OBJECTIVES:We conducted a systematic review and meta-analysis to estimate maternal Group B Streptococcus (GBS) colonization, vertical transmission, serotype distribution, and antimicrobial resistance (AMR) burden of GBS in African pregnant women and newborn infants. METHODS:Literature searches were performed on CINAHL, EMBASE, Cochrane Library, Google Scholar, ProQuest, PsycINFO, MEDLINE/PubMed, WHO IRIS, Scopus, Web of Science, African Journals Online, and University repositories. Heterogeneity was evaluated using Cochrane's chi-square test and the I² statistics, and potential publication bias was assessed using Egger's regression intercept. RESULTS:A total of 116 studies from 29 African countries (n = 53,770 participants) were included. The pooled prevalence of maternal rectovaginal GBS colonization was 18% (95% confidence interval [CI]: 16%-20%). The pooled proportion of vertical transmission was 47% (95% CI: 39%-55%). Among maternal isolates, serotypes V (27%; 95% CI: 23%-32%), Ia (24%; 95% CI: 19%-29%), III (23%; 95% CI: 19%-26%), and II (19%; 95% CI: 13%-25%) were most frequently detected. Among neonatal isolates, serotypes V (31%; 95% CI: 22%-41%), Ia (26%; 95% CI: 17%-35%), III (23%; 95% CI: 15%-31%), and II (15%; 95% CI: 12%-19%) were identified. AMR was high for tetracycline (87%; 95% CI: 85%-88%) and cotrimoxazole (75%; 95% CI: 72%-78%), and low for β-lactams, including ampicillin (10%; 95% CI: 9%-12%) and penicillin (9%; 95% CI: 8%-11%). CONCLUSION:The prevalence of maternal GBS rectovaginal colonization and vertical transmission in Africa is high and comparable to global estimates, but there were marked regional variations. The predominance of serotypes V, Ia, and III supports their inclusion in multivalent vaccine formulations. Widespread resistance to tetracycline and cotrimoxazole and moderate macrolide resistance highlight the need for antimicrobial stewardship and ongoing surveillance.

To characterize real-world COVID-19 vaccine-induced protection, we assessed breakthrough infections and protective antibody thresholds among fully vaccinated nursing home residents (NHR) and staff (NHS). We conducted a cohort study to assess infection rates and risk factors of symptomatic breakthrough infections in 1605 NHR and 1200 NHS (2021) and 483 NHR (2022) in Belgium using multivariable Cox proportional hazards models adjusted for potential confounders. Additionally, a case-control analysis was used to compare pre-infection and peak vaccine response antibody levels between breakthrough cases and matched controls. Receiver Operating Characteristic (ROC) and net benefit models were fitted to evaluate the protective antibody thresholds in clinical decision making. In 2021, breakthrough infection incidence was 10.7 (95

Abstract Manual chemical methods are commonly used for ultrasound probe disinfection but are known to be operator dependent and subject to variability. Automated ultraviolet-C (UV-C) light - emitting diode (LED) disinfection represents a chemical and mercury free alternative, however, clinical evidence comparing UV-C LED to conventional disinfectants for high-level disinfection of ultrasound probes remains limited. In this single-centre non-inferiority study conducted in the departments of gynaecology, obstetrics and urology of a Belgian university hospital, automated UV-C LED disinfection was compared to manual chlorine dioxide disinfection for the reprocessing of transabdominal and transvaginal ultrasound probes. Following clinical use, probes were decontaminated by healthcare practitioners alternating between both reprocessing methods. Samples were collected after treatment and processed for microbial analysis to assess the effectiveness of the decontamination process. A total of 296 probes were included, comprising 86 transabdominal and 210 transvaginal probes. UV-C LED disinfection achieved effectiveness rates of 92.86% for transabdominal probes and 97.17% for transvaginal probes, compared with 79.49% and 93.27%, respectively for chlorine dioxide disinfection. Positive samples following UV-C disinfection yielded single colonies whereas chlorine dioxide treated probes showed greater variability in colony counts and species diversity. Although the total reprocessing time for UV-C disinfection was longer, manual handling times were comparable between both methods. Here, we demonstrated that UV-C LED disinfection was non-inferior to chlorine dioxide disinfection for reprocessing of transabdominal and transvaginal ultrasound probes under routine conditions.

