Ultrasensitive voltammetric detection of SARS-CoV-2 in clinical samples.

In every pandemic, it is critical to test as many people as possible and keep track of the number of new cases of infection. Therefore, there is a need for novel, fast and unambiguous testing methods. In this study, we designed a sandwich-type voltammetric immunosensor based on unlabeled- and labeled with a redox probe antibodies against virus spike protein for fast and ultrasensitive detection of SARS-CoV-2. The process of the preparation of the sensor layer included chemisorption of cysteamine layer and covalent anchoring of antibody specific for the S1 subunit of the S protein. The source of the voltametric signal was the antibody labeled with the redox probe, which was introduced onto biosensor surface only after the recognition of the virus. This easy-to-handle immunosensor was characterized by a wide analytical range (2.0·10-7 to 0.20 mg·L-1) and low detection limit (8.0·10-8 mg·L-1 ≡ 0.08 pg·mL-1 ≡ 4 virions·µL-1). The utility of the designed device was also evidenced by the detection of SARS-CoV-2 in the clinical samples. Moreover, the main advantage and a huge novelty of the developed device, compared to those already existing, is the moment of generating the analytical signal of the redox probe that appears only after the virus recognition. Thus, our diagnostic innovation may considerably contribute to controlling the COVID-19 pandemic. The as-developed immunosensor may well offer a novel alternative approach for viral detection that could complement or even replace the existing methods.

Children were less frequently infected with SARS-CoV-2 than adults during 2020 COVID-19 pandemic in Warsaw, Poland.

Clinical data suggest that during the current COVID-19 pandemic, children are less prone than adults to SARS-CoV-2 infection. Our purpose was to determine the frequency of SARS-CoV-2 in children vs. adults during the 2020 pandemic in Warsaw, Poland, and to investigate whether RSV and/or influenza A/B infections were associated with SARS-CoV-2 infections. We present results of RT-PCR tests for SARS-CoV-2 performed in Warsaw, Poland. Some of the pediatric subjects were also PCR-tested for RSV, and A and B influenza. We compared the test results from the four groups of symptomatic and asymptomatic subjects: 459 symptomatic pediatric patients (children 0-18 years old), 1774 symptomatic adults, 445 asymptomatic children, and 239 asymptomatic adults. 3.26% (15/459) of symptomatic pediatric patients were positive for SARS-CoV-2 in contrast to 5.58% (99/1774) of symptomatic adults (p = 0.0448). There were no SARS-CoV-2 positive cases in the group of asymptomatic children (0/445) and two positive cases in the group of asymptomatic adults (2/239), i.e., 0.83%. In the group of symptomatic pediatric patients, 17.14% (6/35) (p = 0.0002) were positive for RSV, 8.16% (4/49) were positive for influenza A, and 2.04% (1/49), thus 10.20% (5/49) (p = 0.0176) for influenza A/B. Children were less prone to SARS-CoV-2 infection than the adults during the COVID-19 pandemic in Warsaw. Higher percentage of symptomatic children was infected with RSV or influenza A/B than with SARS-CoV-2. This suggests a necessity for the testing for all these viruses for an early identification and isolation of SARS-CoV-2-positive patients for an ensuing 2020 autumn return of COVID-19.

The clinical course of gastroenteritis due to nosocomial and community acquired norovirus infections in immunocompromised and immunocompetent children - single center experience.

