The measurement of the enterovirus and the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in sewage water is relevant in the early detection of the introduction or disappearance of these viruses in the ecosystem. We evaluated the co-circulation of the enteroviruses and SARS-CoV-2 in 81 sewage water samples collected between September 2021 and April 2023 from different regions of north and southeast Romania, at the border with Ukraine. We used, for the molecular detection of the pathogens, the multiplex real-time polymerase chain reaction (PCR) assay produced for respiratory samples and the Respiratory 2.1 Plus panel Biofire Film array. The isolation of enteroviruses was performed on cell culture lines, in accordance with the World Health Organization (WHO) recommendations. By molecular investigations, we detected the SARS-CoV-2 in 22 (27%) samples, and the human rhinovirus/enterovirus in 64 (79%) samples. By isolation on cell culture lines, 27 samples (33,33%) were positive for non-polio enteroviruses, and no poliovirus strains were isolated, proving the maintenance of the polio-free status in Romania. In an emergency situation, the molecular detection of the pathogens in sewage water using a PCR system integrating sample preparation, amplification, detection, and analysis in 1 h could be implemented.

Non-polio enteroviruses (NPEV) cause significant disease worldwide. Population-based sero-surveillance, by measuring antibodies against specific NPEV types, provides additional information on past circulation and the prediction for future upsurges. Virus neutralisation assays (VNA), the current method of choice for measuring NPEV type specific antibodies, are not entirely standardised. Via the European Non-Polio Enterovirus Network, we organised a VNA quality assessment in which twelve laboratories participated. We provided five echovirus (E) types (E1, E18, E30 G2, E30 G6 and E6) and intravenous immunoglobulins (IVIG) as a sample for the NPEV VNA quality assessment. Differences in VNA protocols and neutralising Ab (nAb) titres were found between the participating laboratories with geometric coefficients of variation ranging from 10.3-62.9 %. Mixed-effects regression analysis indicated a small but significant effect of type of cell line used. Harmonisation of cell line passage number, however, did not improve variation between laboratories. Calibration by making use of a reference sample, reduced variation between laboratories but differences in nAb titres remained higher than two log2 dilution steps. In conclusion, sero-surveillance data from different laboratories should be compared with caution and standardised protocols are needed.

Wastewater surveillance (WWS) was developed in the early 1960s for the detection of poliovirus (PV) circulation in the population. It has been used to monitor several pathogens, including non-polio enteroviruses (NPEVs), which are increasingly recognised as causes of morbidity in children. However, when applying WWS to a new pathogen, it is important to consider the purpose of such a study as well as the suitability of the chosen methodology. With this purpose, the European Non-Polio Enterovirus Network (ENPEN) organised an expert webinar to discuss its history, methods, and applications; its evolution from a culture-based method to molecular detection; and future implementation of next generation sequencing (NGS). The first simulation experiments with PV calculated that a 400 mL sewage sample is sufficient for the detection of viral particles if 1:10,000 people excrete poliovirus in a population of 700,000 people. If the method is applied correctly, several NPEV types are detected. Despite culture-based methods remaining the gold standard for WWS, direct methods followed by molecular-based and sequence-based assays have been developed, not only for enterovirus but for several pathogens. Along with case-based sentinel and/or syndromic surveillance, WWS for NPEV and other pathogens represents an inexpensive, flexible, anonymised, reliable, population-based tool for monitoring outbreaks and the (re)emergence of these virus types/strains within the general population.

The place of non-criteria antiphospholipid antibodies (aPLs) in the diagnosis of antiphospholipid syndrome (APS) is still debatable. The aim of this research was to evaluate the correlations between the titres of non-criteria aPLs (anti-phosphatidylethanolamine (aPE), anti-phosphatidylserine (aPS), and anti-prothrombin (aPT) antibodies), and the ones of the already studied criteria aPLs (anti-cardiolipin (aCL) and anti-β2 glycoprotein I-aβ2GPI antibodies). Altogether, 72 APS (30 primary and 42 secondary) patients were included in our study. High correlation coefficients (rs) were found between aPS IgM and aCL IgM, overall (0.77, p < 0.01), as well as in the primary (0.81, p < 0.01), and secondary (0.75, p < 0.01) APS subgroups. Low or statistically insignificant correlations were observed between IgG/IgM isotypes of aPT and aCL, or aβ2GPI, in the entire study population, and when evaluating the subgroups. Therefore, moderate correlations were mainly identified between the tested non-criteria antibodies and the criteria ones, suggesting little added value for the use of the tested non-criteria aPLs, with the exception of aPT, which seems to have different kinetics and might be a promising APS diagnostic tool.