Objectives: To assess the feasibility and reproducibility of predicting antimicrobial resistance (AMR) in Escherichia coli from MALDI-TOF mass spectrometry data using a standardized, open-source machine learning (ML) workflow, we systematically compared four ML algorithms, evaluated the impact of culture conditions, extract storage, and spectral preprocessing on model performance, and validated results through nested cross-validation with statistical significance testing. Methods: A total of 282 clinical E. coli isolates were analyzed. Two MALDI-TOF MS datasets were generated from freshly cultured extracts (T1) and recultured isolates one year later (T3), yielding 4468 spectra. A third dataset from the T1 extracts stored at −20 °C for one year (T2) was evaluated for spectral stability but excluded from primary modeling likely due to storage-induced degradation. Protein spectra (m/z 2000–15,000) were preprocessed using an in-house developed MALDI-TOF preprocessing pipeline (MTPP) comprising variance stabilization, Savitzky–Golay smoothing, SNIP baseline correction, TIC normalization, LOWESS alignment, and MAD-based peak detection (SNR ≥ 3), yielding 121 m/z features. Four classifiers—Random Forest (RF), Logistic Regression, Support Vector Machine, and Gradient Boosting—were trained to predict resistance to 11 antibiotics using nested cross-validation: outer GroupShuffleSplit (5-fold, isolate-level) for evaluation and inner GroupKFold for recursive feature elimination (RFECV) and hyperparameter tuning (RandomizedSearchCV). Classification thresholds were optimized via the precision–recall curve. Model performance was assessed using AUROC, AUPRC, F1-score, Matthews Correlation Coefficient (MCC), and bootstrap 95% confidence intervals (1000 replicates). Pairwise model comparisons were tested with McNemar’s chi-squared test. Results: Among the 12 antibiotics included in the analysis (meropenem excluded for absence of resistance), resistance prevalence ranged from 1.1% (colistin) to 59.9% (amoxicillin). Colistin was subsequently also excluded from ML modeling due to insufficient resistant isolates (n = 3), leaving 11 antibiotics for prediction. The best predictive performance was observed for ciprofloxacin (AUROC 0.76 [95% CI 0.74–0.77]; F1 0.54; MCC 0.38) and ceftazidime (AUROC 0.68 [0.65–0.71]; F1 0.36; MCC 0.29), using 13 and 37 RFECV-selected features, respectively. Amoxicillin achieved the highest F1-score (0.76), driven by high recall (0.98) but modest AUROC (0.58). No meaningful predictive signal was detected for amikacin, cefepime, or tigecycline (AUROC ≤ 0.57, F1 ≤ 0.17), attributable to extreme class imbalance, and no robust multi-peak resistance signature was detected in this dataset. McNemar’s test confirmed that RF significantly outperformed Logistic Regression for all antibiotics (p < 0.01), while Gradient Boosting performed comparably to RF for ciprofloxacin (p = 0.17) and ceftazidime (p = 0.28). Frozen extracts (T2) produced lower spectral similarity and were excluded from model training; the aligned T1+3 dataset yielded the most stable performance across metrics. Conclusions: Machine learning analysis of MALDI-TOF spectra enables reproducible AMR prediction for selected antibiotics in E. coli, with ciprofloxacin and ceftazidime showing the strongest signal. Nested isolate-level cross-validation, multi-model comparison with statistical testing, and open-source code provide a transparent, reproducible foundation for integrating ML-assisted MALDI-TOF analysis into diagnostic AMR surveillance. Extract storage at −20 °C degrades spectral quality and should be avoided in ML training workflows.

Probiotics offer a promising alternative to antibiotics in animal husbandry. This study evaluated the inhibitory potential of live cells and cell-free supernatants (CFS) from 31 putative probiotic strains against two multidrug-resistant (MDR) enterotoxigenic Escherichia coli (ETEC) strains. Methods included antibiogram profiling, zone of inhibition assays, hemolytic activity, and auto-aggregation tests. Results highlighted strain-specific antimicrobial heterogeneity. Lactobacillus acidophilus ATCC 4356 and Pediococcus acidilactici CNCTC 6986 exhibited the strongest contact-dependent inhibition against SP11 and SP31, respectively, while Lactiplantibacillus plantarum FW0BV0887 displayed superior CFS-mediated activity. Notably, L. acidophilus ATCC 4356 and L. helveticus ATCC 15009 showed consistent efficacy in both assays. Safety assessments confirmed non-hemolytic phenotypes for most strains, excluding Enterococcus faecium. Furthermore, at the 24-h plateau, L. crispatus FB141-CAN-1 achieved the highest auto-aggregation capacity (88.03 ± 8.60%), followed closely by L. acidophilus ATCC 4356 (75.88 ± 2.62%), indicating putative adhesive potential. Overall, these findings identify specific probiotics with distinct antimicrobial mechanisms and preliminary safety profiles based on hemolytic activity, supporting their potential as biocontrol agents and warranting further in vivo validation.

