Nasopharyngeal Swab Samples Research Articles

Does SARS-CoV-2 infection cause persistent ocular symptoms?: A cross-sectional study after the lifting of lockdown in Chongqing, China.

To confirm whether ocular symptoms and complaints related to the COVID-19 infection persist after recovery. A cross-sectional study was conducted on people who came to the healthcare center for regular physical examinations when the lockdown lifted for nearly 1 month. Ophthalmologists performed comprehensive ocular examinations. The infection history of COVID-19 was identified by a self-reported reverse transcription-PCR (RT-PCR) test of a nasopharyngeal swab sample for SARS-CoV-2 or a novel coronavirus antigen test with self-reported typical infection symptoms. Demographic data was collected from their healthcare reports. Ocular history and ocular symptoms were collected through face-to-face interviewing. Of a total of 308 participants, 264 (85.7%) reported COVID-19 infection; 73 (27.65%) of infected persons complained of various ocular symptoms during or after infection; and only 15 (5.68%) persons reported ocular symptoms after recovery. Infection significantly increased the complaints of red eye and eye pain compared to the time before knockdown. There were no significant differences between infections and noninfectious infections in various ocular examinations at the time of examination. The duration between the day of infection onset and examination day was unrelated to all ocular examinations. COVID-19 infection can lead to some ocular symptoms, especially conjunctival congestion and ocular pain in the infective stage, but may not cause persistent ocular symptoms in about 1 month after recovery. The results of this study may help relieve public concerns about coronavirus infection in the eyes. However, more studies on various coronavirus infections, with large sample sizes, are warranted in multi-center and community-based populations.

Genetic Characterization of Wild-Type Measles Viruses Isolated in Peshawar Pakistan

Background: Measles is a highly contagious disease, caused by the Measles Virus (MV). Knowledge of genotypes prevalent in any region is important in effective measles surveillance; by mapping distribution of prevalent MV genotypes at different points of time, in a region, gives an idea about the success of measles control through vaccination programs, by documenting the interruption of transmission of the endemic viral genotype. Objectives: To detect genotypes of wild type measles virus isolated from the samples collected of measles patients in Peshawar valley of Khyber Pakhtunkhwa (KPK) province, Pakistan.Material and Methods: This was a cross-sectional descriptive study. Nasopharyngeal and/or throat swab samples were collected during a measles outbreak, from February to March 2014, from clinically diagnosed measles patients (cases who met the clinical case definition of WHO). Collection and processing were done according to the standard protocols. RNA (Ribo-Nucleic Acid) was extracted directly from samples; cDNA (Complimentary Deoxy-Ribonucleic Acid) was synthesized, amplification and sequencing of 528bp of carboxy-terminal of N gene was carried out. Sequences were analyzed and data were compared with the standard reference strains of measles virus.Results: A total of nine sequences obtained were of good quality and enough in length to meet WHO criteria for genotype assignment. It was found that all the nine strains of measles virus belonged to clade B, genotype B3 and cluster B3.1. The rest of the 13 sequences which were shorter in length for genotype assignment also showed close resemblance to Genotype B3.Conclusion: Wild type measles virus strains, isolated from the samples of measles patients, in Peshawar valley of Khyber Pakhtunkhwa province of Pakistan were found to be genotype B3. It shows that genotype B3 may be the predominate genotype prevalent in Peshawar region at the time of this study.Keywords: B3 genotype, Genotyping of measles, Measles Virus, Molecular epidemiology.

Epidemiological surveillance of respiratory viral infections in SARS-CoV-2-negative samples during COVID-19 pandemic in Iran

BackgroundTo improve the patient care, public health surveillance, and infection control, it is crucial to identify the presence and frequency of the common respiratory infections in individuals with COVID-19 symptoms but tested negative for SARS-CoV-2. This study aimed to shed light on this during the COVID-19 pandemic in Iran.MethodsIn this cross-sectional study, a total of 1,002 patients with acute respiratory infection who had negative SARS-CoV-2 test results and referred to Valfajr Health Center, the National Collaborating Laboratory of Influenza and COVID-19 National Reference Laboratory at Pasteur Institute of Iran were recruited between January 2020 and January 2022. Nasopharyngeal and oropharyngeal swab samples were collected to detect 17 common respiratory viruses via TaqMan one-step real-time multiplex PCR. Demographic and clinical data of the participants were obtained from their electronic medical records.ResultsIn total, 218 samples (21.8%) were tested positive for at least one respiratory virus infection. Most of the common investigated respiratory viruses belonged to the years 2020 and 2022. The number of investigated patients in 2021 was few, which highlights the impact of health measures following the COVID-19 pandemic in Iran. Influenza A was the most common virus (5.8%), while adenovirus had the lowest prevalence (0.1%). Although the rate of respiratory virus infection was higher in men (24%) compared to women (19.3%), this difference was not statistically significant (P = 0.069). The prevalence of respiratory viruses had an inverse association with increasing age, with the highest rate (55.6%) observed in the age group below 2 years and the lowest rate (12.7%) in those above 65 years.ConclusionOur findings underscore the significance of adopting a comprehensive approach to respiratory infections detection and management. These results can be employed for the development of syndromic surveillance systems and implementation of the effective infection control measures. Furthermore, the results contribute to better understanding of the dynamics of respiratory viruses, both during pandemic periods and in non-pandemic contexts.