Objectives To assess the frequency of multiple thrombophilia-associated mutations and polymorphisms in a selected population of high-risk pregnancies. Methods Thrombophilia screening was performed for 1,500 pregnant women with prior pregnancy complications or thrombotic events. Nine thrombophilia-associated mutations or polymorphisms were screened: factor V Leiden, factor V H1299R, prothrombin G20210A, MTHFR C677T, MTHFR A1298C, factor XIII V34L, PAI-1 4G/5G polymorphisms, EPCR G4600A, EPCR C4678G. Results Out of the 1,500 patients, 1,291 fulfilled the criteria for data interpretation. All patients had low-risk thrombophilia-associated genetic variants. Only 1.24 % of cases presented high-risk abnormalities (homozygous factor V Leiden/prothrombin G20210A, or both mutations in heterozygous form). Heterozygous factor V Leiden occurred in 10.38 % of cases, while only 5.81 % carried heterozygous prothrombin G20210A mutation. The frequency of prothrombin G20210A mutation was higher (10.37 %) in the subgroup associating factor V Leiden, than in the subgroup lacking it (5.36 %). Low-risk genetic variants occurred with a higher frequency: 23.78 % factor V H1299R, 57.32 % MTHFR C677T, 55.54 % MTHFR A1298C, 44.07 % factor XIII V34L, 73.20 % PAI-1 4G/5G polymorphisms, 69.64 % EPCR G4600A, and 69.63 % EPCR C4678G. Conclusions All patients had at least one prothrombotic genetic mutation or variant. Our data highlight the need for thrombophilia screening, including low-risk genetic variants, in a high-risk population of pregnant women with a history of pregnancy complications or thrombotic events.

Cancer patients have higher prevalences of antiphospholipid antibodies (aPLs), occasionally associated with thrombotic events. A cross-sectional study regarding the presence of criteria (IgG/IgM anti-cardiolipin-aCL, anti-β2 glycoprotein I-aβ2GPI) and non-criteria (IgG/IgM anti-phosphatidylserine-aPS, anti-phosphatidylethanolamine-aPE, anti-prothrombin-aPT) aPLs in 146 patients with involuntary weight loss was performed. None of the patients had thrombotic events during the study. Out of the 36 cancer patients, 33 had non-hematologic malignancies. In the cancer subgroup, 60% of the patients had at least one positive aPL, with significantly more patients being positive for aβ2GPI IgG compared with the non-cancer subgroup—p = 0.03, OR = 2.23 (1.02–4.88). When evaluating the titres, aCL IgG/IgM, aβ2GPI IgG, aPE IgG, and aPS IgG had significantly higher values in cancer patients, the best cancer predictor being aβ2GPI IgG—AUC 0.642 (0.542–0.742). Gastrointestinal cancer patients were studied separately, and aCL IgM positivity was significantly higher—p = 0.008, OR = 6.69 (1.35–33.02). Both the titres of aCL IgM (p = 0.006) and aPS IgM (p = 0.03) were higher in the gastrointestinal cancer subgroup, with aCL IgM being the best predictor for gastrointestinal cancer development—AUC 0.808 (0.685–0.932). Despite criteria and non-criteria aPLs being frequent in cancer, their connection with thrombosis in these patients is probably dependent on other important risk factors and needs further research.

Background: In view of the high recurrence rate of urinary tract infections and the increasing number of germs resistant to multiple antibiotics, the aim of the present study was to evaluate the antibacterial properties of clove, oregano, thyme, eucalyptus, tea tree essential oils (EOs) against 32 isolates of Escherichia coli and 28 isolates of Klebsiella pneumoniae from patients with urinary tract infections (UTI). Methods: The agar disk diffusion method was used to assess the susceptibility of these isolates to essential oils and the minimal inhibitory concentration (MIC), and the minimal bactericidal concentration (MBC) were determined. Results: Our results suggest that volatile phenols (such as carvacrol in oregano EO, thymol in thyme EO, and eugenol in clove EO) are more efficacious as antibacterial than non-aromatic compounds (such as eucalyptol in eucalyptus EO and terpinene derivatives in tea tree EO). Conclusion : The oregano EO, followed by thyme appear to have the highest efficacy against Escherichia coli and Klebsiella pneumoniae isolates investigated.

