveal the resistance mechanisms of the species Pseudomonas aeruginosa.