BACKGROUND: After the introduction of rotavirus vaccines into immunization schedules, noroviruses account for the majority of acute gastrointestinal infections. The aim of the study was to assess the clinical presentation in immunocompromised and immunocompetent children with hospital- and community-acquired norovirus gastroenteritis. MATERIAL AND METHODS: We retrospectively reviewed clinical records of children with noroviral gastroenteritis, hospitalized in the Pediatric Hospital, Medical University of Warsaw, between 2015 and 2018. Acute gastrointestinal tract symptoms and confirmed etiology of noroviral infection were inclusion criteria. The analysis was performed in the subgroups of immunocompetent and immunocompromised patients, during community-acquired and nosocomial infections. RESULTS: A total of 57 children with median age 1.5 year (IQR: 0.7-4.0) were recruited. The majority of patients were immunocompetent (87.7%), and nosocomial infections were predominant (56.1%). Gastrointestinal symptoms included nausea, vomiting and diarrhoea (in approximately 85%), while systemic manifestations such as fever and malaise where observed in only ». Routine laboratory tests were normal in most of the patients. An analysis in the subgroups revealed statistically significant differences in blood pH and serum electrolyte levels - acidosis and electrolyte disturbances were statistically significantly more common in immunocompromised vs immunocompetent patients (p<0.05). CONCLUSIONS: More frequently the clinical presentation includes gastrointestinal symptoms with no differences between immunocompromised and immunocompetent hosts. The median laboratory values were normal in generally healthy children; disturbances were observed only in children with immunodeficiencies. Therefore, prophylactic measures are of particular importance in the latter group, which is especially sensitive to severe and nosocomial infections.

Bartonella and Rickettsia Infections in Haematophagous Spinturnix myoti Mites (Acari: Mesostigmata) and their Bat Host, Myotis myotis (Yangochiroptera: Vespertilionidae), from Poland.

Hematophagous Spinturnix myoti mites and their host, the greater mouse-eared bat (Myotis myotis), were tested for the presence of Bartonella spp., Rickettsia spp., and Anaplasma phagocytophilum. In total, Bartonella spp. DNA was amplified in 28% of 134 mite pools and in 25% of 59 bats tested by PCR targeting a fragment of citrate synthase gltA gen. Adult mites were at least threefold more frequently infected compared to immature stages. The overall infection prevalence among mite pools from cave-dwelling bats was higher than for those collected from attic shelters. Three distinct genotypes were detected. The most prevalent genotype in mites and bats matched closely with Candidatus Bartonella hemsundetiensis identified in bats from Finland and was relatively distant from bat-borne Bartonella strains described in the UK and France. Importantly, most sequences were close to those reported in forest workers from Poland. The presence of identical genotype among S. myoti samples and M. myotis bats suggests that bartonellae can be shared between mites and their bat hosts. In this case, wing mites could serve as vectors, whereas their hosts as reservoirs. One blood sample was positive by PCR for the msp2 gene of A. phagocytophilum. Two mite pools yielded Rickettsia spp. DNA. Both sequences were distinct from any known species but can be classified as spotted fever group Rickettsia spp. Our findings expanded our knowledge on the role of spinturnicid mites in the ecology and epidemiology of bacterial infections associated with vespertilionid bats, especially regarding the genus Bartonella.

Correction to: Bartonella and Rickettsia Infections in Haematophagous Spinturnix myoti Mites (Acari: Mesostigmata) and their Bat Host, Myotis myotis (Yangochiroptera: Vespertilionidae), from Poland.

The original version of this article published online (27 August 2018) unfortunately contained a mistake regarding an affiliation of Dr. Edyta Podsiadly, one of the authors.

Probable Interspecies Transfer of the bla(VIM-4) Gene between Enterobacter cloacae and Klebsiella pneumoniae in a Single Infant Patient.

We report the interspecies transfer of the bla(VLM-4) gene in MBL-producing Enterobacter cloacae and Klebsiella pneumoniae isolates from a newborn patient who had received meropenem therapy. We show evidence that gene bla(VIM-4) was transmitted as a part of the class-1 integron on a ca. -90 kb conjugative plasmid. High homology of nucleotide sequence was observed between the integron found in VIM-4 producing E. cloacae and K. pneumoniae strains tested and class-1 integrons previously reporteded in Pseudomonas aeruginosa from Hungary and Poland. This finding may suggest P. aeruginosa as a potential source of acquired VIM-4 in Enterobacteriaceae.

[Presence of lactoferrin in faeces as the indicator of Clostridium difficile in pediatric patients]. / Obecnosc laktoferyny w kale jako wskaznik zakazenia Clostridium difficile u dzieci.

