bluuebunny/crossref_metadata_2025_split huggingface dataset:
A-233 Wastewater-based surveillance for SARS-CoV-2 Subvariants using the Nanopore Sequencing in South Korea
Authors: Kim Y , Noh T , Lee S , Kim Y , Lee K 
Published: October 2024

Abstract

Background
Genomic tools to identify and characterize SARS-CoV-2 and its various subvariants were adapted from clinical settings. Wastewater-based surveillance (WBS) has since been recognized as tool of transformative potential, providing real-time objective, comprehensive data on the prevalence and distribution of the target pathogen across the space and time. We conducted wastewater-based epidemiology (WBE) at sewage treatment plants to find a more efficient mutation monitoring method, and respond to pre-existing infectious diseases.

Methods
In our study, domestic sewage collected every weeks from 6 sewage treatment plants in urban region from June 2023 to November 2023 was used to concentrate and isolate viruses using wastewater ultra nucleic acid isolation kits with virus enrichment kits. Positive samples confirmed were subjected to SARS-CoV-2 whole genome sequencing (WGS) using GridION (Oxford Nanopore Technologies). The experimental data was aligned through Epi2me, analyzed using Nextclade, that was compared with KCDC official reports and our monitoring results (total 780 cases) ongoing at SCL.

Results
Various mutations were identified at each collection time, occurring new subvariants over time. The mutations (orf1a:A690V, S:F456L) sequentially confirmed from the sample collected on Aug. 23rd are specific mutations of XBB.1.9.2 and EG.5. In addition, the mutations orf1b: D54N, S:L455F, which were sequentially confirmed from the sample collected on Sep. 6th, are HK.3, a descendent lineage of EG.5, and FLip variants, which show growth advantage, were confirmed. Furthermore, the detection rate of FLip variants detected from Sep.6th gradually increased from 17% to 67% (Fig.1). Taken together, we confirmed the similarity between monthly detected mutations and changes in dominant species over time by comparing with ‘monthly COVID-19 mutation trends’ and ‘HK.3 dominance announcement by KCDC’(Fig.2).

Conclusions
WBS would be a useful tool for proactively responding to infectious diseases including COVID-19 by exploring new mutations and predicting changes in dominant species.
A-238 Lyophilized Bead Multiplex Assays for CT/NG/TV Detection
Authors: Moore R , Tang N , Tran E , Fisher T , Kadam R , Wang J , Farmer A 
Published: October 2024

Abstract

Background
Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), and Trichomonas vaginalis (TV) cause the three most prevalent sexually transmitted infectious diseases in developed countries, contributing to approximately 300 million new infections worldwide, annually. Due to the challenges of culturing and using immunoassays to detect these pathogens, DNA-based molecular tests have been instrumental, and current trends in molecular testing for CT/NG/TV have moved toward point-of-care tests.
Lyophilization provides benefits required for point-of-care tests, such as ambient storage, extended shelf life, simplified downstream workflow, and variable formats. A lyophilized-bead format enables an assay to be placed in sealed consumables and stored at room temperature for up to two years.
An additional consideration is the need for endogenous and exogenous controls. Exogenous in-process controls can be used to validate the performance of workflows culminating in a qPCR reaction and provide flexibility to test developers. An ideal exogenous in-process control is nonhomologous to human or human pathogen genome sequences and does not align with primers or probes designed for pathogen detection.
The objective of the current study was to develop a lyophilized-bead-format assay containing an ideal exogenous control and establish whether it is a valid option for the accurate detection of CT, NG, and TV.

Methods
A synthetic DNA control containing targets for CT/NG/TV/RNase P (RP)/exogenous control was used to establish an analytical LOD by qPCR. The 5-plex assay, lyophilized in beads, was hydrated and tested for linearity over an 8-log range of the synthetic DNA control. Further, the qPCR lyophilized bead with assay was tested with a proficiency sample by extracting DNA from ZeptoMetrix NATtrol CT/NG/TV Positive Control and spiking-in the exogenous DNA control. qPCR was performed on the extracted DNA and human cDNA.

Results
The analytical LOD for the control DNA was determined to be 5 copies/reaction. Additionally, linearity was observed by qPCR over an 8-log range of the synthetic DNA control for all five targets in multiplex with good efficiencies and R2 values. Finally, all targets were amplified and detected by qPCR using DNA extracted from ZeptoMetrix NATtrol CT/NG/TV Positive Control with spiked-in exogenous DNA control. The detection of the exogenous sequence in the qPCR multiplex reaction indicated the extraction was successful.

