Twenty-Month Monitoring of Humoral Immune Response to BNT162b2 Vaccine: Antibody Kinetics, Breakthrough Infections, and Adverse Effects.

BACKGROUND: Vaccination is one of the most effective life-saving medical interventions, and the introduction of SARS-CoV-2 vaccines was intended to prevent the serious implications of COVID-19. The objectives of the study were (i) to observe the humoral immune response to the BNT162b2 vaccine and SARS-CoV-2 infection (mainly breakthrough infections), (ii) to demonstrate the persistence of anti-SARS-CoV-2 antibodies over time in relation to the number of received vaccine doses and the course of infection, and (iii) to determine the adverse effects after primary vaccine doses. METHODS: To assess the humoral response, IgG and IgA anti-S1 antibodies were quantified by ELISA assays. In total, the tests were carried out seven times in almost two years. RESULTS: We demonstrated strong immunogenicity (compared to levels before primary vaccination, 150- and 20-fold increases in IgG and IgA, respectively) of the BNT162b2 vaccine. Over time, we observed a systematic decline in antibody levels, which may have contributed to breakthrough infections. Although they caused seroconversion similar to the booster, antibody levels in such patients fell more rapidly than after re-vaccination. On the other hand, in individuals who did not receive booster(s) and who did not present breakthrough infection, anti-SARS-CoV-2 antibodies returned to pre-vaccination levels after 20 months. The most commonly recognized adverse effects were injection site redness and swelling. CONCLUSION: Vaccination is highly effective in preventing the most severe outcomes of COVID-19 and should be performed regardless of prior infection. Booster doses significantly enhance anti-SARS-CoV-2 antibody levels and, in contrast to those obtained by breakthrough infection, they remain longer.

Clinical Validation of GenBody COVID-19 Ag, Nasal and Nasopharyngeal Rapid Antigen Tests for Detection of SARS-CoV-2 in European Adult Population.

Accurate and rapid identification of COVID-19 is critical for effective patient treatment and disease outcomes, as well as the prevention of SARS-CoV-2 transmission. Rapid antigen tests (RATs) for identifying SARS-CoV-2 are simpler, faster and less expensive than molecular assays. Any new product to be considered a medical device is subject to evaluation and data analysis to verify the in vitro diagnostic ability to achieve its intended purpose. Clinical validation of such a test is a prerequisite before clinical application. This study was a clinical validation on adult Europeans of GenBody COVID-19 Ag, nasal and nasopharyngeal RATs. A set of 103 positive and 301 negative from nose and nasopharynx samples confirmed by RT-qPCR were examined. The tests were safe to use and showed 100% specificity in both specimens, and high sensitivity of 94.17% (95%CI 87.75% to 97.83%) and 97.09% (95%CI 91.72% to 99.4%), respectively. The parameters were significantly better for samples with higher virus loads (the highest for CT ≤ 25). The GenBody COVID-19 Ag RATs are inexpensive (compared to RT-qPCR), reliable and rapid with high sensitivity and specificity, making them suitable for diagnosis and timely isolation and treatment of COVID-19 patients, contributing to the better control of virus spread.

Clinically Relevant ß-Lactam Resistance Genes in Wastewater Treatment Plants.

Antimicrobial resistance (AMR) is one of the largest global concerns due to its influence in multiple areas, which is consistent with One Health's concept of close interconnections between people, animals, plants, and their shared environments. Antibiotic-resistant bacteria (ARB) and antibiotic-resistance genes (ARGs) circulate constantly in various niches, sediments, water sources, soil, and wastes of the animal and plant sectors, and is linked to human activities. Sewage of different origins gets to the wastewater treatment plants (WWTPs), where ARB and ARG removal efficiency is still insufficient, leading to their transmission to discharge points and further dissemination. Thus, WWTPs are believed to be reservoirs of ARGs and the source of spreading AMR. According to a World Health Organization report, the most critical pathogens for public health include Gram-negative bacteria resistant to third-generation cephalosporins and carbapenems (last-choice drugs), which represent ß-lactams, the most widely used antibiotics. Therefore, this paper aimed to present the available research data for ARGs in WWTPs that confer resistance to ß-lactam antibiotics, with a particular emphasis on clinically important life-threatening mechanisms of resistance, including extended-spectrum ß-lactamases (ESBLs) and carbapenemases (KPC, NDM).