INTRODUCTION: Number of infection caused by Clostridium difficile in hospitalized children is increasing. Even though children unlike adults seldom develop complications after being ill, cases of persistent diarrhoea triggered by this pathogen and mortality from this origin have been reported. At present the important problem constitute differentiation between the colonization and infection limiting the proper diagnosis of C. difficile infections (CDI) in this age group. The aim of this study was to evaluate the presence of lactoferrin in faeces as the inflammatory marker confirming C. difficile infections in children. METHODS: Seventy seven samples of faeces where examined. Among them in 55 toxin A/B or C. difficile toxinogenic strain and in 15 nontoxinogenic C. difficile had been detected, 7 were collected from healthy children. Stool samples were tested with the use of method routinely applied in laboratory: automatic VIDAS C. difficile Toxin A&B test (bioMerieux, France), culture and GDH test. Lactoferrin in stool has been identified with ELISA IBD-SCAN test (Techlab, Blacksburg, VA). The CRP protein was detected with VITROS 5600. RESULTS: Among 55 children with CDI lactoferrin was detected in 45,5% (25/55) of them. In 30 (54,5%) CDI patients the inflammatory biomarker was not identified. In 15 persons with nontoxinogenic strain cultured, one child had lactoferrin present. CDI was detected most frequently (51%) in 6-11 years old children. The increase of CDI cases was observed in period 2013-2014. Neither differences in frequency of raised CRP level in examined groups of children nor correlation between presence of lactoferrin and CRP was observed. CONCLUSIONS: Lactoferrin an intestinal inflammatory biomarker may be a useful tool in distinguishing between C. difficile infection and colonization. More studies including clinical observations are needed.

Epidemiological Characteristics of Shiga Toxin-Producing Escherichia coli Responsible for Infections in the Polish Pediatric Population.

Shiga toxin-producing Escherichia coli (STEC) are zoonotic pathogens causing hemorrhagic colitis and hemolytic uremic syndrome (HUS) in children and the elderly. Stool samples were collected from 180 children hospitalized in five pediatric centers in Poland in 2018-2022. Direct stx1/stx2 gene detection by PCR in feces and E. coli isolates was performed. Antibiotic susceptibility was tested according to EUCAST v.12. Randomly selected isolates were serotyped with O157 antiserum and genotyped by pulsed-field gel electrophoresis (PFGE). A total of 44 E. coli isolates were confirmed as STEC by PCR. Among them, 84.4% were positive for stx2, and equally 6,8% for only stx1 and both stx1 and stx2 genes. The stx1 gene was also found in one Citrobacter freundii isolate. E. coli serotype O157 was present in 97.6% of the isolates. STEC infections most often occurred between June-October with a peak in July and August (51%). The highest, 77.8% of STEC isolates were found in the 1-5 years old group. No extended-spectrum ß-lactamases (ESBL) were found. Resistance only to amoxicillin/clavulanic acid (24.4%), piperacillin/tazobactam (3%), cefotaxime (6%), gentamicin (6%), ciprofloxacin (3%), azithromycin (3%), trimethoprim/sulfamethoxazole (24,2%) was detected. PFGE analysis showed 18 PFGE types with no clonal distribution. Eight isolates with A, B, and C PFGE types showed genetic relatedness in the type with no detection of transmission way of distribution. STEC strains pose a serious threat to human health, therefore demographic and epidemiological characteristics are crucial for their surveillance.

Analysis of Etiology of Community-Acquired and Nosocomial Urinary Tract Infections and Antibiotic Resistance of Isolated Strains: Results of a 3-Year Surveillance (2020-2022) at the Pediatric Teaching Hospital in Warsaw.

