Biofilm Formation by Hospital-Acquired Resistant Bacteria Isolated from Respiratory Samples.

BACKGROUND: Hospital-acquired resistant infections (HARI) are infections, which develop 48 h or more after admission to a healthcare facility. HARI pose a considerably acute challenge, due to limited treatment options. These infections are associated bacterial biofilms, which act as a physical barrier to diverse external stresses, such as desiccation, antimicrobials and biocides. We assessed the influence of multiple factors on biofilm production by HARI -associated bacteria. METHODS: Bacteria were isolated from samples of patients with respiratory HARI who were hospitalized during 2020-2022 in north Israel. Following antibiotic susceptibility testing by disc diffusion or broth microdilution, biofilm formation capacities of resistant bacteria (methicillin-resistant staphylococcus aureus, extended spectrum beta-lactamase-producing Escherichia coli and Klebsiela pneumonia, and multidrug-resistant Pseudomonas aeruginosa and Acinetobacter baumannii) was assessed using the crystalline violet staining method. Data regarding season, time to infection, bacterial species, patient age and gender, year, and medical department were collected from the patient medical records. RESULTS: Among the 226 study isolates, K. pneumonia was the most prevalent (35.4%) bacteria, followed by P. aeruginosa (23.5%), and methicillin-resistant staphylococcus aureus (MRSA) (21.7%). A significantly higher rate of HARI was documented in 2022 compared to 2020-2021. The majority of isolates (63.3%) were strong biofilm producers, with K. pneumonia (50.3%) being most dominant, followed by P. aeruginosa (29.4%). Biofilm production strength was significantly affected by seasonality and hospitalization length, with strong biofilm production in autumn and in cases where hospitalization length exceeded 30 days. CONCLUSION: Biofilm production by HARI bacteria is influenced by bacterial species, season and hospitalization length.

Clinical isolate characteristics and demographics of patients with C.jejuni and C.coli infections in Northern Israel, 2015-2021.

C.coli is a significant cause of foodborne gastroenteritis worldwide, with the majority of cases attributed to C.jejuni. Although most clinical laboratories do not typically conduct antimicrobial susceptibility testing for C.coli, the rise in resistant strains has underscored the necessity for such testing and epidemiological surveillance. The current study presents clinical isolate characteristics and demographics of 221 patients with C.coli (coli and jejuni) infections in Northern Israel, between 2015 and 2021. Clinical and demographic data were collected from patient medical records. Susceptibility to erythromycin, tetracycline, ciprofloxacin, and gentamicin was assessed using the standard E-test. No significant correlations were found between bacterial species and patient ethnicity, patient gender, or duration of hospitalization. In contrast, significant differences were found between infecting species and patient age and age subgroup (P < 0.001). Furthermore, erythromycin resistance was observed in only 0.5% of the study population, while resistance to ciprofloxacin, tetracycline, and gentamicin was observed in 95%, 93%, and 2.3% of the population, respectively. The presented study underscores the need for routine surveillance of C.coli antibiotic resistance.

Nucleic acid hybridization-based detection of pathogenic RNA using microscale thermophoresis.

Infectious diseases are one of the world's leading causes of morbidity. Their rapid spread emphasizes the need for accurate and fast diagnostic methods for large-scale screening. Here, we describe a robust method for the detection of pathogens based on microscale thermophoresis (MST). The method involves the hybridization of a fluorescently labeled DNA probe to a target RNA and the assessment of thermophoretic migration of the resulting complex in solution within a 2 to 30-time window. We found that the thermophoretic migration of the nucleic acid-based probes is primarily determined by the fluorescent molecule used, rather than the nucleic acid sequence of the probe. Furthermore, a panel of uniformly labeled probes that bind to the same target RNA yields a more responsive detection pattern than a single probe, and moreover, can be used for the detection of specific pathogen variants. In addition, intercalating agents (ICA) can be used to alter migration directionality to improve detection sensitivity and resolving power by several orders of magnitude. We show that this approach can rapidly diagnose viral SARS-CoV2, influenza H1N1, artificial pathogen targets, and bacterial infections. Furthermore, it can be used for anti-microbial resistance testing within 2 h, demonstrating its diagnostic potential for early pathogen detection.