Differentiation of SARS-CoV-2 Variants Using RT-qPCRs by Targeting Recurrent Mutation Sites: A Diagnostic Laboratory Experience from Multi-Center Regional Study, August 2020-December 2021, Poland.

Rapid identification of SARS-CoV-2 variants is essential for epidemiological surveillance. RT-qPCR-based variant differentiation tests can be used to quickly screen large sets of samples for relevant variants of concern/interest; this study was conducted on specimens collected at 11 centers located in Poland during routine SARS-CoV-2 diagnostics between August 2020 and December 2021. A total of 1096 samples (with CT < 30) were screened for Alpha, Beta, Delta, Kappa and Omicron variants using commercial assays targeting repeat mutation sites. Variants were assigned to 434 (39.6%) specimens; the remaining 662 (60.4%) samples were not classified (no tested mutations detected). Alpha (n = 289; 66.59%), Delta (n = 115; 26.5%), Kappa (n = 30; 6.91%) and Omicron (n = 2; 0.46%) variants were identified and their distribution changed over time. The first Alpha variant appeared in October 2020, and it began to gradually increase its proportion of the virus population by June 2021. In July 2021, it was replaced by the Delta variant, which already dominated by the end of the year. The first Kappa was detected in October 2021, while Omicron was found in December 2021. The screening of samples allowed the determination of epidemiological trends over a time interval reflecting the national COVID-19 waves.

Viral Etiological Agent(s) of Respiratory Tract Infections in Symptomatic Individuals during the Second Wave of COVID-19 Pandemic: A Single Drive-Thru Mobile Collection Site Study.

One of the tools to contain the SARS-CoV-2 pandemic was to increase the number of performed tests and to improve the access to diagnostics. To this effect, mobile collection sites (MCSs) were established. This study was performed on samples collected at the MCS between November 2020 and March 2021. We aimed to confirm/exclude SARS-CoV-2, differentiate SARS-CoV-2 variants, and detect other respiratory pathogens. SARS-CoV-2 and other respiratory viruses were identified by RT-qPCRs. A total of 876 (46.35%) SARS-CoV-2 positive specimens in the diagnostic tests were identified. The wild-type variant was determined in 667 (76.14%) samples; the remaining 209 (23.86%) samples specimens were identified as Alpha variant. A total of 51 (5.6%) non-SARS-CoV-2 cases were detected in retrospective studies. These accounted for 33 cases of mono-infection including rhinovirus (RV), human adenovirus (HAdV), human metapneumovirus (HMPV), enterovirus (EV), and influenza virus, and 18 cases of co-infection (SARS-CoV-2 with RV or HAdV or HMPV, and RV with EV). Our research shows that the results obtained from the MCS have value in epidemiological studies, reflecting national trends on a micro scale. Although the spread of COVID-19 is a major public health concern, SARS-CoV-2 is not the only pathogen responsible for respiratory infections.

Impact of the Nucleic Acid Extraction Method and the RT-qPCR Assay on SARS-CoV-2 Detection in Low-Viral Samples.

COVID-19 was initially reported in China at the end of 2019 and soon thereafter, in March 2020, the WHO declared it a pandemic. Until October 2021, over 240 million COVID-19 cases were recorded, with 4.9 mln deaths. In order to stop the spread of this disease, it is crucial to monitor and detect any infected person. The etiologic agent of COVID-19 is a novel coronavirus called SARS-CoV-2. The gold standard for the detection of the virus is the RT-qPCR method. This study evaluated two RNA extraction methods and four commercial RT-qPCR assays routinely used in diagnostic laboratories for detecting SARS-CoV-2 in human specimens from the upper respiratory tract. We analyzed a panel of 70 clinical samples with varying RNA loads. Our study demonstrated the significant impact of the diagnostic methods selected by the laboratory on the SARS-CoV-2 detection in clinical specimens with low viral loads.

Case Report of COVID-19 after Full Vaccination: Viral Loads and Anti-SARS-CoV-2 Antibodies.