Evaluation of Multiplex Rapid Antigen Tests for the Simultaneous Detection of SARS-CoV-2 and Influenza A/B Viruses.

Single-target rapid antigen tests (RATs) are commonly used to detect highly transmissible respiratory viruses (RVs), such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza viruses. The simultaneous detection of RVs presenting overlapping symptoms is vital in making appropriate decisions about treatment, isolation, and resource utilization; however, few studies have evaluated multiplex RATs for SARS-CoV-2 and other RVs. We assessed the diagnostic performance of multiplex RATs targeting both the SARS-CoV-2 and influenza A/B viruses with the GenBody Influenza/COVID-19 Ag Triple, InstaView COVID-19/Flu Ag Combo (InstaView), STANDARDTM Q COVID-19 Ag Test, and STANDARDTM Q Influenza A/B Test kits using 974 nasopharyngeal swab samples. The cycle threshold values obtained from the real-time reverse transcription polymerase chain reaction results showed higher sensitivity (72.7-100%) when the values were below, rather than above, the cut-off values. The InstaView kit exhibited significantly higher positivity rates (80.21% for SARS-CoV-2, 61.75% for influenza A, and 46.15% for influenza B) and cut-off values (25.57 for SARS-CoV-2, 21.19 for influenza A, and 22.35 for influenza B) than the other two kits, and was able to detect SARS-CoV-2 Omicron subvariants. Therefore, the InstaView kit is the best choice for routine screening for both SARS-CoV-2 and influenza A/B in local communities.

On the feasibility of Vis–NIR spectroscopy and machine learning for real time SARS-CoV-2 detection

The pandemic caused by Covid-19 is still present around the world. Despite advances in combating the disease, such as vaccine development, identifying infected individuals is still essential to optimize the control of human-to-human transmission of the virus. The main technique for detecting the virus is the RT-PCR method, which, despite its high relative cost, has a high accuracy in detecting the coronavirus. Given this, a method capable of performing the identification quickly, accurately, and inexpensively is necessary. Thus, this work aimed to analyze the feasibility of a new technique for identifying SARS-CoV-2 through the use of optical spectroscopy in the visible and near-infrared range (Vis–NIR) combined with machine learning algorithms. Spectral signals were obtained from nasopharyngeal swab samples previously analyzed using the RT-PCR method. The specimens were provided by the Molecular Diagnosis Laboratory of Covid-19 at Univasf. A total of 314 samples were analyzed, comprising 42 testing positive and 272 testing negative for Covid-19. Digital signal processing techniques, such as Savitzky–Golay filters and statistical methods were used to eliminate spurious elements from the original data and extract relevant features. Supervised machine learning algorithms such as SVM, Random Forest, and Naive Bayes classifiers were used to perform automatic sample identification. To evaluate the performance of the models, a 5-fold cross-validation technique was applied. With the proposed methodology, it was possible to achieve an accuracy of 75%, a sensitivity of 80%, and a specificity of 70%, in addition to an area under the ROC curve of 0.81, in the identification of nasopharyngeal swab samples from previously diagnosed individuals. From these results, it was possible to conclude that Vis–NIR spectroscopy is a promising, fast and relatively low cost technique to identify the SARS-CoV-2.

Assessment of some hematological factors in patients with COVID-19 and vaccinated individuals in Kirkuk city

Abstract Background: The severe acute respiratory syndrome-coronavirus-2 (SARS-COV-2), which is considered to be the main reason of coronavirus disease-2019 (COVID-19), initiates a multisystem inflammatory condition. The COVID-19 progression is thought to be influenced by hematological abnormalities. A complete blood count (CBC) is a repeatedly used, reasonably priced, and promptly performed testing technique that may be helpful in those processes. The predominance of deep vein thrombosis and pulmonary embolism is determined by d-dimer values. Objective: The aim of this study was to assess the hematological factors in patients with COVID-19 and vaccinated individuals in Kirkuk city. Subject and Methods: Samples were taken from 167 participants aged 20 to 45 years, including men and women, from December 2021 to March 2022; these samples were taken randomly from patients with COVID-19 who were hospitalized and nonhospitalized, those who had received vaccine, and a control group (healthy people who do not have diseases). Samples of nasopharyngeal swabs were also collected from outpatients infected with COVID-19, and the infection was verified using real-time polymerase chain reaction (PCR). Also, PCR was done for control group and performed CBC and d-dimer tests for all samples. Results: Concerning to hematological tests, the outcomes have revealed significant results between study groups with WBC (P < 0.0001), RBC (P < 0.0001), lymphocyte (P < 0.0001), granulocyte (P < 0.0001), HB (P < 0.0001), HCT (P < 0.0001), MCH (P < 0.0017), PLT (P < 0.0455), and d-dimer (P < 0.0001). Conclusion: Patients with COVID-19 have a reliable pattern of CBC parameter changes and normal values in the vaccinated individual.

