RESUMO

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) detection typically relies on reverse transcription-polymerase chain reaction (RT-PCR) technology. However, there is a certain rate of missed detection of SARS-CoV-2 in nasopharyngeal samples, particularly among immunosuppressed individuals. METHODS: In this case, SARS-CoV-2 was detected in nasopharyngeal swabs and bronchoalveolar lavage fluid (BALF) using RT-PCR. Pulmonary imaging was performed using computed tomography (CT). Patient clinical data were retrieved from the Laboratory Information System (LIS). RESULTS: SARS-CoV-2 was negative in two nasopharyngeal tests of the patient, but was finally detected in BALF, confirming that the lung lesions were infected by SARS-CoV-2. CONCLUSIONS: In the post-epidemic era, it is necessary to use BALF to identify SARS-CoV-2 infection in cases where other factors have been ruled out in immunosuppressed individuals with pulmonary infections, especially when the nasopharyngeal test yields a negative result.

Assuntos

Líquido da Lavagem Broncoalveolar , COVID-19 , Nasofaringe , SARS-CoV-2 , Humanos , Líquido da Lavagem Broncoalveolar/virologia , COVID-19/diagnóstico , COVID-19/virologia , Nasofaringe/virologia , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/genética , Tomografia Computadorizada por Raios X , Masculino , Teste de Ácido Nucleico para COVID-19 , Pessoa de Meia-Idade , Feminino , Hospedeiro Imunocomprometido

RESUMO

BACKGROUND: The association of the oral microbiome with SARS-CoV-2 infections and disease progression has been documented in European, Asian, and American populations but not in Africa. METHODS: We conducted a study in Ghana to evaluate and compare the naso-oropharyngeal microbiome in SARS-CoV-2-infected and uninfected persons before (pre-vaccine) and after vaccine availability (post-vaccine) in the country. 16S rRNA V3-V4 variable region was sequenced and analysed from DNA extracted from naso-oropharyngeal swabs. RESULTS: Considering only the infection status, infected and uninfected groups had no difference in their within-group diversity and was evident in the study population pre- and post-vaccine availability. The introduction of vaccines reduced the diversity of the naso-oropharyngeal microbiome particularly among SARS-CoV-2 positive persons and, vaccinated individuals (both infected and uninfected) had higher microbial diversity compared to their unvaccinated counterparts. SARS-CoV-2-positive and -negative individuals were largely compositionally similar varying by 4-7% but considering vaccination*infection statuses, the genetic distance increased to 12% (P = 0.003) and was mainly influenced by vaccination. Common among the pre- and post-vaccine samples, Atopobium and Finegoldia were abundant in infected and uninfected individuals, respectively. Bacteria belonging to major butyrate-producing phyla, Bacillota (particularly class Clostridia) and Bacteroidota showed increased abundance more strikingly in infected individuals before vaccines were available. They reduced significantly after vaccines were introduced into the country with Fusobacterium and Lachnoanaerobaculum being the only common bacteria between pre-vaccine infected persons and vaccinated individuals, suggesting that natural infection and vaccination correlate with high abundance of short-chain fatty acids. CONCLUSION: Our results show, in an African cohort, the abundance of bacteria taxa known for their protective pathophysiological processes, especially during infection, suggesting that this population is protected against severe COVID-19. The immune-related roles of the members of Bacillota and Bacteroidota that were found associated with infection and vaccination require further studies, and how these may be linked to ethnicity, diet and age. We also recommend expansion of microbiome-disease association studies across Africa to identify possible bacterial-mediated therapeutics for emerging infections.

Assuntos

Bactérias , COVID-19 , Orofaringe , SARS-CoV-2 , Humanos , COVID-19/microbiologia , Masculino , Adulto , Feminino , SARS-CoV-2/genética , Gana/epidemiologia , Pessoa de Meia-Idade , Bactérias/classificação , Bactérias/genética , Bactérias/isolamento & purificação , Orofaringe/microbiologia , Orofaringe/virologia , Butiratos/metabolismo , Microbiota , RNA Ribossômico 16S/genética , Índice de Gravidade de Doença , Vacinas contra COVID-19 , Nasofaringe/microbiologia , Nasofaringe/virologia , Idoso , Adulto Jovem