Associations of motility and auto-aggregation with biofilm-formation capacity levels in Clostridioidesdifficile.

Clostridioides difficile (C. difficile) is responsible for one of the most common nosocomial infections worldwide. This work assessed associations between biofilm-formation capacity levels of C. difficile and cell viability, motility, flagella, motility and auto-aggregation in 118 clinical isolates. Biofilm production was assessed by the crystal violet method. Cell viability was determined by BacTiter-Glo™ Microbial Cell Viability Assay and live-imaging microscopy. Expression levels of LuxS, Cwp84, Spo0A, PilA, and FliC were measured by real-time PCR. Motility was visually assessed in agar tubes. Auto-aggregation levels were determined by OD600 measurements. Out of 118 isolates, 66 (56 %) were biofilm producers, with most being strong or moderate producers. Cell viability, motility and auto-aggregation positively correlated with biofilm-production capacity (p = 0.0001, p = 0.036 and p < 0.0001, respectively). Positive associations were found between pilA, fliC and luxS expression levels and biofilm-production capacity (p = 0.04, p = 0.01, p = 0.036, respectively). This is the first report of associations between biofilm-formation capacity and cell viability, pilA, fliC, and luxS gene expression, auto-aggregation and motility. These correlations should be further explored to expand knowledge on the regulation of C. difficile biofilm formation, and pathogenesis, which will have notable implications on treatment options.

A Multi-Laboratory Evaluation of Commercial Monkeypox Virus Molecular Tests.

In this report, we describe the first national scale multi-laboratory evaluation of monkeypox virus (MPXV) DNA commercial PCR kits. The objective of this study was to evaluate 2 kits by different diagnostic laboratories across Israel. Ten standardized samples were tested simultaneously using the Novaplex (15 laboratories) and Bio-Speedy (seven laboratories) kits. An in-house assay based on previously published reactions was used as reference. Comparison of the results showed high intra-assay agreement between laboratories, with small variations for most samples. The in-house assay had an analytical detection limit of less than 10 copies per reaction. While the 2 commercial kits were able to detect specimens with low viral loads similarly to the in-house assay, significant differences were observed, in the Cq values and relative fluorescence (RF), between the assays. The RF signal of the in-house and Bio-Speedy assays ranged between 5,000 and 10,000 RFU, while the signal in the Novaplex assay was less than 600 RFU. Due to the kit measurement protocol, the Cq values of the Bio-Speedy kit were 5 to 7.5 cycles lower than those of the in-house assay. On the contrary, the Cq values of the Novaplex kit were significantly higher than those of the in-house assay, with differences of 3 to 5 cycles per sample. Our results suggest that while all assays were similar in their overall sensitivity, direct comparison of Cq values between them may be misleading. To our knowledge, this is the first methodical evaluation of commercial MPX test kits. We therefore anticipate that this study would help diagnostic laboratories in choosing a specific MPX detection assay. IMPORTANCE To the best of our knowledge, this study is the first methodical evaluation of commercial kits designed for Monkeypox virus detection. This was done by performing the same tests using the same sample set in multiple laboratories, simultaneously, on a national scale. It therefore provides important and unique information on the performance of such kits and provides a guideline for choosing the assay of choice for monkeypox virus diagnosis in a standard diagnostic laboratory. It also demonstrates potential complications when trying to compare the results of different assays, even when testing exactly the same samples, under identical conditions.

Altered somatic hypermutation patterns in COVID-19 patients classifies disease severity.