Bacterial nasopharyngeal colonisation in children in South Africa before and during the COVID-19 pandemic: an observational study

Deployment of non-pharmaceutical interventions such as face masking and physical distancing during the COVID-19 pandemic could have altered the transmission dynamics and carriage of respiratory organisms. We evaluated colonisation with Streptococcus pneumoniae and other upper respiratory tract bacterial colonisers before and during the COVID-19 pandemic. We did two cross-sectional surveys in Soweto, South Africa from July 3 to Dec 13, 2018 (pre-COVID-19 period) and from Aug 4, 2021, to March 31, 2022 (COVID-19 period) in healthy children (aged ≤60 months) who had recorded HIV status and had not received antibiotics in the 21 days before enrolment. At enrolment, we collected nasopharyngeal swab samples from child participants. Following nucleic acid extraction, nanofluidic quantitative PCR was used to screen all samples for 92 S pneumoniae serotypes and 14 other bacteria. The primary objective was to compare the prevalence and density of pneumococcal nasopharyngeal colonisation, overall and stratified by 13-valent pneumococcal conjugate vaccine (PCV13) serotypes and non-vaccine serotypes. Secondary study objectives included a comparison of serotype-specific pneumococcal colonisation and density, as well as colonisation by the 14 other bacteria in the COVID-19 versus pre-COVID-19 period. We used an adjusted multiple logistic and linear regression model to compare the colonisation prevalence and density between study periods. We analysed nasopharyngeal swabs from 1107 children (n=571 in the pre-COVID-19 period; n=536 in the COVID-19 period). We observed no change in overall pneumococcal colonisation between periods (274 [51%] of 536 in the COVID-19 period vs 282 [49%] of 571 in the pre-COVID-19 period; adjusted odds ratio [aOR] 1·03 [95% CI 0·95-1·12]). The prevalence of PCV13 serotypes was lower in the COVID-19 than in the pre-COVID-19 period (72 [13%] vs 106 [19%]; 0·87 [0·78-0·97]), whereas the prevalence of non-typeable S pneumoniae was higher (34 [6%] vs 63 [12%]; 1·30 [1·12-1·50]). The mean log10 density for overall pneumococcal colonisation was lower in the COVID-19 period than in the pre-COVID-19 period (3·96 [95% CI 3·85-4·07] vs 4·72 [4·63-4·80] log10 genome equivalents per mL; p<0·0001). A lower density of non-vaccine serotypes (3·63 [3·51-3·74] vs 4·08 [3·95-4·22] log10 genome equivalents per mL; p<0·0001) and non-typeable S pneumoniae (3·11 [2·94-3·29] vs 4·41 [4·06-4·75] log10 genome equivalents per mL; p<0·00001) was also observed in the COVID-19 period. There was no difference in the density of PCV13 serotypes between the periods. The prevalence of colonisation during the COVID-19 versus pre-COVID-19 period was lower for non-typeable Haemophilus influenzae (280 [49%] vs 165 [31%]; aOR 0·77 [95% CI 0·71-0·84]), Moraxella catarrhalis (328 [57%] vs 242 [45%]; 0·85 [0·79-0·92]), and Neisseria lactamica (51 [9%] vs 13 [2%]; 0·64 [0·52-0·78]), but higher for Acinetobacter baumannii (34 [6%] vs 102 [19%]; 1·55 [1·35-1·77]) and Staphylococcus aureus (29 [5%] vs 52 [10%]; 1·28 [1·10-1·50]). There were variable effects on the colonisation prevalence and density of bacterial organisms during the COVID-19 compared with the pre-COVID-19 period. The lower prevalence of PCV13 serotype together with other respiratory organisms including non-typeable H influenzae and M catarrhalis could have in part contributed to a decrease in all-cause lower respiratory tract infections observed in South Africa during the initial stage of the COVID-19 pandemic. The pathophysiological mechanism for the increase in A baumannii and S aureus colonisation warrants further investigation, as does the clinical relevance of these findings. The Bill & Melinda Gates Foundation.

Nasopharyngeal Viral Load Is the Major Driver of Incident Antibody Immune Response to SARS-CoV-2 Infection.