Urinary tract infections (UTIs) remain the most common infections diagnosed in outpatients and hospitalized patients. This study was designed to determine the patterns of antibiotic resistance and the prevalence of uropathogens causing UTIs in pediatric patients hospitalized between 1 January 2020 and 31 December 2022 at Teaching Hospital in Warsaw. The most frequent species isolated from urine samples were E. coli (64.5%), Klebsiella spp. (11.6%), and Enterococcus spp. (6.1%). UTIs caused by Enterobacter spp., Enterococcus spp., and Klebsiella spp. were significantly more common in children younger than three months of age than in children older than three months (p < 0.001). Trimethoprim and trimethoprim-sulfamethoxazole were the least active compounds against Enterobacterales with the resistance of E. coli, Klebsiella spp., P. mirabilis, and Enterobacter spp. in the range of 26.7/25.2%, 48.4/40.4%, 51.1/40.4%, and 15.8/13.2% respectively. Ampicillin was also found to have resistance rates for E. coli of 54.9% and P. mirabilis of 44.7%. Cefalexin and cefuroxime were highly active towards Enterobacterales except for Klebsiella spp., in which the resistance level reached 40%. Regarding third- and fourth- generation cephalosporins, resistance in E. coli and P. mirabilis was observed in approximately 2-10% of the isolates, but in Klebsiella spp. and Enterobacter spp. ranged over 30%. The resistance of Enterobacterales to carbapenems, nitrofurantoin, and fosfomycin was below 1%. The quinolones resistance was very high for Klebsiella spp. (31.1%) and P. mirabilis (29.8%) and three times lower for E. coli (11.9%), P. aeruginosa (9.3%), Enterobacter spp. (2.6%), and E. faecalis (4.6%). Resistance to multiple antibiotic classes was identified in 396 Enterobacterales strains, 394 of which were multi-drug resistant (MDR) and 2 were extensive drug-resistant (XDR). In the case of E. coli, 30% of isolates were MDR, with the proportion of strains having this exact resistance pattern similar in all of the analyzed years; no E. coli XDR strains were isolated. The number of Klebsiella spp. MDR strains was much higher in 2022 (60%) than in 2021 (47.5%). In the analyzed time, only one strain of K. pneumonia XDR, producing New Delhi metallo-ß-lactamase, was isolated. Monitoring infection trends is essential to improve control and limit the rise of bacterial resistance.

Epidemiology and Risk Factors of UTIs in Children-A Single-Center Observation.

Urinary tract infections (UTIs) are one of childhood's most common bacterial infections. The study aimed to determine the clinical symptoms, laboratory tests, risk factors, and etiology of different UTIs in children admitted to pediatric hospitals for three years. Methods: Patients with positive urine cultures diagnosed with acute pyelonephritis (APN) or cystitis (CYS) were analyzed for clinical symptoms, laboratory tests, risk factors, and etiology, depending on their age and sex. Results: We studied 948 children with UTIs (531 girls and 417 boys), with a median age of 12 (IQR 5−48 months). A total of 789 children had clinical symptoms; the main symptom was fever (63.4% of patients). Specific symptoms of UTIs were presented only in 16.3% of patients. Children with APN had shown significantly more frequent loss of appetite, vomiting, lethargy, seizures, and less frequent dysuria and haematuria than children with CYS. We found significantly higher median WBC, CRP, and leukocyturia in children with APN than with CYS. The risk factors of UTIs were presented in 46.6% of patients, of which 35.6% were children with APN and 61.7% with CYS. The main risk factor was CAKUT, more frequently diagnosed in children with CYS than APN, mainly in children <2 years. The most commonly isolated bacteria were Escherichia coli (74%). There was a higher percentage of urine samples with E. coli in girls than in boys. Other bacteria found were Klebsiella species, Pseudomonas aeruginosa, Proteus mirabilis, and Enterococcus species. Conclusions: Patients with APN were younger and had higher inflammatory markers. Often, fever is the only symptom of UTI in children, and other clinical signs are usually non-specific. The most common UTI etiology is E. coli, regardless of the clinical presentation and risk factors.

Monte Carlo Regression for Evaluating Children's Role in the Pandemic Spread on the Example of Delta COVID-19 Wave.