Introduction: The success of the human body in fighting SARS-CoV2 infection relies on lymphocytes and their antigen receptors. Identifying and characterizing clinically relevant receptors is of utmost importance. Methods: We report here the application of a machine learning approach, utilizing B cell receptor repertoire sequencing data from severely and mildly infected individuals with SARS-CoV2 compared with uninfected controls. Results: In contrast to previous studies, our approach successfully stratifies non-infected from infected individuals, as well as disease level of severity. The features that drive this classification are based on somatic hypermutation patterns, and point to alterations in the somatic hypermutation process in COVID-19 patients. Discussion: These features may be used to build and adapt therapeutic strategies to COVID-19, in particular to quantitatively assess potential diagnostic and therapeutic antibodies. These results constitute a proof of concept for future epidemiological challenges.

The effect of different C. difficile MLST strains on viability and activity of macrophages.

Objectives: Clostridioides difficile is the most common infectious agent of nosocomial diarrhea. C. difficile infection (CDI) pathogenesis and disease severity depend on its toxins (toxins A, B and binary) and on the host's immune response, especially the innate immune system. The current study examined the efficacy of macrophage activity, macrophages viability and cytokine secretion levelsin response to different sequence type (ST) strains of C. difficile. Methods: RAW 264.7 macrophages were exposed to six different strains of C. difficile as well as to both toxins A and B and macrophage viability was measured. The levels of four secreted cytokines were determined by RT-PCR and ELISA. Morphological changes to the macrophages were investigated by fluorescent microscopy. Results: Strains ST37 and ST42 affected macrophages' vitality the most. Toxins A and B led to a significant reduction in macrophages' vitality at most time points. In addition, starting at 30-min post-exposure to 5 ng/µl of both toxins led to significant differences in macrophage viability versus at lower concentrations. Furthermore, cytokine secretion levels, including IL-12, IL-6 and TNF-α, increased dramatically when macrophages were exposed to strains ST42 or ST104. Finally, gene expression surveys point to increases in IL-12 gene expression in response to both ST42 and ST104. Conclusions: C. difficile strains with higher toxins levels induced an increased activation of the innate immune system and may activate macrophages more profoundly resulting in secretion of higher levels of pro-inflammatory cytokines. However, higher toxin levels may also damage macrophages' normal skeletal structure, reducing macrophage viability.

The Impact of a Positive Biofire® FilmArray® Gastrointestinal Panel Result on Clinical Management and Outcomes.

The gold standard diagnostic method for gastrointestinal infections is stool culture, which has limited sensitivity and long turnaround time. Infection diagnosis recently shifted to syndrome-based panel assays. This study employed the FilmArray® Gastrointestinal Panel, which detects 22 pathogens simultaneously, to investigate gastrointestinal infection and pathogen distribution in 91 stool samples of patients hospitalized at the Tzafon Medical Center, Israel, during 2020, and to compare the clinical and demographic data of negative vs. positive samples. Among the 61 positive samples (67%), the most common pathogen was Campylobacter (34.4%). Positive test results were associated with a slightly younger patient age (p = 0.012), significantly higher post-diagnosis use of antibiotics (63.9% vs. 36.7%; p = 0.014), and shorter length of stay and time to discharge (p = 0.035, p = 0.003, respectively) than negative test results. To conclude, the FilmArray® Gastrointestinal Panel enabled the early identification of causative infectious agents and enhanced clinical management and outcomes.

Trend of Changes in Chloramphenicol Resistance during the Years 2017-2020: A Retrospective Report from Israel.

Resistant bacteria limit treatment options. This challenge has awakened interest in antibiotics that are no longer in use due to side effects, such as chloramphenicol. This work investigated trends in chloramphenicol resistance rates during 2017-2020 in bacteria isolated from diverse clinical samples at the Baruch Padeh Medical Center, Poriya, Israel. Bacteria were isolated from 3873 samples and identified using routine methods, including matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) technology. Chloramphenicol susceptibility was tested using a VITEK II instrument or by the Kirby-Bauer disk-diffusion test. The average chloramphenicol resistance rate was 24%, with no significant differences between study years. Chloramphenicol resistance was associated with sample origin (p < 0.001); isolates originating from sputum samples showed 49.8% resistance rate, compared to 2.3% of the body fluid isolates, 10.4% of the ear/eye isolates and 22.5% of the blood isolates. Furthermore, there was a significant decrease in chloramphenicol resistance among blood and ear/eye isolates during the study period (p = 0.01, p < 0.001, respectively). The highest resistance rate was among Pseudomonas aeruginosa isolates (50.5%). In conclusion, since chloramphenicol susceptibility seems to be retained, its comeback to the clinical world should be considered.