Virologic determinants of seroconversion to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection were defined in a post hoc analysis of prospectively studied vaccine- and infection-naïve individuals at high risk for coronavirus disease 2019 (COVID-19). This phase 3 COVID-19 prevention trial (NCT04452318) with casirivimab and imdevimab was conducted in July 2020-February 2021, before widespread vaccine availability. Placebo-treated participants who were uninfected (SARS-CoV-2 quantitative reverse transcription polymerase chain reaction [RT-qPCR] negative) and seronegative were assessed weekly for 28 days (efficacy assessment period [EAP]) for COVID-19 symptoms and SARS-CoV-2 infection by RT-qPCR of nasopharyngeal swab samples and for serostatus by antinucleocapsid immunoglobulin (Ig) G. Regression-based modeling, including causal mediation analysis, estimated the effects of viral load on seroconversion. Of 157/1069 (14.7%) uninfected and seronegative (for antispike IgG, antispike IgA, and antinucleocapsid IgG) participants who became infected during the EAP, 105 (65%) seroconverted. The mean (SD) maximum viral load of seroconverters was 7.23 (1.68) log10 copies/mL vs 4.8 (2.2) log10 copies/mL in those who remained seronegative; viral loads of ∼6.0 log10 copies/mL better predicted seroconversion. The mean of the maximum viral load was 7.11 log10 copies/mL in symptomatic participants vs 5.58 log10 copies/mL in asymptomatic participants. The mean duration of detectable viral load was longer in seroconverted vs seronegative participants: 3.24 vs 1.63 weeks. Maximum SARS-CoV-2 viral load is a major driver of seroconversion and symptomatic COVID-19, with high viral loads (∼6.0 log10 copies/mL) better predicting seroconversion. Serology underestimates infection rates, incidence, and prevalence of SARS-CoV-2 infection.

Respiratory torque teno virus load at emergency department visit predicts intensive care unit admission of SARS-CoV-2 infected patients.

Accurate prediction of COVID-19 severity remains a challenge. Torque teno virus (TTV), recognized as a surrogate marker of functional immunity in solid organ transplant recipients, holds the potential for assessing infection outcomes. We investigated whether quantifying TTV in nasopharyngeal samples upon emergency department(ED) admission could serve as an early predictor of COVID-19 severity. Retrospective single-center study in the ED of Saint-Louis Hospital in Paris, France. TTV DNA was quantified in nasopharyngeal swab samples collected for SARS-CoV-2 testing. Among 295 SARS-CoV-2 infected patients, 92 returned home, 160 were admitted to medical wards, and 43 to the intensive care unit (ICU). Elevated TTV loads were observed in ICU patients (median: 3.02 log copies/mL, interquartile range [IQR]: 2.215-3.825), exceeding those in discharged (2.215, [0; 2.962]) or hospitalized patients (2.24, [0; 3.29]) (p = 0.006). Multivariate analysis identified diabetes, obesity, hepatitis, fever, dyspnea, oxygen requirement, and TTV load as predictors of ICU admission. A 2.91 log10 copies/mL TTV threshold independently predicted ICU admission. Nasopharyngeal TTV quantification in SARS-CoV-2 infected patients is linked to the likelihood of ICU admission and might reflect respiratory immunosuppression.

480. Duration of Infectious Virus Shedding by SARS-CoV-2 Omicron Variant among Immunocompromised Patients

Abstract Background Immunocompromised patients with coronavirus disease (COVID-19) shed SARS-CoV-2 RNA longer than usual, but the duration of viral shedding and criteria for nosocomial de-isolation in these patients remains unclear. Methods A prospective cohort study was performed at two tertiary medical centers in Japan during the Omicron epidemic waves from July 8, 2022, to January 30, 2023. Nasopharyngeal swab samples were serially collected from immunocompromised COVID-19 patients. We included populations with either hematological malignancies, solid tumors, autoimmune diseases, or human immunodeficiency virus infection. Patients were classified as severely or moderately immunocompromised based on the national COVID-19 guidelines in Japan. The relationship among patients' characteristics, immune status, quantitative reverse transcription PCR (qRT-PCR), and duration of infectious virus shedding between the two groups was assessed using Mann-Whitney U and Fisher's exact tests. Results Among 41 patients with 163 samples, nine patients and 47 samples were severely immunocompromised, and 32 patients and 116 samples were moderately immunocompromised. The median ages of the two groups were 70 (IQR: 56-71) and 70.5 (IQR: 60.75-75) years, respectively. In the severely immunocompromised group, 87.2% of the samples (41/47) were PCR-positive, with a median Cq value of 26.1 (IQR: 20.5-30.2), and 36.2% (17/47) were culture-positive. In the moderately immunocompromised group, 75.0% of the samples (87/116) were PCR-positive, with a median Cq value of 21.9 (IQR: 18.4-28.6), and 38.8% (45/116) were culture-positive (Table 1). Of the 62 culture-positive samples, five (8.1%) were culture-positive from day 20 onwards, all from severely immunocompromised patients (P = 0.001). In moderately immunocompromised patients, none were culture-positive on day 12 or later (Figure 1). Conclusion In COVID-19 patients, particularly those severely immunocompromised, the duration of viral shedding should be closely monitored. Further research is needed to establish a safe period for in-hospital de-isolation of immunocompromised patients. Disclosures All Authors: No reported disclosures