Conclusions
We have developed a multiplex qPCR assay in lyophilized-bead format that can identify infectious agents, such as CT/NG/TV, in a single reaction. We have included RP as a human host control, plus an exogenous in-process control, for a total of 5-plex in separate fluorescent channels. We have shown that our lyophilized beads with assay produce a positive qPCR result when combined with a proficiency sample.
A-240 Validation of the OSOM Flu SARS-CoV-2 Combo Home Test
Authors: Yaji S , Shimizu K , Fong D , Gonzalez E , Aihara T , Delgado Flores N , Dennis K , Nguyen-Brown T , Cox K , Baytan A , Schenken J , Morita M , Chun A , Nair-Schaef D 
Published: October 2024

Abstract

Background
COVID-19 and Influenza respiratory infections often cause similar symptoms, however, treatment is different for each infection. Delay in diagnosis can be fatal in some cases, therefore rapid detection and differentiation are critical for appropriate medical treatment. The OSOM Flu SARS-CoV-2 Combo Home Test is a lateral flow immunoassay intended for the qualitative and differential detection of nucleocapsid protein from SARS-CoV-2 and influenza A/B proteins using self-collected anterior nasal swab samples (self-collection is only recommended for those 14 years of age and older). An interpretation card, included with the test, is used to reliably distinguish among possible testing results.

Methods
A prospective study was conducted at multiple geographically distinct sites in the U.S during the 2023-2024 season for the validation of the OSOM Flu SARS-CoV-2 Combo Home Test to detect SARS-CoV-2/Flu A/Flu B in self-collected anterior nasal swab samples. The study evaluated the performance in symptomatic individuals only. A total of 703 prospective samples were evaluable for SARS. For Flu A/B, four additional subject samples were unevaluable due to comparator results not being available and a total of 699 samples were included in the data analysis. The comparator tests were FDA 510(k)-cleared molecular assays.

Results
Influenza A was detected with a sensitivity of 93.1% (95% CI: 84.8%-97.0%) and a specificity of 99.5% (95% CI: 98.6% - 99.8%). Influenza B was detected with a sensitivity of 89.1% (95% CI: 77.0% - 95.3%) and a specificity of 99.7% (95% CI: 98.9% - 99.9%). SARS CoV-2 was detected with a sensitivity of 87.0% (95% CI 78.6% - 92.4%) and a specificity of 99.1% (95% CI: 97.9% - 99.6%) when Ct values are standardized and 10% of samples are considered low positives, having a Ct value of ≥ 30.

Conclusions
With respiratory infections often showing indistinguishable symptoms and the possibility of co infection, lowering the testing hurdles for individuals to be able to perform a rapid differential test at home without the aid from a healthcare worker is the most effective way to mitigate spread and patient management from a public health perspective. Comparing the OSOM Flu SARS-CoV-2 Combo Home test to the FDA cleared comparators in a clinical setting, the OSOM Flu SARS-CoV-2 Combo Home test retains acceptable sensitivity and specificity. Using self-collected anterior nasal swabs which are minimally invasive alongside the low cost of a lateral flow immunochromatographic assay remains the most effective way to perform the serial screening testing recommended by the CDC in situations where transmission and severe outcome risk is high.
A-242 Assessing the efficacy of the one-step diagnostic algorithm for Hepatitis C
Authors: Bérgamo-Vázquez S , González-García L , González-Fernández C , Rives-Jimenez J , Prado-Ramos M , Guzmán-Olivares B , Enriquez-Marroquin C , Huerta-Velez A , Lao-López C , Álvarez-González M , Trapé-Pujol J 
Published: October 2024

Abstract

Background
Hepatitis C virus (HCV) infection can be asymptomatic and may go into spontaneous remission. However, in 60-80% of cases, it becomes chronic and progresses into cirrhosis or hepatocarcinoma. The virus is mainly transmitted parenterally, although cases of sexual and transplacental transmission have also been found. Since the development of direct-acting antiviral (DAA) therapy that can achieve a complete cure in 95% of patients, eradication strategies based on rapid diagnosis and treatment have been promoted. Therefore, the World Health Organization (WHO) has set a goal to eliminate HCV infection as public health threat by 2030. This is a retrospective study on the usefulness and effectiveness of the one-step HCV diagnostic algorithm application from May 2022 to December 2023 in our hospital.