Background: Despite widespread Plasmodium falciparum resistance, intermittent preventive treatment in pregnancy (IPTp) with sulfadoxine–pyrimethamine is more protective against adverse pregnancy and birth outcomes compared to IPTp with dihydroartemisinin–piperaquine, even though dihydroartemisinin–piperaquine is superior against malaria-related outcomes. We assessed whether adding metronidazole to sulfadoxine–pyrimethamine is more protective against adverse pregnancy and birth outcomes compared to IPTp with dihydroartemisinin–piperaquine in an area of high malaria transmission and parasite resistance to sulfadoxine–pyrimethamine, and where curable sexually transmitted infections (STIs) in pregnancy, including bacterial vaginosis, are common. Methods: From December 2019 to March 2022, we enrolled pregnant women without HIV into ASPIRE, an individually randomised, double-blinded, three-arm, partly placebo-controlled trial in Zambia. Participants received monthly IPTp with standard sulfadoxine–pyrimethamine (3 tablets 500mg sulfadoxine and 25mg pyrimethamine) plus placebo metronidazole in Arm 1, or standard sulfadoxine–pyrimethamine plus metronidazole (4 tablets 500mg) in Arm 2, or dihydroartemisinin-piperaquine plus metronidazole (3 tablets 40mg dihydroartemisinin and 320mg piperaquine once daily for 3 consecutive days) plus metronidazole (4 tablets 500mg) in Arm 3. The primary outcome was any adverse pregnancy or birth outcome (spontaneous abortion, stillbirth, low birthweight, preterm birth, or neonatal death). Secondary outcomes included components of the primary outcome, small-for-gestational age, and day 28 treatment efficacy of IPTp regimens against asymptomatic Plasmodium falciparum malaria infection and asymptomatic curable STIs. We used a modified intention-to-treat analysis for the primary outcome. ClinicalTrials.gov: NCT04189744. Findings: 5436 pregnant women were allocated to three treatment-arms. Overall, 16·3% (288/1763) of women in Arm 1 (reference) had adverse pregnancy and birth outcomes compared to 16·6% (290/1745) in Arm 2 (p=0·82), and 16·0% (281/1755) in Arm 3 (p=0·79). Of individual components of adverse pregnancy and birth outcomes in the primary outcome, none differed across arms. However, women in Arm 3 were at greater risk of delivering small-for-gestational age newborns relative to Arm 1, 28·4% (476/1678) vs 24·5% (408/1663) (p=0·01). In Arms 1, 2, and 3, the day 28 clearance of asymptomatic peripheral malaria detected by PCR was 72·1% (555/770), 71·8% (541/754), and 93·5% (722/772), respectively. In Arm 1, day 28 clearance of asymptomatic STIs by PCR, and bacterial vaginosis by microscopy using the Nugent score, were: Chlamydia trachomatis 98·4% (60/61), Neisseria gonorrhoea 84·1% (95/113), Trichomonas vaginalis 75·0% (96/128), and bacterial vaginosis 34·3% (193/563). In Arm 2, day 28 clearance rates were: C. trachomatis 89·5% (51/57), N. gonorrhoea 84·2% (96/114), T. vaginalis 87·5% (105/120), and bacterial vaginosis 53·9% (289/536). In Arm 3, day 28 clearance rates were: C. trachomatis 80·8% (42/52), N. gonorrhoea 84·9% (84/99), T. vaginalis 89·8% (115/128), and bacterial vaginosis 62·2% (335/536). Interpretation: Neither Arm 2 nor 3 were superior to Arm 1 for protection against adverse pregnancy and birth outcomes. Arm 3 was superior to Arms 1 and 2 against malaria infection, whereas all IPTp regimens reduced curable STIs, including bacterial vaginosis. Sulfadoxine-pyrimethamine may serve as an important tool fo