Different bile acids have versatile effects on sporulation, toxin levels and biofilm formation of different Clostridioides difficile strains.

Clostridioides difficile infection develops following ingestion of virulent stains by a susceptible host. Once germinated, toxins TcdA and TcdB, and in some of the strains binary toxin, are secreted, eliciting disease. Bile acids play a significant role in the process of spore germination and outgrowth, with cholate and its derivative enhancing colony formation, while chenodeoxycholate inhibit germination and outgrowth. This work investigated bile acids' impact on spore germination, toxin levels and biofilm formation in various strain types (STs). Thirty C. difficile isolates (A+ B+ CDT-\+) of different STs were exposed to increasing concentrations of the bile acids, cholic acid (CA), taurocholic acid (TCA) and chenodeoxycholic acid (CDCA). Following treatments, spore germination was determined. Toxin concentrations were semi-quantified using the C. Diff Tox A/B II™ kit. Biofilm formation was detected by the microplate assay with crystal violet. SYTO® 9 and propidium iodide staining were used for live and dead cell detection, respectively, inside the biofilm. Toxins levels were increased by 1.5-28-fold in response to CA and by 1.5-20-fold in response to TCA, and decreased by 1-37-fold due to CDCA exposure. CA had a concentration-dependent effect on biofilm formation, with the low concentration (0.1%) inducing- and the higher concentrations inhibiting biofilm formation, while CDCA significantly reduced biofilm production at all concentrations. There were no differences in the bile acids effects on different STs. Further investigation might identify a specific bile acids' combination with inhibitory effects on C. difficile toxin and biofilm production, which could modulate toxin formation to reduce the likelihood of developing CDI.

Impact of Helicobacter pylori virulence markers on clinical outcomes in adult populations.

BACKGROUND: In recent years, associations between specific virulence markers of Helicobacter pylori (H. pylori) and gastrointestinal disorders have been suggested. AIM: To investigate the presence of virulence factors including vacuolating cytotoxin A genotypes (s1m1, s1m2, s2m1, and s2m2), cytotoxin-associated gene A (CagA), and urease activity in H. pylori strains isolated from Arab and Jewish populations in northern Israel and to assess associations between these factors and patients' demographics and clinical outcomes. METHODS: Patients (n = 108) who underwent gastroscopy at the Baruch Padeh Medical Center, Poriya due to symptomatic gastroduodenal pathologies as part of H. pylori diagnosis were enrolled in the study. Gastric biopsy specimens were collected from the antrum of the stomach. Clinical condition was assessed by clinical pathology tests. Bacteria were isolated on modified BD Helicobacter Agar (BD Diagnostics, Sparks, MD, United States). Bacterial DNA was extracted, and PCR was performed to detect CagA and vacuolating cytotoxin A genes. Urease activity was assessed using a rapid urease test. RESULTS: A significant correlation was found between disease severity and patient ethnicity (P = 0.002). A significant correlation was found between CagA presence and the s1m1 genotype (P = 0.02), which is considered the most virulent genotype. Further, a higher level of urease activity was associated with isolates originating from the Jewish population. Moreover, higher urease activity levels were measured among CagA-/s1m1 and CagA-/s2m2 isolates. CONCLUSION: Our study highlights the importance of incorporating molecular methods for detection of virulence markers of H. pylori in order to tailor optimal treatments for each patient. Further investigation should be performed regarding associations between H. pylori virulence factors and ethnicity.

Should pregnant women be screened for SARS-CoV-2 infection? A prospective multicenter cohort study.