The introduction of effective vaccines against SARS-CoV-2 is expected to prevent COVID-19. However, sporadic cases of infection in vaccinated persons have been reported. We describe a case of a double-dose vaccinated woman with COVID-19. All stages of infection were observed, from no identification of virus, then the start of the infection, a high viral load, coming out of viraemia, and finally no detection of the virus. Despite the high viral load, the woman demonstrated mild COVID-19 symptoms, manifested only by a sore throat. The antibody results showed that she produced both post-infectious and post-vaccination immune responses. Phylogenetic analysis of the obtained viral genome sequence indicated that the virus belonged to the UK SARS-CoV-2 lineage B.1.1.7 (GR 501Y.V1; 20I/S:501Y.V1; Alpha variant).

Occurrence of Beta-Lactamases in Colistin-Resistant Enterobacterales Strains in Poland - a Pilot Study.

Sixty-five colistin-resistant Enterobacterales isolates recovered from different clinical specimens were analyzed. The strains were collected in 12 hospitals all over Poland within a period of nine months. Strains were analyzed for eight genes from the mcr family. The presence of mcr-1 gene was detected in three Escherichia coli strains. The 45/65 isolates were identified as ESBL producers. CTX-M-1-like enzymes were the most common ESBLs (n = 40). One E. coli and seven Klebsiella pneumoniae strains produced carbapenemases, with the NDM being produced by five isolates. Among all the strains tested, four and five were resistant to new drugs meropenem/vaborbactam and ceftazidime/avibactam, respectively.

Substituent Effect in the Cation Radicals of Monosubstituted Benzenes.

In 30 monosubstituted benzene cation radicals, studied at the ωB97XD/aug-cc-pVTZ level, the phenyl rings usually adopt a compressed form, but a differently compressed form-equivalent to an elongated one-may coexist. The computational and literature ionization potentials are well correlated. The geometrical and magnetic aromaticity, estimated using HOMA and NICS indices, show the systems to be structurally aromatic but magnetically antiaromatic or only weakly aromatic. The partial charge is split between the substituent and ring and varies the most at C(ipso). In the ring, the spin is 70%, concentrated equally at the C(ipso) and C(p) atoms. The sEDA(D) and pEDA(D) descriptors of the substituent effect in cation radicals, respectively, were determined. In cation radicals, the substituent effect on the σ-electron system is like that in the ground state. The effect on the π-electron systems is long-range, and its propagation in the radical quinone-like ring is unlike that in the neutral molecules. The pEDA(D) descriptor correlates well with the partial spin at C(ipso) and C(p) and weakly with the HOMA(D) index. The correlation of the spin at the ring π-electron system and the pEDA(D) descriptor shows that the electron charge supplied to the ring π-electron system and the spin flow oppositely.

Clinical and Molecular Findings of Infections Caused by Extended-Spectrum ß-Lactamase-Producing Enterobacterales in Patients with Solid Tumors: A Single-Center Study.

Infectious complications caused by multidrug-resistant bacteria are a serious clinical and therapeutic problem. Our study aimed to analyze the genetic characteristics of extended-spectrum ß-lactamase-producing Enterobacterales (ESBL-E) that cause multidrug-resistant infections in patients with solid tumors. Identification of ESBL-encoding genes was performed by polymerase chain reaction (PCR) and sequencing. The clonal relationship of the isolates was evaluated by pulsed-field gel electrophoresis. Multilocus sequence typing (MLST) was carried out for selected Escherichia coli and Klebsiella pneumoniae isolates. All E. coli strains were classified into phylogenetic groups using the PCR-based approach. There were 735 patients with clinical symptoms of infections tested, of which 44 (6.0%) were positive for ESBL-E on genotypic testing. The most frequent organism was E. coli (n = 24, 54.5%), followed by K. pneumoniae (n = 13, 29.5%), Proteus mirabilis (n = 3, 6.8%), Enterobacter cloacae cplx (n = 2, 4.5%), and Klebsiella oxytoca (n = 2, 4.5%). Overall, 31 (70.5%) of the ESBL-E isolates carried only blaCTX-M-1-like genes, and the genes were found to be blaCTX-M-15 (n = 30, 68.2%) or blaCTX-M-3 (n = 1, 2.3%). Eleven strains (25%) had blaCTX-M-9-like genes, mostly blaCTX-M-27 (n = 10, 22.7%) and unique blaCTX-M-65 (n = 1, 2.3%). One isolate possessed both blaCTX-M-15 and blaCTX-M-27 genes, and another one produced TEM-12 ESBL. MLST analysis revealed E. coli sequence type (ST) 131 and ST361, and K. pneumoniae ST16, ST307, and ST437. Among E. coli isolates, the B2 phylogenetic group was predominant. Most of the strains showed resistance to third-generation cephalosporins and fluoroquinolones, and susceptibility to aminoglycosides and carbapenems. Patients with solid cancer and ESBL-E infections require special management since they are a population with a high threat of antibiotic-resistant infections. Carbapenems and aminoglycosides remain active antibiotics against these infections.