In 2018, an upsurge in echovirus 30 (E30) infections was reported in Europe. We conducted a large-scale epidemiologic and evolutionary study of 1,329 E30 strains collected in 22 countries in Europe during 2016-2018. Most E30 cases affected persons 0-4 years of age (29%) and 25-34 years of age (27%). Sequences were divided into 6 genetic clades (G1-G6). Most (53%) sequences belonged to G1, followed by G6 (23%), G2 (17%), G4 (4%), G3 (0.3%), and G5 (0.2%). Each clade encompassed unique individual recombinant forms; G1 and G4 displayed >2 unique recombinant forms. Rapid turnover of new clades and recombinant forms occurred over time. Clades G1 and G6 dominated in 2018, suggesting the E30 upsurge was caused by emergence of 2 distinct clades circulating in Europe. Investigation into the mechanisms behind the rapid turnover of E30 is crucial for clarifying the epidemiology and evolution of these enterovirus infections.

Due to the outbreak of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), combined with the risk of polio importation from Ukraine, we evaluated the presence of SARS-CoV-2 and enteroviruses in 25 sewage water samples from Romania, concentrated using the WHO method between January 2020 and January 2021. Surveillance for enteroviruses and SARS-CoV-2 are relevant in the calculation of prevalence estimates as well as early detection of the introduction or disappearance of these viruses. For SARS-CoV-2 detection, we used two immunochromatographic nucleocapsid antigenic tests as well as real-time PCR assays, produced for respiratory samples. The isolation of cell culture lines, in accordance with the WHO recommendations, was carried out for enterovirus detection. Twenty-three of the samples investigated were positive in rapid tests for SARS-CoV-2, while the RNA of SARS-CoV-2, detected with Respiratory 2.1 plus a panel Biofire Film array, was present in eight samples. The Allplex 2019-nCoV assay was used for the validation of the tests. There were three genes detected in one sample, E, RdPR, and N, two genes, E and RdPR, in one sample, two genes, RdPR and N, in four samples, one gene, RdPR, in five samples and one gene, N, in one sample. Eight samples were positive for non-polio enteroviruses, and no poliovirus strains were isolated. This study suggests the presence of SARS-CoV-2 and enteroviruses in Romanian sewage water in 2020. As such, our results indicate that a rapid, more specific test should be developed especially for the detection of SARS-CoV-2 in sewage water.

Objective: The aim of the current study is to assess the prevalence of hepatitis B and the risk of hepatitis reactivation in carriers of hepatitis B virus (HBV) cancer patients who underwent chemotherapy for gynecologic and/or breast cancers in a single institution, during a period of five years, and to identify a relationship to some particular chemotherapy regimen, more prone to lead to reactivation. Material and methods: We conducted a retrospective chart review on all consecutive oncological patients treated for a gynecologic and/or breast cancers who presented for the first time to the Gynecologic Oncology Department of Filantropia Hospital, Bucharest, Romania, between January 2016 and December 2020. Results: A total of 1 895 patients diagnosed with ovarian, cervical, endometrial or breast cancers were admitted to hospital for systemic therapy during the mentioned period. Among these, only four patients (two patients with breast cancers, one cervical cancer and one endometrial carcinoma) were chronic carriers of HBV surface antigen (HBsAg positive). Patients received a variety of chemotherapeutic regimens including corticosteroids, gemcitabine, cisplatin, carboplatin, taxanes and anthracyclines. We report one reactivation that occurred in one occult carrier of hepatitis B virus diagnosed with breast cancer (HBsAg negative, hepatitis B core antibody positive - HBcAb), initially excluded from this study, as being screened negative for HBV, treated with taxanes-based chemotherapy and corticosteroids. Conclusion: HBV reactivation had a low incidence in our population of patients diagnosed with gynecologic or breast cancer who received systemic chemotherapy. The HBV reactivation risk was positively correlated with breast cancer and to taxanes-based regimens and glucocorticoids. Further studies to identify additional risk factors of HBV infection reactivation in gynecologic oncology patients and possible risk reducing measures are warranted.