506. Resistance Analyses from the Remdesivir Phase 3 REDPINE Study in Participants with Severely Reduced Kidney Function Who Were Hospitalized for COVID-19

Abstract Background Remdesivir (RDV) is a broad-spectrum nucleotide analog prodrug approved for the treatment of COVID-19 in nonhospitalized and hospitalized adult and pediatric patients with clinical benefit demonstrated in multiple Phase 3 trials. Here we present SARS-CoV-2 resistance analyses from the Phase 3 REDPINE study, which demonstrated the safety of RDV in participants with severely reduced kidney function. Methods REDPINE (NCT04745351) was a double-blind, placebo-controlled trial in participants hospitalized for COVID-19 with severely reduced kidney function who were randomized 2:1 to receive RDV or placebo for 5 days. Full-genome deep sequencing of SARS-CoV-2 was performed on nasopharyngeal swab samples collected on Days 1 (baseline), 3, 5, 7, 14, 21, and/or 29. Emergent amino acid substitutions from RDV-treated participants were tested for RDV susceptibility in a SARS-CoV-2 replicon system. Results Of the 243 participants randomized and treated, 82 (RDV, 55; placebo, 27) met the resistance analysis criteria (all participants with all-cause death or invasive mechanical ventilation [IMV] through Day 29 or with SARS-CoV-2 RNA above the viral load assay lower limit of quantitation on Day 14). Sequencing data at both baseline and postbaseline timepoints were obtained from 60 participants (RDV, 41; placebo, 19). Among these, emergent amino acid substitutions in Nsp12 were observed in 8/41 (19.5%) in the RDV group and 1/19 (5.3%) in the placebo group. The proportions of participants with all-cause death or IMV through Day 29 were comparable between participants in the RDV group with emergent Nsp12 substitutions (5/8, 62.5%) or without emergent Nsp12 substitutions (30/33, 90.9%). In 4 participants in the RDV group, Nsp12 substitutions with low-level reduced susceptibility to RDV were identified in samples collected 9 days after cessation of RDV treatment (2.9- to 3.4-fold change in EC50): M794I (n = 2), C799F (n = 1), and E136V (n = 1). Three of these 4 participants had received solid organ transplants and were receiving concomitant immunosuppressive therapies during the study. Conclusion The results of resistance analyses in participants with severely reduced kidney function hospitalized for COVID-19 confirm a high barrier to clinically meaningful resistance to RDV in COVID-19 patients. Disclosures Kristen Andreatta, MS, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Jiani Li, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Lauren Rodriguez, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Dong Han, MS, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Simin Xu, MS, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Jason K. Perry, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Yiannis Koullias, MD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Robert H. Hyland, DPhil, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds Charlotte Hedskog, PhD, Gilead Sciences, Inc.: Employee|Gilead Sciences, Inc.: Stocks/Bonds

The evaluation of a rapid microfluidic immunofluorescence antigen test in detecting the infectiousness of COVID-19 patients

BackgroundA test-based strategy against coronavirus disease 2019 (COVID-19) is one of the measures to assess the need for isolation and prevention of infection. However, testing with high sensitivity methods, such as quantitative RT-PCR, leads to unnecessary isolation, whereas the lateral flow antigen test shows low sensitivity and false negative results. The purpose of this study was to evaluate the performance of the LumiraDx SARS-CoV-2 Ag test (Lumira Ag), a rapid microfluidic immunofluorescence method, in assessing infectivity.MethodsThis study was performed from March 2022 to July 2022. A pair of nasopharyngeal swab samples were obtained from each patient with mild COVID-19. One swab was used for Lumira Ag testing, and the other for quantitative RT-PCR testing and virus culture.ResultsA total of 84 patients were included in the study. Among them, PCR, Lumira Ag test, and virus culture indicated positivity for 82, 66, and 24 patients, respectively. When comparing the Lumira Ag test to virus culture, its sensitivity was 100.0% (24/24), specificity, 30.0% (18/60); positive predictive value, 36.3% (24/66); and negative predictive value (NPV), 100.0% (18/18). The positive sample for virus culture was observed until the ninth day from the onset of symptoms, while the Lumira Ag test was observed until day 11.ConclusionsThe Lumira Ag test showed high sensitivity and NPV (100% each) compared to virus culture. A test-based strategy using the Lumira Ag test can effectively exclude COVID-19 infectiousness.

Robotic system for nasopharyngeal swab sampling based on remote center of motion mechanism.