Epidemic Territorial Spread of IncP-2-Type VIM-2 Carbapenemase-Encoding Megaplasmids in Nosocomial Pseudomonas aeruginosa Populations.

In 2003 to 2004, the first five VIM-2 metallo-ß-lactamase (MBL)-producing Pseudomonas aeruginosa (MPPA) isolates with an In4-like integron, In461 (aadB-blaVIM-2-aadA6), on conjugative plasmids were identified in three hospitals in Poland. In 2005 to 2015, MPPA expanded much in the country, and as many as 80 isolates in a collection of 454 MPPA (∼18%) had In461, one of the two most common MBL-encoding integrons. The organisms occurred in 49 hospitals in 33 cities of 11/16 main administrative regions. Pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) classified them into 55 pulsotypes and 35 sequence types (STs), respectively, revealing their remarkable genetic diversity overall, with only a few small clonal clusters. S1 nuclease/hybridization assays and mating of 63 representative isolates showed that ∼85% of these had large In461-carrying plasmids, ∼350 to 550 kb, usually self-transmitting with high efficiency (∼10-1 to 10-2 per donor cell). The plasmids from 19 isolates were sequenced and subjected to structural and single-nucleotide-polymorphism (SNP)-based phylogenetic analysis. These formed a subgroup within a family of IncP-2-type megaplasmids, observed worldwide in pseudomonads from various environments and conferring resistance/tolerance to multiple stress factors, including antibiotics. Their microdiversity in Poland arose mainly from acquisition of different accessory fragments, as well as new resistance genes and multiplication of these. Short-read sequence and/or PCR mapping confirmed the In461-carrying plasmids in the remaining isolates to be the IncP-2 types. The study demonstrated a large-scale epidemic spread of multidrug resistance plasmids in P. aeruginosa populations, creating an epidemiological threat. It contributes to the knowledge on IncP-2 types, which are interesting research objects in resistance epidemiology, environmental microbiology, and biotechnology.

Characteristics of ESBL-Producing Enterobacterales Colonizing the Gastrointestinal Tract in Patients Admitted to the Oncological Hospital.

We analyzed the prevalence and genetic characteristics of the extended-spectrum ß-lactamases (ESBLs)-producing Enterobacterales isolated from adult patients hospitalized in the oncological center in 2019. Out of 9372 patients admitted to the hospital, 1373 had been in various medical facilities during the last year, which was an indication to perform a screening test for ESBL-producing Enterobacterales colonizing their gastrointestinal tract. In eighty-three patients (6.1%), 85 ESBL producers were detected. These isolates included the following: Escherichia coli (n = 67; 78.8%), Klebsiella pneumoniae (n = 14; 16.5%), Enterobacter cloacae cplx (n = 3; 3.5%), and Klebsiella oxytoca (n = 1; 1.2%). CTX-M-1-like enzymes were the most common ESBLs (n = 67; 78.8%). Two K. pneumoniae isolates (2/85; 2.4%) additionally produced New Delhi-metallo-ß-lactamases (NDM). All isolates, except for K. oxytoca, were typed by pulsed-field gel electrophoresis (PFGE) and demonstrated high genetic diversity. The most prevalent phylogroups of E. coli were B2 group (n = 30; 44.8%), followed by A group (n = 25; 37.3%). These observations have motivated us to investigate the link between ESBL-E colonization and infection among patients with solid tumors.