Anaerobic bacteremia due to Prevotella spp. is uncommon, and "Candidatus Prevotella massiliensis" has rarely been reported from clinical specimens. We describe a case of bacteremia caused by "P. massiliensis" in an 18-year-old woman presenting with fever and abdominal pain following medical abortion and subsequent suction curettage. An anaerobic blood-culture bottle yielded small Gram-negative rods that could not be identified by MALDI-TOF MS using a routine reference database. Near-complete 16S rRNA gene sequencing demonstrated 100% sequence identity to the reference sequence associated with "P. massiliensis," supporting species-level identification. Antimicrobial susceptibility testing showed resistance to clindamycin and susceptibility to metronidazole and penicillin. To our knowledge, this is the first report of "P. massiliensis" bacteremia in the context of medical abortion. This case demonstrates both the pathogenic potential of this rarely reported species and the limitations of routine MALDI-TOF MS identification for recently described or database-limited Prevotella spp., underscoring the value of 16S rRNA gene sequencing when routine methods are inconclusive.

Bacterial vaginosis (BV) and vaginal candidiasis are prominent causes of vaginal infections, leading to discomfort and negative pregnancy outcomes. Despite their significance, there is limited data on the prevalence, progression, and contributing factors of BV and vaginal Candida colonization among pregnant women in low-resource settings such as Ethiopia. Thus, the aim of this paper is to determine the prevalence and associated factors of BV and vaginal Candida colonization, and antifungal susceptibility patterns in Eastern Ethiopia. Exploratively, we assessed the relationship between these infections and adverse pregnancy outcomes among pregnant women. A total of 217 pregnant women, ranging from 12 to 22 weeks of gestation, were enrolled and followed until birth or pregnancy termination. Data on sociodemographic information, pregnancy history, and current conditions were gathered through interviews. Two vaginal swabs were collected for microbiological analysis, using Nugent scoring for BV, culture and Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry for Candida detection. Data were collected using Redcap and analyzed using STATA version 17. Multivariable logistic regression was employed to identify associated factors at a significance level of p < 0.05. This study indicated that 43% (95% confidence interval (CI): 36.6%, 50.2%) of the participants had BV or vaginal Candida colonization or both. The overall prevalence of BV and vaginal Candida colonization were 27.7% (95% CI: 21.8%, 34.1%) and 23.5% (95% CI: 18.4%, 29.6%), respectively. Factors associated with BV included antibiotic use (AOR = 9.47, 95% CI: 4.09-21.94) and vaginal douching (AOR = 6.93, 95% CI: 2.86-16.77). Similarly, antibiotic use (AOR = 4.18, 95% CI: 1.78-9.80) and vaginal douching (AOR = 5.48, 95% CI: 2.39-12.56) were significantly associated with Candida colonization. BV increased the likelihood of adverse birth outcomes by 1.89 times and preterm birth alone by 3.89 times. BV and vaginal Candida colonization are highly prevalent among pregnant women in Eastern Ethiopia and are associated with modifiable behavioral and clinical factors. The findings underscore the importance of improved understanding of vaginal microbiota dynamics during pregnancy and their potential links with adverse maternal and neonatal outcomes.

Human papillomavirus (HPV) genotype classification relies on DNA sequence similarity to reference (prototype) sequences. Most HPV assays used for cervical cancer screening were clinically validated against European HPV prototypes. However, the impact of HPV sequence polymorphisms on test performance remains unexplored. We evaluated whether sequence variation in E6/E7 relative to HPV prototypes affects test performance by analyzing cervicovaginal samples from 990 women enrolled (2019-2022) across Belgium, Portugal, Brazil and Ecuador. Samples were tested using cobas, Anyplex, and next-generation sequencing (Ampliseq/Ion Torrent targeting E6/E7). Sequence variation was defined as the proportion of single nucleotide polymorphisms across E6/E7 relative to reference sequence. Sequence variation was, on average, higher in HPV-negative than HPV-positive samples for HPV16 (0.46% vs 0.13%) and HPV18 (0.44% vs 0.37%) using cobas. Similar patterns were observed with Anyplex (HPV16: 0.78% vs 0.13%, HPV33: 0.66% vs 0.40%, HPV58: 0.79% vs 0.53, and HPV66: 1.14% vs 0.25%). For HPV45, sequence variation was, on average, higher in HPV-positive than HPV-negative samples when tested with Anyplex (0.87% vs 0.43%). For HPV types 18, 31, 35, 39, 51, 52, 56, 59 and 68, the mean sequence variation was similar between HPV-negative and HPV-positive samples using Anyplex. Our findings show that sequence variation relative to prototypes may impact test performance.