OBJECTIVE: Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), ranges from asymptomatic to severe infection. We aimed to compare the prevalence of COVID-19 in asymptomatic pregnant versus nonpregnant women in order to establish recommendations for a COVID-19 screening strategy. METHODS: A prospective multicenter cohort study was conducted. Asymptomatic pregnant or nonpregnant women after March 2020 (the time when COVID-19 was first detected in north Israel) were tested for SARS-CoV-2 using nasopharyngeal reverse transcription polymerase chain reaction test, anti-nucleocapsid IgG, and anti-spike IgG. Diagnosis was made if at least one test result was positive. Pregnant women were tested between 34 and 42 weeks, mostly at birth. RESULTS: Among the 297 participating women, 152 were pregnant and 145 were nonpregnant. The prevalence of asymptomatic COVID-19 was similar between the groups (4 [2.6%] and 8 [5.5%], respectively; P = 0.2). All women with COVID-19 delivered healthy appropriate-for-gestational-age babies without malformations, at term. CONCLUSIONS: The rate of asymptomatic COVID-19 in pregnant women is low and comparable to the rate among nonpregnant women. Pregnancy outcomes are favorable. Future screening programs should consider that one of 25 screened asymptomatic women will be positive.

Sexually Transmitted Diseases in Northern Israel: Insights From a Large Referral Laboratory.

OBJECTIVES: The aims of the study are to investigate the distribution and frequency of different sexually transmitted diseases (STDs) among a large study population of individuals undergoing STD investigation both in inpatient and STD clinic settings and to evaluate influence of test anonymity on the positivity rate of pathogens. MATERIAL AND METHODS: A retrospective study retrieved epidemiologic data from the following 3 sources: a secondary referral hospital and 2 STD clinics in Northern Israel. Positivity rate of Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Mycoplasma genitalium , and Trichomonas vaginalis (TV) was assessed and stratified based on age, sex, site of sampling, and anonymity of test. Adjusted odds ratios (ORs) were calculated by multivariable logistic regression. RESULTS: Overall, 3,753 assays were performed on 2,407 patients who were screened for STD. Chlamydia trachomatis (4.8%) was the most frequently detected STD, followed by NG (2.1%), MG (1.9%), and TV (0.6%). Mycoplasma genitalium (OR, 4.32; 95% CI, 1.70-10.97; p = .001) and NG (OR, 6.08; 95% CI, 2.18-16.96; p < .001) were significantly associated with male sex, while TV was more frequently encountered among female individuals (OR, 4.34; 95% CI, 1.49-12.50; p = .003). Mycoplasma genitalium infection was detected most commonly by urine samples, while rectal swabs were the leading source of positive tests for CT. Compared with fully identified patients, those tested anonymously were 6-fold more likely to be tested positive for TV (adjusted OR, 6.49; 95% CI, 2.06-20.42; p = .001). CONCLUSIONS: Chlamydia trachomatis and NG are the leading non-HIV STDs in Northern Israel. Anonymous tests predict higher positivity of TV. Rectal sampling should be increasingly used because of its efficacy in detecting CT infections.

Characterization of community-acquired Clostridioides difficile strains in Israel, 2020-2022.

Background: The prevalence of community-acquired Clostridioides difficile infection (CA-CDI) has been rising, due to changes in antibiotics prescribing practices, emergence of hypervirulent strains and improved diagnostics. This study explored CA-CDI epidemiology by examining strain diversity and virulence factors of CA-CDI isolates collected across several geographical regions in Israel. Methods: Stool samples of 126 CA-CDI patients were subjected to PCR and an immunoassay to identify toxin genes and proteins, respectively. Toxin loci PaLoc and PaCdt were detected by whole-genome sequencing (WGS). Biofilm production was assessed by crystal violet-based assay. Minimum inhibitory concentration was determined using the Etest technique or agar dilution. WGS and multi-locus sequence typing (MLST) were used to classify strains and investigate genetic diversity. Results: Sequence types (ST) 2 (17, 13.5%), ST42 (13, 10.3%), ST104 (10, 8%) and ST11 (9, 7.1%) were the most common. All (117, 92.8%) but ST11 belonged to Clade 1. No associations were found between ST and gender, geographic area or antibiotic susceptibility. Although all strains harbored toxins genes, 34 (27%) produced toxin A only, and 54 (42.9%) strains produced toxin B only; 38 (30.2%) produced both toxins. Most isolates were biofilm-producers (118, 93.6%), primarily weak producers (83/118, 70.3%). ST was significantly associated with both biofilm and toxin production. Conclusion: C. difficile isolates in Israel community exhibit high ST diversity, with no dominant strain. Other factors may influence the clinical outcomes of CDI such as toxin production, antibiotic resistance and biofilm production. Further studies are needed to better understand the dynamics and influence of these factors on CA-CDI.