Methods
The current HCV testing algorithm consists in a reflex HCV RNA testing, in which anti-HCV antibody-positive samples are automatically referred for HCV RNA testing on the same specimen. We used a serological electrochemiluminescence (ECLIA) assay for the detection of anti-HCV antibodies (Cobas e801, Roche Diagnostics®) in serum samples; and then a confirmation of HCV viremia by nucleic acid testing of HCV RNA in plasma samples (Cobas 4800, Roche Diagnostics®).

Results
A total of 22713 HCV serologies were performed with a total of 221 positive results (1%). Of the HCV cases, 21 patients had a newly diagnosis of active infection (10%) for whom we recommended derivation to the Digestive Service to initiate elimination treatment. 71% were men, with a mean age of 53 years and a standard deviation of 17. The mean age of women was 56 years with a standard deviation of 18. Coinfection with HIV or hepatitis B virus (HBV) was also assessed. Patients with HIV-HCV coinfection represented 9% of cases and were on average 12 years younger than HCV-monoinfected individuals. We did not identify any case of HBV-HCV co-infection.

Conclusions
The pattern we observed in terms of incidence by age and sex is similar to that reported in the literature, as well as the incidence of new HCV cases. One-step diagnosis enables the laboratory to identify the need for referring to a specialist, reducing the time to treatment for new and known but untreated cases. Therefore, global access to HCV diagnostics will be a keystone to success.
A-243 Quantitative Bacteriology Parameters for Seal-Packaged Agar Plates Challenged with Experimental Low-resource Tropical Field Clinic Stress
Authors: Font B K , Vaughn P 
Published: October 2024

Abstract

Background
InTray® Mueller Hinton Chocolate (MHC) and Colorex™ Screen (CS) plates are in vitro diagnostic devices combining prepared bacteriological agar-media with Biomed’s ‘InTray®’ format (a double sealed-packaging cassette system that provides extended shelf-life 12 months past the date of manufacture). The devices are intended for use with clinical specimens to culture, isolate and identify potentially pathogenic bacteria. This study evaluated MHC and CS device performance at 7 and 13 months past the date of manufacture after product was experimentally challenged with stress replicating a low-resource tropical field clinic scenario.

Methods
MHC and CS plates were exposed to high temperature (40.5°C ± 0.5) and high humidity (74.5 ± 2.5% relative humidity) for an extended time-period (18 days). After the 18 days of heat treatment, the stress-group plates were stored in the lab at working room temperature (20-25°C) for the remainder of the study. Quantitative differences between control-groups (stored under refrigeration: 2-8°C) and heat-stress treatment groups for both MHC and CS were examined via CFU/mL measurements comparing three different lab-strain bacteria chosen based on clinical sub-culturing prevalence from blood bottles; Escherichia coli (ATCC 25922), Streptococcus pneumoniae (ATCC 6305), and Staphylococcus aureus (ATCC 25923). Briefly, 24-hour (18-30 h) old bacterial colony suspensions (grown on MHC) were prepared in 0.85% saline solution, adjusted to a 0.5 McFarland standard (here defined as 0.08-0.96 Absorbance @ 625 nm), measured in triplicate and diluted. A 20 μL volume from the lowest dilution (2.5 x 103 CFU/mL) was then spread-plated to each plate in order to count colony-forming units (CFU) in the acceptable range of 25-250.

Results
Quantitative CFU/mL differences between control-groups (stored under refrigeration: 2-8°C) and heat-stress-treatment groups (&gt;40°C and high humidity) for MHC and CS for three different test bacteria (listed above) were not statistically significant when tested at 13 months. The CS stress and control group comparison for S. pneumoniae with a p value &lt; 0.05 at 7 months had a p value &gt; 0.05 at 13 months, suggesting that the difference between the CFU counts may not be significant in biological terms.

Conclusions
In conclusion, control and heat-stress treatment groups for both MHC and CS met acceptable performance criteria as expected for the products, 13 months after the date of manufacture.
A-250 Evaluation of Liver Fibrosis in Schistosomiasis Mansoni Using the ELF (Enhanced Liver Fibrosis) Score
Authors: Ozaki B C , Thorp R D , Batista A D , d. Mariz C , Diniz G T , Barreto A V , de Morais C N , Domingues A C , Lopes E P 
Published: October 2024

Abstract

Background
Measuring the presence and pattern of periportal fibrosis (PPF) in schistosomiasis mansoni (SM) is essential for diagnosing its clinical forms and detection of portal hypertension. For these evaluations, the gold-standard has been ultrasound scan (US) using Niamey classification, however it requires sophisticated equipment and well-trained operator. ELF (Enhanced Liver Fibrosis) is an index capable of quantifying liver fibrosis in chronic liver diseases, although there have been few studies evaluating PPF in SM.