In the context of the current Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) pandemic, a very common problem is the choice of optimal disinfectant for the decontamination / disinfection of contaminated surfaces, medical salons, medical equipment, multi purpose protective equipment, medical instruments and areas frequented by persons. The paper presents tests for possibly contaminated the virucidal and bactericidal effect of new solutions based on silver and gold nanoparticles (AgNPs / AuNPs) and quaternary ammonium salts against SARS-CoV-2, by a surrogate method that quantifies the cytopathic effect of a Sabin attenuated LSc/2ab strain of poliovirus on an immortal rhabdomyosarcoma cell line.

OBJECTIVE To identify the osteoprotegerin (OPG) correlates with antiphospholipid syndrome (APS) parameters. METHODS Our cohort included 40 patients with primary APS disease associated with systemic lupus erythematosus (SLE) (mean age, 43.7 years; 87% female). Data on cardiovascular risk factors and specific clinical events in APS were collected. Then we tested OPG and 10 criteria and noncriteria antiphospholipid antibodies (aPLs) on preserved specimens in all cases. RESULTS A total of 26 patients (65%) had high serum OPG levels. Patients with high OPG were mostly overweight. In patients with SLE, the OPG levels were associated with anti-double-stranded DNA (anti-dsDNA) and anti-Sm titers. However, we did not find significant correlations of the OPG with any of the 10 aPLs tested. Also, we found no relationship regarding venous APS events. CONCLUSION In APS, high OPG levels are not linked to serum aPL expression.

The co-circulation of the enterovirus and norovirus species was evaluated by culture and molecular methods, in the framework of the risk assessment for importation of vaccine-derived poliovirus type 1(VDPV1), from Ukraine in Romania.56 sewage water samples collected from the north and southeast Romania, between April and August 2017 were investigated.Xpert EV assay and Xpert Norovirus assay, standardized methods for the detection of human enterovirus (HEV) in the cerebrospinal fluid respectively for the detection of norovirus genogroups in stool samples, were used for the molecular detections of these viruses in the concentrated sewage water.Additionally, the cell culture lines were used for human enterovirus isolation.18 human enteroviruses (32,14%) were detected by using the Xpert EV assay and 17 (30,35%) nonpolio enteroviruses (NPEV) were isolated on cell culture lines.Norovirus genogroup G II was detected in 20 samples (35,7%), followed by norovirus genogroups GI and GII, in 8 samples (14,28%).Taking into account our results, a new diagnostic algorithm could be evaluated in an emergency situation, the sewage water concentration by using the WHO method, and screening by molecular detection.

Carbapenemase-producing Klebsiella pneumoniae (CPKP) is one of the important causes of nosocomial infection worldwide. A number of 61 CPKP strains isolated in 2016, from clinical specimens of individual patients in an emergency university hospital from Romania were evaluated by molecular methods. The most isolates of Klebsiella pneumoniae were sensitive to amikacin (77%), and fosfomycinum and nitrofurantoin (55% each). OXA-48 was the most frequently identified genotype in 73,77% of cases (45 isolates), 13,11% (eight isolates) presented the combinations bla OXA-48 and bla NDM, 1,63 % (one isolate) presented blaOXA-48 and bla KPC, 8,19 % (five isolates) presented bla KPC, and in 3,27 %, (two isolates) was recorded bla NDM, all by use of the Xpert Carba-R Assay. The XbaI PFGE profile-based dendrogram of 20 K. pneumoniae strains selected from 61 strains extended to the 58% similarity level overall. However, the 88% criterion resolved 8 pulsotypes with 4 single-strain pulsotypes (D, E, F, and H) and 4 multiple-strain pulsotypes (A, B, C, and G). Our results confirm an epidemic expansion of a major OXA-48 positive clone in hospitals from Romania. The prudent use of antibiotics and the introduction of laboratory screening of patients for colonisation with carbapenem resistant bacteria will help to control the spread of these bacteria in the hospital.

Backgroud. Which factors determine venous thrombotic events in some antiphospholipid syndrome (APS) patients and arterial thrombosis or conditions related to pregnancy in others has not been established yet. Purpose. The aim of this study was to search the antiphospholipid antibodies (APLAs) correlates in regard to deep vein thrombosis (DVT) in patients with systemic lupus erythematosus (SLE) and APS. Methods. Twenty-nine patients fulfilling the criteria of both SLE and APS were included. Complete anamnesis and clinical examination was performed on inclusion. Also, for all patients, disease activity was assessed by the SLEDAI score. An extended APLAs profile, ten Abs, was searched. Results. The titers of IgG anticardiolipin (aCL), IgG anti-β2 glycoprotein I (aβ2GPI), IgG antiphosphatidylethanolamine (aPE), and also of IgG antiprothrombin (aPT) were significant higher in patients with DVT history. After analysis by ROC curve and univariate logistic regression, the strongest association was found for IgG aPE. Also, in multivariate analysis, SLEDAI score correlated with the DVT antecedents. Conclusions. IgG aPE might be involved in DVT pathogenic pathways in patients with SLE and APS as their titers remain significantly higher in patients with previous DVT. Lupus patients with DVT events represent a subgroup of patients with more severe underlying pathology.