Comparison of FT-IR with whole-genome sequencing for identification of maternal-to­neonate transmission of antibiotic-resistant bacteria.

Following a previous study in which we evaluated the carriage rates of extended spectrum ß-lactamase (ESBL) -producing Enterobacterales (ESBL+ E), carbapenem-resistant Enterobacterales (CRE), vancomycin-resistant Enterococci (VRE), and methicillin-resistant Staphylococcus aureus (MRSA) among pregnant women and their neonates, in the current study we used, for the first time, Fourier transform infrared spectrometry (FT-IR) in order to determine whether antibiotic-resistant bacteria colonization in neonates has resulted from a vertical transmission from the mothers. To this end, 28 pairs of maternal and neonatal isolates of antibiotic-resistant bacteria, including ESBL-producing E. coli (ESBL+E.coli), ESBL-producing K. pneumoniae (ESBL+K. pneumoniae) and MRSA isolates, were subjected to a FT-IR analysis to assess the similarity between maternal and new-born isolates. We compared the FT-IR analysis results with whole genome sequencing of the isolates, in order to define whether FT-IR spectroscopy can be applied for bio-typing of bacteria and for assessment of mother-to­neonate transmission. The FT-IR analysis showed that all neonatal isolates were similar to their corresponding maternal isolates and belonged to the same cluster. Alignments of the DNA sequences of the maternal and neonatal isolates pairs revealed above 99% identity, thus confirming the FT-IR results. In conclusion, FT-IR spectroscopy can be applied to monitor bacterial transmission and specifically maternal-to­neonate transmission.

Antibiotic resistance patterns of Helicobacter pylori in North Israel - A six-year study.

BACKGROUND: One main challenge in Helicobacter pylori (H. pylori) eradication is its increasing antibiotic resistance. Additionally, resistance rates vary between geographic areas and periods. However, data are limited since susceptibility testing is not routinely performed. Thus, it is valuable to gather data regarding H. pylori's resistance rates in Israel that would aid in better adjustment of treatment. MATERIALS AND METHODS: The study included 540 H. pylori isolates, recovered from gastric biopsy samples of patients who had undergone endoscopy, during 2015-2020, at the Padeh Poriya Medical Center. Antibiotic susceptibility testing to amoxicillin, clarithromycin, metronidazole, levofloxacin, rifampicin, and tetracycline was performed using the Etest technique. Data regarding participants' sex, age, and ethnic group were collected. For every antibiotic and for multi-resistance, generalized linear models were used to estimate crude and adjusted estimated differences in mean MIC and odds ratios (ORs) for every year, compared with the reference year 2015. RESULTS: The highest resistance rates were for clarithromycin and metronidazole (46.3% and 16.3%, respectively). Patients above 18 had higher resistance rate to rifampicin and multi-resistance (3.3% and 14.8%), compared with patients under 18 (0.5% and 8.4%, respectively). Resistance rates for levofloxacin, rifampicin, and multi-resistance were significantly higher among Arab patients, compared with Jewish patients. During the 6-year surveillance, a significant annual trend in MIC for metronidazole and in ORs for metronidazole, levofloxacin, and multi-resistance were observed (after adjustment). During 2020 compared with 2015, significant increased ORs were observed for levofloxacin and metronidazole [5.72 (1.03-31.84); 4.28 (1.30-14.14), respectively]. CONCLUSIONS: In light of the remarkable changes in antibiotic resistance of H. pylori during the study's period and the increasing resistance rates to various antibiotics, it is very important to continuously monitor H. pylori antibiotic susceptibly. In order to increase eradication rates of this bacterium, therapy regimes must be based on an updated antibiotic resistance data.