In this study, a robotic system is proposed for nasopharyngeal (NP) swab sampling with high safety and efficiency. Most existing swab-sampling robots have more than six degrees of freedom (DOFs). However, not all six DOFs are necessarily required for NP swab sampling. A high number of DOFs can cause safety problems, such as collisions between the robot and patient. We developed a new type of robot with four DOFs for NP swab sampling that consists of a two DOFs remote center of motion (RCM) mechanism, a two DOFs insertion mechanism, and a nostril support unit. With the nostril support unit, the robot no longer needs to adjust the insertion position of the swab. The proposed robot enables the insertion orientation and depth to be adjusted according to different postures or facial shapes of the subject. For intuitive and precise remote control of the robot, a dedicated master device for the RCM and a visual feedback system were developed. The effectiveness of the robotic system was demonstrated by repeatability, RCM accuracy, tracking accuracy, and in vitro phantom experiments. The average tracking error between the master device and the robot was less than 2mm. The contact force exerted on the swab prior to reaching the nasopharynx was less than 0.04 N, irrespective of the phantom's pose. This study confirmed that the RCM-based robotic system is effective and safe for NP swab sampling while using minimal DOFs.

Screening of pathogenic molecular markers of Staphylococcus aureus in children based on whole genome sequencing technology

To explore the molecular characteristics of Staphylococcus aureus (S. aureus) in children, and to compare the molecular characteristics of different types of strains (infection and colonization strains) so as to reveal pathogenic molecular markers of S. aureus. A cross-sectional study design was used to conduct nasopharyngeal swab sampling from healthy children in the community and clinical samples from infected children in the hospital. Whole genome sequencing was used to detect antibiotic resistance genes and virulence genes. A random forest method to used to screen pathogenic markers. A total of 512 S. aureus strains were detected, including 272 infection strains and 240 colonization strains. For virulence genes, the carrying rates of enterotoxin genes (seb and sep), extracellular enzyme coding genes (splA, splB, splE and edinC), leukocytotoxin genes (lukD, lukE, lukF-PV and lukS-PV) and epidermal exfoliating genes (eta and etb) in infection strains were higher than those in colonization strains. But the carrying rates of enterotoxin genes (sec, sec3, seg, seh, sei, sel, sem, sen, seo and seu) were lower in infection strains than in colonization strains (P<0.05). For antibiotic resistance genes, the carrying rates of lnuA, lnuG, aadD, tetK and dfrG were significantly higher in infection strains than in colonization strains (P<0.05). The accuracy of cross-validation of the random forest model for screening pathogenic markers of S. aureus before and after screening was 69% and 68%, respectively, and the area under the curve was 0.75 and 0.70, respectively. The random forest model finally screened out 16 pathogenic markers (sem, etb, splE, sep, ser, mecA, lnuA, sea, blaZ, cat(pC233), blaTEm-1A, aph(3')-III, ermB, ermA, ant(9)-Ia and ant(6)-Ia). The top five variables in the variable importance ranking were sem (OR=0.40), etb (OR=3.95), splE (OR=1.68), sep (OR=3.97), and ser (OR=1.68). The random forest model can screen out pathogenic markers of S. aureus and exhibits a superior predictive performance, providing genetic evidence for tracing highly pathogenic S. aureus and conducting precise targeted interventions.

Exploring Viral Metagenomics in Pediatric Patients with Acute Respiratory Infections: Unveiling Pathogens beyond SARS-CoV-2.

The emergence of SARS-CoV-2 and the subsequent pandemic have prompted extensive diagnostic and clinical efforts to mitigate viral spread. However, these strategies have largely overlooked the presence of other respiratory viruses. Acute respiratory diseases in pediatric patients can be caused by a diverse range of viral agents, and metagenomics represents a powerful tool for their characterization. This study aimed to investigate the viral abundance in pediatric patients with acute respiratory symptoms who tested negative for SARS-CoV-2 during the Omicron pandemic wave. To achieve this, viral metagenomics and next-generation sequencing were employed on 96 nasopharyngeal swab samples, which were organized into 12 pools, with each pool consisting of eight individual samples. Metagenomic analysis revealed that the most prevalent viruses associated with acute disease in pediatric patients were respiratory syncytial virus (detected in all pools) and enteroviruses, which are known to cause significant morbidity and mortality in children. Additionally, clinically significant viruses such as mumps orthorubulavirus, human metapneumovirus, influenza A, and a wide array of human herpesviruses (1, 3-7) were identified. These findings highlight the extensive potential of viral metagenomics in identifying viruses other than SARS-CoV-2 that contribute to acute infections in children. Consequently, this methodology should garner clinical attention in terms of differential diagnosis and the development of public policies to address such conditions in the global pediatric population.