This study investigates the degradation of resistant α-1,4-glucan by vaginal bacterial species, with a focus on Gardnerella spp., to elucidate its role in bacterial vaginosis (BV). The ability of Gardnerella vaginalis, Gardnerella swidsinskii, Gardnerella leopoldii, Gardnerella piotii, Lactobacillus iners, and Lactobacillus crispatus was assessed to metabolize an ungelatinized, labeled form of raw amylose, a degradation-resistant α-1,4-glucan. The enzymatic activity of these species was evaluated in vitro, and its association with BV was examined in vaginal swabs. Gardnerella vaginalis, G. swidsinskii, and G. leopoldii demonstrated the best ability to degrade resistant α-1,4-glucan in vitro. Unlike the cell-bound, S-layer-associated glycogen-degrading activity in L. crispatus, this α-glucosidase activity in Gardnerella was also extracellular, but not cell-bound and not repressed by glucose. Vaginal swabs showing high rates of resistant α-1,4-glucan degradation activity were associated with BV, particularly in the concurrent presence of G. leopoldii, G. swidsinskii, and G. vaginalis. These findings suggest a role of α-1,4-glucan degradation in BV pathogenesis mediated by Gardnerella species. The results indicate the potential of targeting bacterial amylase activity as therapeutic strategy for BV prevention and treatment.

With the onset of the severe acute respiratory syndrome coronavirus-2 pandemic, controversial theories emerged regarding the potential consequences of the virus on pregnant women and delivery outcomes. Over the past three years, various studies have reported various data on pregnant women infected with covid and pregnancy-related complications, including preterm birth, stillbirth, preeclampsia, cesarean delivery. However; the exact influence of the virus and possibility of vertical transmission remained unclear. Here, we described the clinical features and delivery outcomes in 16 laboratory confirmed COVID-19-infected mothers who referred to a hospital in northern Iran from August 2020 to December 2021. We collected Clinical records, laboratory results, and chest CT scans,as well as samples as maternal peripheral blood, umbilical cord blood, placental blood, vaginal secretions, placental tissue, breast milk after the first lactation, neonatal throat swab, and peripheral blood. We evaluated all the aforementioned specimens based on molecular and serological assays to answer the questions raised on the possibility of vertical transmission of COVID-19 and transfer of maternal immunity to the neonates. SARS-CoV-2 RNA was not detected in vaginal secretions and placental tissue. SARS-CoV-2 IgG and IgM antibodies were detected in 15 and 4 maternal blood samples, respectively, in one breast milk sample (IgM), two umbilical cord blood (IgG) samples, two placental blood (IgG) and two neonatal blood (IgG) samples. Chest CT scans of abnormal cases revealed typical signs of viral pneumonia. The current study suggests an association between SARS-CoV-2 infection and an increased risk of preterm birth. However, no intrauterine vertical transmission of SARS-CoV-2 was identified. These results also suggest passive IgG transfer from infected mothers to their neonates.

Aptamer lateral flow assay (ALFA) for testing T. vaginalis (Tv) in cervicovaginal lavage (CVL).