BACKGROUND: The children's role in transmitting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the familial settings is uncertain. We aimed to assess how often children were the index cases transmitting SARS-CoV-2 into their households during the Delta wave, and to identify risk factors of children being the index case. METHODS: In this prospective survey study, we collected information regarding household members of SARS-CoV-2-positive children tested in a single tertiary hospital. Some patients were tested with polymerase chain reaction and those samples were typed and classified as Delta or non-Delta variant. We have used the Monte Carlo approach to assess predictors of children being the index case in the household. RESULTS: We surveyed 629 families and 515 of them fulfilled inclusion criteria. The child was the index case in 359 (69.71%) households. Attending childcare facilities in all age groups was positively associated with being the index case in the household [nursery, estimate = 1.456, 95% confidence interval (CI): 1.456-1.457, P < 0.001; kindergarten, estimate = 0.899, 95% CI: 0.898-0.900, P = 0.003; school, estimate = 1.23, 95% CI: 1.229-1.231, P = 0.001]. The same association was present in the subgroup of the families with the predominant Delta variant, but not in the subgroup with the predominant non-Delta variant. CONCLUSIONS: Attending childcare and educational facilities might be a significant predictor of a child being the SARS-CoV-2 index case in their household. Children's role in driving the SARS-CoV-2 pandemic changes in consecutive waves. The Monte Carlo approach can be applied to assess risk factors of infectious agents' spread in future epidemics.

Ukrainian War Refugee Children With Particularly Severe Viral Infections: A Case Series Report.

The humanitarian crisis in Ukraine in 2022 led to a massive migration of refugees to Poland. Immigrant children, living in overcrowded humanitarian hubs, were exposed to multiple stressful factors likely affecting their immune systems. This case series study aimed to describe a particularly severe course of common viral infections, in Ukrainian refugee children. We present 2 case series of Ukrainian refugee children: 5 hospitalized due to either adenovirus (AdV) and 8 with rotavirus (RV) infection, admitted within 3 months in each case series, recruited retrospectively. Most patients lived in humanitarian hubs and were neglected on admission (dehydrated, with poor hygiene and anxious). All RV infection cases had symptoms of severe gastroenteritis requiring intravenous rehydration. Metabolic acidosis was present in 6 children, and hypoglycemia in 4 participants. None of them were vaccinated against RV. All children with AdV infection had prolonged fever, dyspnea requiring oxygen therapy and hyperinflammation. In 2 AdV infection cases with no clinical improvement and increasing inflammatory markers, intravenous immunoglobulins and glucocorticosteroids were used. The combination of stressful factors and living in overcrowded hubs during the high prevalence of viral infections led to a particularly severe course of viral infections in Ukrainian refugee children.

Susceptibility of Polish Bartonella henselae strains.

Due to the fastidious nature of B. henselae and the limited number of available isolates worldwide, there are few data on its in vitro susceptibility to antibiotics. We determined the minimal inhibitory concentrations (MIC) of ten antimicrobial agents against 11 feline isolates of B. henselae by Etest method. The lowest MICs were obtained for rifampicin < or = 0.002 mg/L. MICs of all isolates were < 0.016 mg/L for ampicilin, amoxicillin/clavulanic acid, tetracycline and ranged from 0.016 to 0.032 mg/L for azithromycin. The MICs for two tested fluoroquinolones: ciprofloxacin and levofloxacin ranged from 0.016 to 0.125 mg/L. The highest MICs were obtained for gentamicin ranging from 0.025 to 2.0 mg/L. Sulphonamide resistance genes sul 1, sul 2, sul 3 were not found in any of the tested isolates. Etest methodology seems to be a reliable method for determination of B. henselae susceptibility, however standardization is strongly desired.

Infectious Etiology of Vomiting in Children With Presumed Acute Gastroenteritis in the Absence of Diarrhea: Protocol for a Cohort Study.

NCT05270291, https://clinicaltrials.gov/ct2/show/. Objectives: In children with acute gastroenteritis (AGE), vomiting often precedes diarrhea. To establish the diagnosis of AGE, enteropathogen detection typically relies on diarrheal stool samples. However, testing requires sufficient stool sample, which may not be easily available. Recent studies suggest that in children presenting to emergency departments with presumed AGE with isolated vomiting, an enteropathogen can be identified using rectal swabs and molecular diagnostic tests. The rate of enteropathogen detection in children with isolated vomiting due to AGE may differ in various populations. Using rectal swabs and molecular diagnostic tests, we plan to assess the proportion of children with isolated vomiting with presumed AGE in whom an enteropathogen can be identified. Methods: This will be a cohort study conducted in the emergency department(s) of one or more pediatric hospital(s) in Poland. Children younger than 5 years with the presence of ≥3 episodes of vomiting due to presumed AGE, lasting no longer than 7 days before enrollment, will be recruited. The primary outcome will be the proportion of children with isolated vomiting in whom an enteropathogen is detected. In all eligible participants, rectal swabs will be taken to perform molecular testing for detection of typical viral and bacterial enteropathogens. All children will be followed-up at 14 days after the initial contact to classify them into one of three groups (i.e., vomiting only, vomiting and diarrhea, and diarrhea only).