169 Effects of Intranasal Prebiotics, Probiotics and Synbiotics on the Nasopharyngeal Microbiota of Feedlot Cattle

Abstract The emergence of antibiotic-resistant pathogens associated with bovine respiratory disease (BRD) presents a significant challenge to the beef industry, as antibiotic administration is commonly used to prevent and control BRD in feedlot cattle. Alternatives to antibiotics are therefore needed as part of new management strategies to reduce antibiotic use and BRD. We recently developed probiotics comprised of six Lactobacillus strains that inhibited the BRD pathogen Mannheimia haemolytica, and conferred longitudinal modulation of the nasopharyngeal microbiota in calves when a single dose was administered intranasally. In the present study, we evaluated the effects of a prebiotic (lactulose) alone and in combination with probiotic bacteria on the nasopharyngeal microbiota of feedlot calves. The probiotic bacteria consisted of 6 strains of Lactobacillus originated from the nasopharyngeal microbiota of healthy feedlot cattle. The calves were assigned to one of five treatments (n = 8 calves per treatment) and house individually: 1) Syringe-Probiotic group received intra-nasal probiotic cocktail via syringe that was attached to a 25-cm long catheter; 2) MAD-Probiotic group received an intra-nasal probiotic cocktail with a mucosal atomization device; 3) MAD-Synbiotic group received an intra-nasal probiotic cocktail containing prebiotic lactulose (0.5%, suspended in PBS) with a mucosal atomization device; 4) MAD-Prebiotic group received intra-nasal prebiotic lactulose (0.5%, suspended in PBS) with a mucosal atomization device; and 5) MAD-PBS (Control) group received only PBS with a mucosal atomization device. Utilization of a syringe versus mucosal atomization device was tested to determine if mode of probiotic bacteria delivery had an impact on the respiratory microbiota. Deep nasopharyngeal swab samples were collected on day 0 (24 h prior to treatment administration), 2 (24h post intranasal treatment), 3, 6, 9 and 14 days. Genomic DNA was extracted from the swabs and processed for nasopharyngeal microbiota characterization by sequencing 16S rRNA gene amplicon sequencing. When administered to calves intranasally, lactulose had no effect on the diversity or community structure of bacteria (PERMANOVA, P &amp;gt; 0.05) and had limited prolonged effect on specific genera. In addition, when administered as a synbiotic with a multi-strain cocktail of Lactobacillus probiotic bacteria, colonization was not enhanced. However, probiotic bacteria did affect the bacterial structure of NP bacteria (P &amp;lt; 0.05) and increased microbial network interactions. The strongest effect was observed when probiotic bacteria were administered alone, using an atomization device. Overall, this study showed that the bovine respiratory microbiota can be altered by administration of probiotics and may therefore provide new opportunities to enhance microbiome-mediated respiratory resistance against BRD pathogens. While colonization and growth of probiotic bacteria in the respiratory tract are unlikely to be modified by prebiotics delivered in PBS solution, the delivery method and time of application are important factors that need to be evaluated to have optimal efficacy of biologicals.

Diversity of upper respiratory tract pathogens in patients having flu-like symptoms during Covid-19 pandemic in a referral hospital

Background: Upper respiratory tract infections (URTIs) are caused by a wide range of viruses and bacteria, however, produce similar symptoms. Routine molecular tests are not performed and empirical use of antibiotics in treating URTIs is a major public health concern. In attempt to unveil the diversity of upper respiratory tract pathogens in the community during COVID-19 pandemic, we have screened 153 nasopharyngeal swab samples from patients having flu like symptoms. Materials and methods: We tested nasopharyngeal swabs by real-time multiplex PCR for 19 viruses and 3 bacteria using cartridge based rapid PCR platform. Results: Of 153 patients sample tested, 103 (67.32%) had a laboratory-confirmed respiratory pathogen. Of the 153 swabs tested rhinovirus/enterovirus was found 25(16.34%), influenza 18(11.77%), RSV 13(8.5%), SARS-CoV2- 15(11.11%), other coronaviruses 11(7.19%), parainfluenza 16(10.45%), human metapneumovirus 8(5.23%). Out of 18 influenza cases influenza A was 17(11.12%) and influenza B was 1(0.65%). Among 17 influenza A viruses H1N1pdm09 strain was 9(5.88%), H3 was 5(3.27%). These data shows that even in COVID-19 pandemic period rhinovirus/ enterovirus and Influenza dominated over all other respiratory viruses and as a causative agent bacteria might play very insignificant role in URTIs. Conclusion: Our data provides strong evidence against empiric antibiotic use for treating URTIs and highlights a strong need for improving the diagnostic capacity for URTIs by using more molecular testing in the country. J Bangladesh Coll Phys Surg 2023; 41(4): 256-261

Nucleocapsid single point-mutation associated with drop-out on RT-PCR assay for SARS-CoV-2 detection