Background/Objectives: DNA extraction from dried blood spot (DBS) samples is often applied in neonatal screening programs. Although various methods to extract DNA from DBSs have been described, the optimal approach remains unclear. Therefore, this study aimed to compare and optimize extraction methods to establish a reliable and efficient protocol for human DNA extraction from DBSs. Methods: We conducted a back-to-back comparison of five different DNA extraction methods on 20 DBS samples: three column-based kits (QIAamp DNA mini kit, High Pure PCR Template Preparation kit, DNeasy Blood & Tissue kit) and two in-house boiling methods (one using TE buffer, one using Chelex-100 resin). DNA recovery was measured with DeNovix DS-11 and ACTB qPCR. Further optimization of elution volumes and starting material was performed on the best-performing methods (sample size = 5). Additionally, T-cell receptor excision circle (TREC) DNA was assessed by qPCR as an application. Results: The Chelex boiling method yielded significantly (p < 0.0001) higher ACTB DNA concentrations compared to the other methods. Column-based methods showed low DNA recovery, except for Roche, which showed significantly (p < 0.0001) higher DNA concentrations than the other column-based methods, as measured by DeNovix DS-11. Decreasing elution volumes (150 vs. 100 vs. 50 µL) increased ACTB DNA concentrations significantly, while increasing starting material (two vs. one 6 mm spot) did not. Conclusions: We identified an easy and cost-effective optimized DNA extraction method using Chelex from DBSs, with an elution volume of 50 µL and 1 × 6 mm DBS punch, which is particularly advantageous for research in low-resource settings and large populations, such as neonatal screening programs.

Background/Objectives: This study mapped antibody dynamics across three COVID-19 vaccination rounds (primary course, first, and second booster with BNT162b2) in Belgian nursing home residents (NHRs). Methods: Within a national SARS-CoV-2 serosurveillance study (February 2021–September 2022) across Belgian nursing homes, dried blood spots were collected, on which anti-spike SARS-CoV-2 IgG antibodies were quantified by ELISA in international units/mL (IU/mL). Sociodemographic data were collected at the study start and infection history and vaccination data at each sampling round. Results: Infection-naïve NHRs had low antibody levels after primary course vaccination (geometric mean concentration (GMC) 292 IU/mL, 95% confidence interval (95% CI): 197–432), but increased tenfold after first booster (GMC 2168 IU/mL, 95% CI: 1554–3027). While antibodies among NHRs significantly declined within six months after primary vaccination (p < 0.0001), they remained stable for nine months post-booster (p > 0.05). Among primary vaccine non-responders, 92% (95% CI: 82–97%) developed antibodies after the first booster (GMC 594 IU/mL, 95% CI: 416–849), though tenfold lower than initial responders (GMC 4642 IU/mL, 95% CI: 3577–6022). Conclusions: These findings demonstrate that NHRs require tailored vaccination, prioritizing repeated immunization to improve serological outcomes in poor responders such as infection-naive NHRs. Regular immune monitoring could aid in implementing evidence-based vaccine strategies, ensuring optimal protection for vulnerable populations against SARS-CoV-2 and other infectious threats.

In line with the global research priorities on sexually transmitted infections (STIs), we developed a molecular point-of-care test (POCT) for the parasite Trichomonas vaginalis that causes the STI trichomoniasis. Trichomoniasis remains the most common curable nonviral STI. We report on the use of specific T. vaginalis single-stranded-tailed DNA primers in combination with recombinase polymerase amplification (RPA) in a protein-free nucleic acid lateral flow (NALF) device for use at the point of care. The use of aminated DNA probes eliminates the need for proteins (such as streptavidin or hapten-specific antibodies) for detection of the DNA amplicons, simplifying the manufacturing process and improving reproducibility and cost-effectiveness. The estimated shelf life of the NALF devices is at least 6.6 months at 22 °C, and the devices exhibited high reproducibility. The RPA-NALF requires three simple operator steps with minimal instrumentation and takes approximately 30 min from sample preparation to interpretation of the result. It is specific to T. vaginalis and can detect 1.3 × 103 cells/mL with visual readout or 282 cells/mL with the aid of a portable LFA reader. Analysis of the two sets of clinical genomic DNA extracts showed that RPA-NALF is positive for all samples positive by culture assay and for samples with Cq ≤ 23 with the S-DiaMGTV qPCR. Finally, RPA-NALF is positive on biobanked vaginal swabs with T. vaginalis Cq ≤ 25 with the Allplex qPCR assays. These results demonstrate that RPA-NALF can specifically detect a moderate load of T. vaginalis even in the presence of other microbial DNA and cells and other components of clinical samples.