Streptococcus intermedius Acute Meningitis in an Immunocompetent Child.

Streptococcus intermedius is a commensal bacterium increasingly associated with brain abscesses. It can infect the brain through a direct extension of an inflammatory process or a hematogenous route. Meningitis due to this pathogen is sporadic and has not been described in children before. We report a case of an immunocompetent 6-year-old boy who developed bacterial meningitis due to S. intermedius .

The Impact of the COVID-19 Pandemic Lockdown on Pediatric Infections-A Single-Center Retrospective Study.

Since the SARS-CoV-2 outbreak, many countries have introduced measures to limit the transmission. The data based on ICD-10 codes of lower respiratory tract infections and microbiological analysis of respiratory and gastrointestinal infections were collected. The retrospective five-year analysis of the medical records revealed a substantial decrease in respiratory tract infections during the pandemic year (from April 2020 to March 2021). We noted an 81% decline in the LRTI-associated hospital admissions based on the ICD-10 analysis (from a mean of 1170 admissions per year in the previous four years to 225 admissions between April 2020 through March 2021). According to microbiological analysis, there were 100%, 99%, 87%, and 47% drops in influenza virus, respiratory syncytial virus, rotavirus, and norovirus cases reported respectively during the pandemic season until April 2021 in comparison to pre-pandemic years. However, the prevalence of gastrointestinal bacterial infections was stable. Moreover, in August 2021, an unexpected rise in RSV-positive cases was observed. The measures applied during the COVID-19 pandemic turned out to be effective but also had a substantial contribution to the so-far stable epidemiological situation of seasonal infections.

Fresh Versus Frozen Stool for Fecal Microbiota Transplantation-Assessment by Multimethod Approach Combining Culturing, Flow Cytometry, and Next-Generation Sequencing.

The objective of this work was to compare the quality of FMT preparations made from fresh feces with those made from feces frozen at -30°C without any pre-processing or cryopreservation additives. The research hypothesis was that such preservation protocol (frozen whole stool, then thawed and processed) is equipotent to classical fresh FMT preparation. For that, three complementary methods were applied, including: (i) culturing in aerobic and anaerobic conditions, (ii) measuring viability by flow cytometry, and (iii) next-generation sequencing. Flow cytometry with cell staining showed that the applied freezing protocol causes significant changes in all of the observed bacterial fractions. Alive cell counts dropped four times, from around 70% to 15%, while the other two fractions, dead and unknown cell counts quadrupled and doubled, with the unknown fraction becoming the dominant one, with an average contribution of 57.47% per sample. It will be very interesting to uncover what this unknown fraction is (e.g., bacterial spores), as this may change our conclusions (if these are spores, the viability could be even higher after freezing). Freezing had a huge impact on the structure of cultivable bacterial communities. The biggest drop after freezing in the number of cultivable species was observed for Actinobacteria and Bacilli. In most cases, selected biodiversity indices were slightly lower for frozen samples. PCoA visualization built using weighted UniFrac index showed no donor-wise clusters, but a clear split between fresh and frozen samples. This split can be in part attributed to the changes in the relative abundance of Bacteroidales and Clostridiales orders. Our results clearly show that whole stool freezing without any cryoprotectants has a great impact on the cultivability and biodiversity of the bacterial community, and possibly also on the viability of bacterial cells.