Aim
To assess serum ELF levels according to the PPF patterns established by ultrasound using the Niamey classification in patients with schistosomiasis.

Methods
A cross-sectional, bidirectional analytical study was carried out with patients from an endemic area (patterns A, B, C and D) and from an SM reference outpatient clinic (patterns D, E and F). Clinical data and 10 mL of blood were collected, centrifuged and frozen to measure and calculate the score by the ELF™ Test (Atellica® IM), according to the equation: [ELF = 2.278 + 0.851 ln(HA) + 0.751 ln(PIIINP) + 0.394 ln(TIMP-1)] and expressed as a numerical value without unit. US using GE Healthcare Logic E (endemic area) or Siemens Acuson S2000 (outpatient clinic) equipment were performed by a single examiner.

Results
Of the 196 patients evaluated, 114 (58.2%) were women with a mean age of 45 ± 15.42 years. They had the following PPF patterns: 18 pattern A+B; 64 pattern C; 56 pattern D; 49 pattern E and 9 pattern F. The median ELF values were higher according to PPF progression in these 5 groups (p = 0.0021), respectively: (A+B = 8.36); (C = 8.40); (D = 8.64); (E = 8.99) and (F = 9.75). The ELF was able to differentiate patients without PPF, with mild, moderate and advanced PPF: A+B vs C vs D vs E+F (p = 0.0024), as well as patients with advanced PPF from those without fibrosis or with moderate fibrosis: A+B vs E+F (p = 0.0464) and C+D vs E+F (p = 0.0007). The index was also able to differentiate patients with absent or mild PPF from those with moderate or advanced PPF: A+B+C vs D+E+F (p = 0.0015).

Conclusions
The ELF showed great potential as a tool for assessing PPF in patients with schistosomiasis mansoni, being able to differentiate those with little fibrosis from those with moderate or advanced fibrosis.
A-251 Simplification of Molecular Diagnostics for Sexually Transmitted Infection Point-of-Care Testing
Authors: Seely S O , Roberts J , Collins D , Zingg J , Deo S , Daunert S 
Published: October 2024

Abstract

Background
HPV-caused cancers contribute to a large disease burden, accounting for 5% of all cancers. It is estimated that over 700,000 people develop HPV-related cancers each year with the majority being cervical cancer. The burden of this disease is disproportionately carried by low-income countries with the highest rates of HPV cervical infections primarily found in sub-Saharan Africa, Latin America, Eastern Europe, and South-East Asia. Many regions in low-resource countries lack adequate access to sensitive point-of-care screening tools, preventing timely diagnosis and treatment. Current low-cost methods, such as Pap smears and visual inspection using acetic acid suffer from low sensitivity, specificity, and reproducibility which limit their effectiveness as a screening modality. Many countries use nucleic acid amplification tests (NAATs) to identify key HPV infections which may progress towards cervical cancer. However, the expenditure for NAAT-capable equipment and reagents are cost-prohibitive for rural and underserved communities. To reduce screening barriers, we developed an isothermal HPV molecular test that can detect all 14 cancer-causing HPV strains using a novel amplification methodology combined with G-quadruplex DNAzyme colorimetric detection.

Methods
We developed a novel isothermal amplification technique that is related to loop-mediated isothermal amplification (LAMP) but uses modified looped primers to enable exponential amplification at 60°C for 1 hour. The modified primers only require two binding sites compared to 4-6 in LAMP. To detect the amplified products, we used G-quadruplex peroxidase-like DNAzyme probes which reduce ABTS to produce a visually interpreted result. Analytical validation evaluated the method’s limit of detection, specificity, and accuracy. As proof of concept in a relevant matrix, clinical cervical brushings collected ThinPrep Pap media were tested with the new method and compared to an FDA-approved reference qPCR test.

Results
Repeated experiments demonstrate that the lowest detectable HPV DNA concentration was approximately 1000 genomic copies/µL with no amplification with other pathogens. As proof of concept, the comparison to the reference standard demonstrated high accuracy.