IntroductionThe patients with antiphospholipid syndrome (APS) associate an increased risk of atherosclerosis.ObjectiveTo determine the predictors of an abnormal ankle‐brachial index (ABI), surrogate measure of atherosclerosis, in patients with APS.MethodsThe ABI was measured according to standard recommendations in 106 patients. Traditional cardiovascular risk factors were assessed in all cases. A large spectrum of APS antibodies was determined in 73 patients.ResultsA total of 106 patients diagnosed with APS were included. 28.3% patients included were found to have low ABI. Anti‐beta 2‐glycoprotein I (aβ2GPI) IgG antibodies [4.00 (1.00‐79.00) vs 3.00 (0.00‐29.00) U/mL, P = 0.02] and antiprothrombin (aPT) IgM antibodies [4.50 (0.00‐82.00) vs 3.00 (0.00‐14.00) U/mL, P = 0.05] titers were found to be higher in patients with abnormal ABI. However, after multivariate regression analysis, only the aβ2GPI IgG titer remained predictor of low ABI (P = 0.04).Conclusionsaβ2GPI IgG associated with impaired ABI in patients with APS. This relation might reflect their involvement in the atherosclerosis occurrence.

Thrombophilia is the increased tendency to form blood clots. The condition can be acquired or inherited. The risk of venous thrombosis and pulmonary embolism in healthy women is highest during pregnancy. The most common forms of inherited thrombophilia are factor V Leiden mutation and prothrombin gene mutation. The aim of this study was to perform a screening of pregnant women hospitalized in our unity for hereditary thrombophilia. Our study group included 107 pregnant women. The study is retrospective. The incidence of high-risk inherited thrombophilias is low in our study, so the frequency of homozygous factor V Leiden mutation is equal with the incidence of homozygous prothrombin G20210A mutation - 0.9%.

Introduction . Two cases of circulating vaccine-derived poliovirus type 1(cVDPV1), from southwestern Ukraine, bordering Romania, were confirmed in 2015 and the environmental enterovirus surveillance was enhanced in our country. The molecular detection of human enteroviruses as a screening test followed by isolation on cell culture lines or sequencing could be proposed as a new diagnosis algorithm. Material and Methods . The sensitivity of two molecular methods for the detection of enterovirus strains in 10 mL of sewage water (15 samples) was studied with Film Array ME panel BioFire (Biomerieux, France) and Xpert EV assay (Cepheid, USA). These are standardized methods for the detection of microorganisms in the cerebrospinal fluid (CSF). Results . Of the 15 samples, six enterovirus strains were detected using Film Array ME, four enterovirus strains were detected using Xpert EV assay, while only two nonpolio enterovirus strains were isolated on RD cell line, using the standard WHO algorithm. However, only one of the strains detected by the standard WHO algorithm was detected by one of the molecular methods. Conclusions . The molecular methods for enterovirus detection are more sensitive than the virus isolation on cell culture lines, but in one case the virus isolated on RD cell line was not detected by the molecular methods. The results could be influenced by the small number of the samples investigated, by the volume and the concentration method used for samples tested, and by the limits of detection (LoD) of the enterovirus species in the samples, depending on the method used.

Thrombophilia is the increased tendency to form blood clots. The condition can be acquired or inherited. The risk of venous thrombosis and pulmonary embolism in healthy women is highest during pregnancy. The most common forms of inherited thrombophilia are factor V Leiden mutation and prothrombin gene mutation. The aim of this study was to perform a screening of pregnant women hospitalized in our unity for hereditary thrombophilia. Our study group included 107 pregnant women. The study is retrospective. The incidence of high-risk inherited thrombophilias is low in our study, so the frequency of homozygous factor V Leiden mutation is equal with the incidence of homozygous prothrombin G20210A mutation - 0.9%.