[Fenotypic and genotypic characterization of Shiga toxin-producing Escherichia coli O111 strain isolated from patient with hemolytic-uremic syndrome]. / Charakterystyka fenotypowych i genotypowych wlasciwosci werotoksycznego szczepu E. coli O111 wyizolowanego od pacjenta z zespolem hemolityczno--mocznicowym.

Shiga toxin-producing E. coli O157 and non-O157 are important emergance pathogens that can cause diarrhea and hemorrhagic colitis with life-threatening complications, such as hemolytic uremic syndrome (HUS). A few cases of EHEC infections are documented per year in Poland. Among them only one patient with EHEC O157 infection developed HUS. We characterized the first VTEC non-O157 strain isolated from child with HUS in Poland. The VTEC O111 strain produced Stx2 which was cytotoxic for Vero cell. Using DNA microarray analysis we have found set of virulence genes in VTEC O111 strain as: stx2A, stx2B, ehly, eae, tir tccP espA, espJ, cif nleA, nleB, lpfA, iha, efa1, cba. The strain was fenotypic resistant to streptomycin, tetracyclin and sulphonamides (strA, tetA, sul2 genes were detected).

Decontamination of microbiologically contaminated abiotic porous surfaces in an oral surgery clinic using vaporised hydrogen peroxide (VHP).

PURPOSE: The aims of the study were to identify microorganisms, including those in the VBNC state, inhabiting porous surfaces in oral surgery offices and to assess the biocidal effectiveness and impact of 300 ppm vaporised hydrogen peroxide (VHP) for 20 min on decontaminated materials. METHODS: From the surfaces of textured armrests of dental chairs, pinewood doors and window frames and cotton medical aprons, 30 swabs were taken with moistened sponges. The identification of isolated microorganisms was performed using molecular methods with MALDI-TOF MS, DNA Sanger sequencer and Illumina MiSeq. To evaluate the impact of VHP decontamination (independent variable) on the number of microorganisms (response variable) ANOVA and LSD tests were used. After application of 10 processes of VHP decontamination, changes in the properties of the materials were assessed using FTIR spectroscopy, SEM microscopy and XPS spectrometry. RESULTS: The concentration of microorganisms was 101-104 CFU/100 cm2 on the tested surfaces and 102 CFU/m3 in the air. Twenty species of bacteria, one yeast and 16 filamentous fungi were identified, with the predominance of Bacillus, Staphylococcus, Alternaria, Aspergillus and Penicillium. Moreover, Janthinobacterium, Acremonium, Aureobasidium, Coprinellus and Cosmospora in the VBNC state were metagenomically detected. VHP decontamination resulted in a reduction in the majority of tested microbial strains by a minimum of 3 log, and all tested mixed cultures inhabiting porous surfaces were above 98% and in the air, 100%. VHP decontamination did not affect the structural and morphological properties of cotton fibres, wood or stainless steel. CONCLUSIONS: VHP decontamination at a concentration of 300 ppm for 20 min can be used for the holistic disinfection of air, surfaces and equipment in oral surgery offices.

Multimodal Approach to Assessment of Fecal Microbiota Donors based on Three Complementary Methods.

Methods of stool assessment are mostly focused on next-generation sequencing (NGS) or classical culturing, but only rarely both. We conducted a series of experiments using a multi-method approach to trace the stability of gut microbiota in various donors over time, to find the best method for the proper selection of fecal donors and to find "super-donor" indicators. Ten consecutive stools donated by each of three donors were used for the experiments (30 stools in total). The experiments assessed bacterial viability measured by flow cytometry, stool culturing on different media and in various conditions, and NGS (90 samples in total). There were no statistically significant differences between live and dead cell numbers; however, we found a group of cells classified as not-dead-not-alive, which may be possibly important in selection of "good" donors. Donor C, being a regular stool donor, was characterized by the largest number of cultivable species (64). Cultivable core microbiota (shared by all donors) was composed of only 16 species. ANCOM analysis of NGS data highlighted particular genera to be more abundant in one donor vs. the others. There was a correlation between the not-dead-not-alive group found in flow cytometry and Anaeroplasma found by NGS, and we could distinguish a regular stool donor from the others. In this work, we showed that combining various methods of microbiota assessment gives more information than each method separately.