BackgroundSince its beginning, the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has been a challenge for clinical and molecular diagnostics, because it has been caused by a novel viral agent. Whole-genome sequencing assisted in the characterization and classification of SARS-CoV-2, and it is an essential tool to genomic surveillance aiming to identify potentials hot spots that could impact on vaccine immune response and on virus diagnosis. We describe two cases of failure at the N2 target of the RT-PCR test Xpert® Xpress SARS-CoV-2.MethodsTotal nucleic acid from the Nasopharyngeal (NP) and oropharyngeal (OP) swab samples and cell supernatant isolates were obtained. RNA samples were submitted to random amplification. Raw sequencing data were subjected to sequence quality controls, removal of human contaminants by aligning against the HG19 reference genome, taxonomic identification of other pathogens and genome recovery through assembly and manual curation. RT-PCR test Xpert® Xpress SARS-CoV-2 was used for molecular diagnosis of SARS-CoV-2 infection, samples were tested in duplicates.ResultsWe identified 27 samples positive for SARS-CoV-2 with a nucleocapsid (N) gene drop out on Cepheid Xpert® Xpress SARS-CoV-2 assay. Sequencing of 2 of 27 samples revealed a single common mutation in the N gene C29197T, potentially involved in the failed detection of N target.ConclusionsThis study highlights the importance of genomic data to update molecular tests and vaccines.

Performance of a microfluidic immunofluorescence assay kit for equine influenza virus antigen detection

Equine influenza virus (EIV) infection is one of the most important respiratory diseases in the equine industry around the world. Rapid diagnosis, facilitated by point-of-care testing, is essential to implement movement restrictions and control disease outbreaks. This study evaluated a microfluidic immunofluorescence assay kit, which detects influenza virus and SARS-CoV-2 antigens in human specimens with a 12 min turnaround time, for its potential use in detecting EIV. The microfluidic immunofluorescence assay kit succeeded in detecting 11 EIV strains. Using the real-time reverse transcription polymerase chain reaction as a reference assay, the microfluidic immunofluorescence assay kit showed a sensitivity of 60.7% when evaluating nasopharyngeal swab samples of three horses experimentally infected with EIV. Comparing with the other two rapid antigen detection kits based on immunochromatography and silver amplification immunochromatography, the microfluidic immunofluorescence assay kit exhibited higher sensitivity than the former assay (53.6%) and the same sensitivity as the latter (60.7%). The microfluidic immunofluorescence assay kit did not detect nine non-EIV viruses including one equine coronavirus strain and seven bacteria, suggesting a high specificity for EIV antigens. Similar to other rapid antigen detection kits, the microfluidic immunofluorescence assay kit could be an effective diagnostic tool to detect EIV in the field.

Association of HLA-DRB1*11 and HLA-DRB1*12 gene polymorphism with COVID-19 in Burkina Faso

BackgroundThe clinical manifestations of coronavirus disease (COVID-19) can vary widely, ranging from asymptomatic to severe, and may be influenced by the host genetic background. The aim of the present study was to determine the frequencies of HLA-DRB1*11 and HLA-DRB1*12 allele polymorphisms and their associations with COVID-19.MethodsIn this cross-sectional study, 198 subjects were enrolled, including 150 COVID-19 positive cases and 48 subjects who tested negative for COVID-19. Participants were recruited from the emergency, intensive care, and infectious diseases departments of the Bogodogo Centre University Hospital (CHU-B) or the routine laboratory of Centre de Recherche Biomoléculaire Pietro Annigoni (CERBA). Genomic DNA was extracted from nasopharyngeal swabs samples and multiplex PCR-SSP was used to detect the HLA-DRB1*11 and HLA-DRB1*12 alleles. The study was approved by CERS (№ 2021-02-033).ResultsThe positive cases were categorized into 38 asymptomatic (CC+), 60 symptomatic (NC+), and 52 severe cases (SC+). Females were more frequent in the overall study population (53.0%, 105/198) as well as in the negative group’s CC- (68.75%, 33/48) and SC+ (57.69%, 30/52 negative groups, whereas males were more frequent in the CC+ (63.16%, 24/38) and NC+ (53.33%, 32/60) groups. The highest mean age was observed in the SC + group. A frequency of 19.19% (38/198) and 14.65% (29/198) was found for the HLA-DRB1*11 and HLA-DRB1*12 alleles, respectively. Individuals carrying the HLA-DRB1*11 allele had an approximately sixfold higher risk of asymptomatic SARS-CoV-2 infection (OR = 5.72 [1.683–19.442], p = 0.005) based on the association analysis.ConclusionsAltogether, the present study reports high frequency of HLA-DRB1*11 and HLA-DRB1*12 alleles within a population from Ouagadougou, Burkina Faso. The results suggest that individuals carrying the HLA-DRB1*11 allele are more susceptible to COVID-19 infection but may not display symptoms.