Thallium(I) Tropolonates: Synthesis, Structure, Spectral Characteristics, and Antimicrobial Activity Compared to Lead(II) and Bismuth(III) Analogues.

Synthesis, single-crystal X-ray determination diffraction and FT-IR, NMR (1H, 13C, 19F and 205Tl), UV-vis, and luminescence spectra characteristics were described for series of thallium(I) compounds: thallium(I) triflate (Tl(OTf)), 1:1 co-crystals of thallium(I) triflate and tropolone (Htrop), Tl(OTf)·Htrop, as well as simple thallium(I) chelates: Tl(trop) (1), Tl(5-metrop) (2), Tl(hino) (3), with Htrop, 5-methyltropolone (5-meHtrop), 4-isopropyltropolone (hinokitiol, Hhino), respectively, and additionally more complex {Tl@[Tl(hino)]6}(OTf) (4) compound. Comparison of their antimicrobial activity with selected lead(II) and bismuth(III) analogs and free ligands showed that only bismuth(III) complexes demonstrated significant antimicrobial activity, from two- to fivefold larger than the free ligands.

The genetic background of antibiotic resistance among clinical uropathogenic Escherichia coli strains.

The spreading mechanisms of antibiotic resistance are related to many bacterial and environment factors. The overuse of antibiotics is leading to an unceasing emergence of new multidrug resistant strains. This problem also concerns uropathogenic Escherichia coli strains, which is the most common pathogen causing urinary tract infections. The aim of this study was the genetic analysis of antibiotic resistance in comparison to the phenotypic background of E. coli strains. The characterized collection of E. coli strains isolated 10 years ago from the urine samples of patients with urinary tract infections was used for antimicrobial susceptibility testing (the disc diffusion method) and analysis of antibiotic resistance genes (PCR reaction, sequencing). Additionally, the presence of ESBL strains was analyzed. Fourteen genes were associated with resistance to beta-lactams, aminoglycosides, sulfonamides and quinolones. The genetic analysis revealed that blaTEM-1 and sul2 were present in almost all of the studied strains. Other drug-resistance genes were very rare or non-existent. Otherwise, the phenotypic resistance to fluoroquinolones was well correlated with the genotypic background of the studied bacteria. The presence of particular genes and specific mutations indicate a high bacterial potential to multidrug resistance. On the other hand, it needs to be emphasized that the standard disk diffusion test for the routine antimicrobial susceptibility analysis is still the best way to estimate the current situation of bacterial drug-resistance.

Multiregional dissemination of KPC-producing Klebsiella pneumoniae ST258/ST512 genotypes in Poland, 2010-14.

Objectives: In 2008-09, the KPC carbapenemase epidemiology in Poland was dominated by a Klebsiella pneumoniae ST258 KPC-2 outbreak in Warsaw and its administrative region. The aim of this study was to analyse the situation in 2010-14, with a focus on new outbreaks in other parts of the country. Methods: KPCs were detected in all suspected isolates by PCR. The detailed study was performed on 173 isolates from 2010 to 2012, and included PFGE and MLST, PCR identification of K. pneumoniae clonal group CG258 clades and potential specificity markers ( pilv-1 , IS 66 and prp ), PCR mapping of Tn 4401 transposons, and plasmid analysis by nuclease S1 profiling and PCR-based replicon typing. Results: Six hundred and eight KPC cases were identified in Poland in 2010-14, almost half of which occurred in the Warsaw region, and another half in four other areas. The new outbreaks were caused by four K. pneumoniae CG258 genotypes, different from each other and from the organisms spreading in Warsaw. The new lineages were ST258 or ST512 of clade II, and had specific compositions of potential ST258/ST512 clonal markers. The isolates produced KPC-3 encoded by Tn 4401 a or Tn 4401 b elements on plasmids with single or multiple replicons, including I2, FII K (+/-FIB K ), 'FII Y -like', X3 and R. Of other species, Citrobacter freundii ST17 and Enterobacter cloacae ST254 with KPC-2 were identified in a Warsaw hospital. Conclusions: The study showed remarkable changes in the KPC epidemiology in Poland in 2010-14, which, following the localized regional spread in the early phase, has converted into multiregional dissemination.