Assessing the impact of a positive Biofire® FilmArray® Meningitis/Encephalitis Panel result on clinical management and outcomes.

A rapid and accurate diagnosis of meningitis/encephalitis (ME) is required for early and effective intervention or adjustment of empirical treatment. This study retrospectively analyzed 485 records of patients hospitalized at the Padeh Poriya Medical Center during 2016-2020, due to a suspicion of ME. Pathogen distribution in cerebrospinal fluid samples, as determined using the BioFire® FilmArray ME Panel (MEP), is presented, as well as comparison of demographic and clinical characteristics, clinical management and outcomes of MEP+ (105) vs MEP- (380) patients. Pathogen distribution correlated with that reported in the literature, with Enterovirus (62%) being the most common causative agent. MEP+ patients were significantly younger than MEP- patients. Antibiotics use prior to lumbar puncture was significantly higher among MEP+ patients. MEP+ was associated with more frequent antibiotic change, compared to MEP-. While MEP+ contributed to early treatment adjustment or cessation, it did not necessarily impact the length of stay or patient outcomes.

Rapid SARS-CoV-2 Detection Using the Lucira™ Check It COVID-19 Test Kit.

The need for the early identification of SARS-CoV-2 has let to a quest for reliable tests that meet the standards of polymerase chain reaction (PCR) tests, on the one hand, and are low-cost, easy-to-use, and fast, on the other hand. One such test is the Lucira™ Check It COVID-19 Test kit ("Lucira") (Lucira Health, Inc., Emeryville, CA, USA), which utilizes real-time loop-mediated isothermal amplification technology, developed for at-home use. This study evaluated the clinical sensitivity and specificity of Lucira in identifying the virus in 190 nasopharyngeal samples collected between January and October 2021. Each sample was also subjected to RT-PCR. All negative RT-PCR results were paralleled by a negative Lucira result. Out of 90 participants who had a positive RT-PCR result, 82 (91.1%) tested positive by Lucira. Among the 72 symptomatic participants, 67 (93%) tested positive by Lucira. All samples with a positive RT-PCR result with a threshold cycle (Ct) > 36, yielded a negative Lucira result. In addition, a significant positive correlation was found between Ct and time-to-positivity with Lucira (R = 0.8612, p < 0.0001). The implementation of such a portable and affordable assay may aid in breaking the COVID-19 transmission chain.

High throughput optical modulation biosensing for highly sensitive and rapid detection of biomarkers.

Rapid, highly sensitive, and high-throughput detection of biomarkers at low concentrations is invaluable for early diagnosis of various diseases. In many highly sensitive immunoassays, magnetic beads are used to capture fluorescently labeled target molecules. The target molecules are then quantified by detecting the fluorescent signal from individual beads, which is time consuming and requires a complicated and expensive detection system. Here, we demonstrate a high-throughput optical modulation biosensing (ht-OMB) system, which uses a small permanent magnet to aggregate the beads into a small detection volume and eliminates background noise by steering a laser beam in and out of the cluster of beads. Shortening the aggregation, acquisition, and well-to-well scanning transition times enables reading a 96-well plate within 10 min. Using the ht-OMB system to detect human Interleukin-8, we demonstrated a limit of detection of 0.14 ng/L and a 4-log dynamic range. Testing 94 RNA extracts from 36 confirmed RT-qPCR SARS-CoV-2-positive patients (Ct≤40) and 58 confirmed RT-qPCR SARS-CoV-2-negative individuals resulted in 100% sensitivity and 100% specificity.