In line with the global research priorities on sexually transmitted infections (STIs), we developed a molecular point-of-care test (POCT) for the parasite Trichomonas vaginalis that causes the STI trichomoniasis. Trichomoniasis remains the most common curable nonviral STI. We report on the use of specific T. vaginalis single-stranded-tailed DNA primers in combination with recombinase polymerase amplification (RPA) in a protein-free nucleic acid lateral flow (NALF) device for use at the point of care. The use of aminated DNA probes eliminates the need for proteins (such as streptavidin or hapten-specific antibodies) for detection of the DNA amplicons, simplifying the manufacturing process and improving reproducibility and cost-effectiveness. The estimated shelf life of the NALF devices is at least 6.6 months at 22 °C, and the devices exhibited high reproducibility. The RPA-NALF requires three simple operator steps with minimal instrumentation and takes approximately 30 min from sample preparation to interpretation of the result. It is specific to T. vaginalis and can detect 1.3 × 103 cells/mL with visual readout or 282 cells/mL with the aid of a portable LFA reader. Analysis of the two sets of clinical genomic DNA extracts showed that RPA-NALF is positive for all samples positive by culture assay and for samples with Cq ≤ 23 with the S-DiaMGTV qPCR. Finally, RPA-NALF is positive on biobanked vaginal swabs with T. vaginalis Cq ≤ 25 with the Allplex qPCR assays. These results demonstrate that RPA-NALF can specifically detect a moderate load of T. vaginalis even in the presence of other microbial DNA and cells and other components of clinical samples.

OBJECTIVES:Vulvovaginal candidiasis is the second most common cause of vaginitis among women of childbearing age worldwide and is often self-diagnosed and self-treated by women. However, self-management of vaginal symptoms has been shown to have very poor outcomes. Hence, reliable self-tests are essential for improving the management of VVC. Here, we evaluated and compared the diagnostic performance of four marketed Candida self-tests. METHODS:Remnant diagnostic material was collected from 140 vaginal samples analysed by culture in the Netherlands. This was used in four commercially available Candida self-tests: Candida albicans Rapid Test Beright, StrongStep Candida albicans Antigen Rapid Test, Tigsun VVC-TV-GV Combo Antigen Test and VagiQUICK. Furthermore, DNA was extracted and used to quantify Candida using qPCR. The performance of each test was compared with culture and qPCR results using appropriate statistical methods. RESULTS:The Beright, StrongStep, Tigsun and VagiQUICK tests exhibited sensitivities of 47.3%, 30.8%, 73.0% and 66.3% and specificities of 100%, 100%, 100% and 97.4%, respectively, compared with culture. Compared with qPCR, the tests exhibited a sensitivity of 43.4%, 60.2%, 60.8% and 54.7% and a specificity of 89.7%, 73.3%, 80.0% and 75.9%, respectively. Sensitivity improved with increasing Candida concentration thresholds. CONCLUSIONS:The investigated self-tests demonstrated varying performances, with both sensitivity and specificity being consistently lower than manufacturer claims. These findings underscore the urgent need for developing and validating more reliable self-tests to support adequate management of women's health. The sensitivity of currently marketed tests is only slightly better than empirical or self-diagnosis, which limits their clinical utility.

Background: T-cell receptor excision circles (TRECs) and kappa-deleting recombination excision circles (KRECs) are markers of recent thymic and bone marrow output, respectively. As they have previously been associated with immunosenescence, we aimed to investigate their association with anti-spike SARS-CoV-2 (S1RBD) IgG antibody response after COVID-19 vaccination in nursing home residents (NHRs) and staff (NHS). Methods: We measured TREC and KREC levels and S1RBD IgG antibody levels from dried blood spots (DBSs) using in-house qPCRs and a commercial ELISA kit, respectively, in 200 participants (50 NHRs and 150 NHS). DBSs were collected in April 2021, approximately two months after primary course COVID-19 vaccination (BNT162b2). We assessed the association between TREC and KREC as dependent variables and age, sex, infection-priming status, and post-vaccination S1RBD-specific IgG concentrations as independent variables by simple and multiple linear regression. Results: TREC and KREC levels were significantly lower in NHRs compared with NHS and were negatively correlated with age (p < 0.001). Neither TREC nor KREC levels were significantly associated with SARS-CoV-2 antibody concentrations (p > 0.05). Conclusions: In our study population, TREC and KREC levels decreased with age and were statistically significantly lower in NHRs than NHS. They were, however, not associated with the antibody response after COVID-19 vaccination. Yet, additional research is warranted to explore their potential relevance in cellular immune responses or in combination with other biomarkers of immune function.