Conclusions
The application of our assay is intended as a near-patient screening tool with further evaluation by a clinician for confirmation. With an accurate and rapid screening tool, we envision low-resource regions across the world will have the tools to rapidly detect and treat cervical cancers in their early stages.
A-271 Expansion of sample viability for self-collected urine specimens containing CT/NG & TV/MG utilizing the Roche cobas® 6800 System
Authors: Muldrow R , Sicari B , Caro Azoy E , Beal S 
Published: October 2024

Abstract

Background
The CDC estimates that 1 in 5 Americans have an STI. Home collection for STIs could help mitigate the stigma, cost, and accessibility, which contribute to low diagnosis rates and disparities in diagnosis and treatment. Home self-collected specimens occasionally arrive in the lab with volumes higher or lower than what is required by the assay manufacturer’s specifications for self-collection. This study demonstrates that sample stability is retained after temperature fluctuation via shipping - even when the correct ratio of buffer to sample has been disturbed, a precise and accurate measurement persists. Our goal is to improve accessibility to discreet, patient collected STI tests.

Methods
Positive samples were prepared by spiking negative urine with organisms near the manufacturer’s limit of detection (LoD). Properly collected samples were aliquoted into tubes with 5mL volumes as per manufacturer specifications. Improperly collected samples were either overfilled (9mL for CT/NG, 7mL for TV/MG) or underfilled (1mL for CT/NG, 2mL for TV/MG). Samples underwent temperature cycling to mimic summer or winter shipping conditions. Ambient temperature studies were conducted on over/underfilled specimens. All samples were baseline tested and compared qualitatively to the manufacturer's claimed LoD using the Roche c6800 and the IVD assays for CT/NG & TV/MG.

Results
Improperly filled samples were stable at ambient temperature for 63 days for CT/NG, and 59 days for TV/MG. See chart for temperature cycling results. All samples (n=164) showed 100% qualitative agreement with their baseline result after summer and winter shipping challenges.

Conclusions
Urine samples that fail to satisfy the manufacturer’s self-collection specifications or have endured temperature fluctuations still yield accurate CT/NG & TV/MG results. These results indicate the potential to enhance sample viability and patient accessibility utilizing validated IVD assays.
A-276 Unraveling Pathogen Dynamics In Pediatric Meningitis Complications During The Omicron Variant Outbreak In Taiwan
Authors: Huang C , Lin Y , Hsieh T , Tsao K , Shih S , Chiu C 
Published: October 2024

Abstract

Background
Taiwan, once celebrated for its remarkable control over COVID-19, faced an unexpected surge in pediatric cases during the first wave of the omicron variant in April 2022. Alarming data revealed acute encephalitis emerging in half of the severe or fatal COVID-19 cases in children within the initial two months of the omicron epidemic. This study delves into the intricate microbiota patterns and viral pathogens contributing to encephalitis complications in affected children.

Methods
The study enrolled 36 pediatric patients, comprising 11 patients with severe encephalitis, 12 patients with mild encephalitis, and 13 normal controls. Nasal swabs were collected for pathogen detection. For respiratory microbiota analysis, 16S rRNA metagenomic sequencing was employed, while the Illumina Respiratory Pathogen ID/AMR Enrichment Panel was used for respiratory pathogen identification.

Results
Microbiota analysis of nasal swabs from COVID-19 pediatric patients with meningitis complications revealed prevalent Corynebacterium, Staphylococcus, Dolosigranulum, Acinetobacter, Moraxella, and Streptococcus. Conversely. No predominant bacterial species were detected in the normal control group. Nasal swab respiratory pathogen detection identified coinfections, particularly Herpesviridae viruses such as human herpesvirus 6, Epstein-Barr virus, and Cytomegalovirus in pediatric patients with meningitis complications. Notably, two severe encephalitis cases exhibited coinfections involving rhinovirus, SARS-CoV-2, and Herpesviridae viruses (HHV-6, EBV). In one fatal case, distribution of Streptococcus genes among total geneomes reaches 40.69%, 7 drug resistant genes were detected.

Conclusions
This study suggests a potential link between latent Herpesviridae virus coinfections (HHV-6, CMV, EBV) and meningitis complications in pediatric patients. Furthermore, the presence of a secondary respiratory virus (rhinovirus) coinfection appears to be associated with severe complications. These findings shed light on the complex pathogen dynamics contributing to the emergence of encephalitis in pediatric COVID-19 cases during the omicron variant outbreak in Taiwan.