KPC-2-producing Klebsiella pneumoniae ST11 in a Children's Hospital in Poland.

Four Klebsiella pneumoniae isolates from children hospitalized over 10 months in an intensive care unit in a children's teaching hospital in Poland were analyzed. All of the isolates belonged to a single pulsotype and sequence type (ST) 11, and produced the KPC-2 carbapenemase and extended-spectrum ß-lactamase (ESBL) CTX-M-15. They were resistant to a variety of antimicrobials, and their ß-lactam resistance patterns were typical for KPC producers. This is one of few cases of identification of KPC (or carbapenemase)-producing K. pneumoniae in a pediatric center in Poland.

Usefulness of CHROMagar Candida Medium, Biochemical Methods--API ID32C and VITEK 2 Compact and Two MALDI-TOF MS Systems for Candida spp. Identification.

This study was conducted to compare of the yeasts identification results obtained with two new systems using the MALDI-TOF MS technique with the ones obtained using the routine identification methods of Candida spp. in clinical microbiology laboratories. All 124 Candida spp. isolates were recovered from the routine examination of clinical specimens in microbiological laboratories and collected in the Centre of Quality Control in Microbiology in Warsaw (Poland). Our findings confirm the high agreement (98%) of fungal identification using the standard, biochemistry laboratory methods and mass spectrometry technique.

Sequence types 235, 111, and 132 predominate among multidrug-resistant pseudomonas aeruginosa clinical isolates in Croatia.

A population analysis of 103 multidrug-resistant Pseudomonas aeruginosa isolates from Croatian hospitals was performed. Twelve sequence types (STs) were identified, with a predominance of international clones ST235 (serotype O11 [41%]), ST111 (serotype O12 [15%]), and ST132 (serotype O6 [11%]). Overexpression of the natural AmpC cephalosporinase was common (42%), but only a few ST235 or ST111 isolates produced VIM-1 or VIM-2 metallo-ß-lactamases or PER-1 or GES-7 extended-spectrum ß-lactamases.

Characterization of two new CTX-M-25-group extended-spectrum ß-lactamase variants identified in Escherichia coli isolates from Israel.

OBJECTIVES: We characterized two new CTX-M-type extended-spectrum ß-lactamase (ESBL) variants in Escherichia coli isolates from stool samples of two elderly patients admitted at the Tel Aviv Sourasky Medical Center, Israel. Both patients underwent treatment with cephalosporins prior to isolation of the E. coli strains. METHODS: ESBLs were detected by the double-disk synergy test and PCR-sequencing of ß-lactamase genes. The bla(CTX-M) genes were cloned into the pCR-BluntII-TOPO vector in E. coli TOP10. The role of amino-acid substitutions V77A and D240G was analyzed by site-directed mutagenesis of the bla(CTX-M-94) and bla(CTX-M-100) genes and comparative characterization of the resulting E. coli recombinants. MICs of ß-lactams were determined by Etest. Plasmid profiling, mating experiments, replicon typing and sequencing of bla(CTX-M) flanking regions were performed to identify the genetic background of the new CTX-M variants. RESULTS: The novel CTX-M ß-lactamases, CTX-M-94 and -100, belonged to the CTX-M-25-group. Both variants differed from CTX-M-25 by the substitution V77A, and from CTX-M-39 by D240G. CTX-M-94 differed from all CTX-M-25-group enzymes by the substitution F119L. Glycine-240 was associated with reduced susceptibility to ceftazidime and leucine-119 with increased resistance to ceftriaxone. bla(CTX-M-94) and bla(CTX-M-100) were located within ISEcp1 transposition units inserted into ∼93 kb non-conjugative IncFI and ∼130 kb conjugative IncA/C plasmids, respectively. The plasmids carried also different class 1 integrons. CONCLUSIONS: This is the first report on CTX-M-94 and -100 ESBLs, novel members of the CTX-M-25-group.