RESUMO

Oral fluids provide ready detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and host responses. This study sought to evaluate relationships between oral virus, oral and systemic anti-SARS-CoV-2-specific antibodies, and symptoms. Oral fluids (saliva/throat wash (saliva/TW)) and serum were collected from asymptomatic and symptomatic, nasopharyngeal (NP) SARS-CoV-2 RT-qPCR+ human participants (n = 45). SARS-CoV-2 RT-qPCR and N-antigen detection by immunoblot and lateral flow assay (LFA) were performed. RT-qPCR for subgenomic RNA (sgRNA) was sequence confirmed. SARS-CoV-2-anti-S protein RBD LFA and ELISA assessed IgM and IgG responses. Structural analysis identified host salivary molecules analogous to SARS-CoV-2-N-antigen. At time of enrollment (baseline, BL), LFA-detected N-antigen in 86% of TW and was immunoblot-confirmed. However, only 3/17 were saliva/TW qPCR+ . Sixty percent of saliva and 83% of TW demonstrated persistent N-antigen at 4 weeks. N-antigen LFA signal in three anti-spike sero-negative participants suggested potential cross-detection of 4 structurally analogous salivary RNA binding proteins (alignment 19-29aa, RMSD 1-1.5 Angstroms). At enrollment, symptomatic participants demonstrated replication-associated sgRNA junctions, were IgG+ (94%/100% in saliva/TW), and IgM+ (63%/54%). At 4 weeks, SARS-CoV-2 IgG (100%/83%) and IgM (80%/67%) persisted. Oral and serum IgG correlated 100% with NP+ PCR status. Cough and fatigue severity (p = 0.010 and 0.018 respectively), and presence of weakness, nausea, and composite upper respiratory symptoms (p = 0.037, 0.005, and 0.017, respectively) were negatively associated with saliva IgM but not TW or serum IgM. Throat wash IgM levels were higher in women compared to men, although the association did not reach statistical significance (median: 290 (female) versus 0.697, p = 0.056). Important to transmission and disease course, oral viral replication and persistence showed clear relationships with select symptoms and early oral IgM responses during early infection. N-antigen cross-reactivity may reflect mimicry of structurally analogous host proteins.

Assuntos

Anticorpos Antivirais , COVID-19 , SARS-CoV-2 , Saliva , Humanos , COVID-19/imunologia , COVID-19/virologia , COVID-19/diagnóstico , SARS-CoV-2/imunologia , Saliva/virologia , Saliva/imunologia , Feminino , Masculino , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Adulto , Pessoa de Meia-Idade , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Imunoglobulina M/imunologia , Fosfoproteínas/imunologia , RNA Viral , Nasofaringe/virologia , Proteínas do Nucleocapsídeo de Coronavírus/imunologia , Idoso

RESUMO

BACKGROUND: Adults living with human immunodeficiency virus (ALWHIV) receiving antiretroviral therapy (ART) exhibit higher pneumococcal carriage prevalence than adults without HIV (HIV-). To assess factors influencing high pneumococcal carriage in ALWHIV, we estimated pneumococcal carriage acquisition and clearance rates in a high transmission and disease-burdened setting at least 10 years after introducing infant PCV13 in routine immunisation. METHODS: We collected longitudinal nasopharyngeal swabs from individuals aged 18-45 in Blantyre, Malawi. The study group included both HIV- individuals and those living with HIV, categorised based on ART duration as either exceeding 1 year (ART > 1y) or less than 3 months (ART < 3 m). Samples were collected at baseline and then weekly for 16 visits. To detect pneumococcal carriage, we used classical culture microbiology, and to determine pneumococcal serotypes, we used latex agglutination. We modelled trajectories of serotype colonisation using multi-state Markov models to capture pneumococcal carriage dynamics, adjusting for age, sex, number of under 5 year old (< 5y) children, social economic status (SES), and seasonality. RESULTS: We enrolled 195 adults, 65 adults in each of the study groups. 51.8% were females, 25.6% lived with more than one child under 5 years old, and 41.6% lived in low socioeconomic areas. The median age was 33 years (IQR 25-37 years). The baseline pneumococcal carriage prevalence of all serotypes was 31.3%, with non-PCV13 serotypes (NVT) at 26.2% and PCV13 serotypes (VT) at 5.1%. In a multivariate longitudinal analysis, pneumococcal carriage acquisition was higher in females than males (hazard ratio [HR], NVT [1.53]; VT [1.96]). It was also higher in low than high SES (NVT [1.38]; VT [2.06]), in adults living with 2 + than 1 child < 5y (VT [1.78]), and in ALWHIV on ART > 1y than HIV- adults (NVT [1.43]). Moreover, ALWHIV on ART > 1y cleared pneumococci slower than HIV- adults ([0.65]). Residual VT 19F and 3 were highly acquired, although NVT remained dominant. CONCLUSIONS: The disproportionately high point prevalence of pneumococcal carriage in ALWHIV on ART > 1y is likely due to impaired nasopharyngeal clearance, which results in prolonged carriage. Our findings provide baseline estimates for comparing pneumococcal carriage dynamics after implementing new PCV strategies in ALWHIV.

Assuntos

Portador Sadio , Infecções por HIV , Nasofaringe , Infecções Pneumocócicas , Vacinas Pneumocócicas , Streptococcus pneumoniae , Humanos , Malaui/epidemiologia , Feminino , Adulto , Infecções por HIV/epidemiologia , Masculino , Vacinas Pneumocócicas/administração & dosagem , Infecções Pneumocócicas/epidemiologia , Infecções Pneumocócicas/prevenção & controle , Portador Sadio/epidemiologia , Portador Sadio/microbiologia , Streptococcus pneumoniae/isolamento & purificação , Adulto Jovem , Pessoa de Meia-Idade , Adolescente , Nasofaringe/microbiologia , Nasofaringe/virologia , Lactente , Vacinas Conjugadas/administração & dosagem , Estudos Longitudinais

RESUMO

At the beginning of the COVID-19 pandemic, the Georgia Institute of Technology made the decision to keep the university doors open for on-campus attendance. To manage COVID-19 infection rates, internal resources were applied to develop and implement a mass asymptomatic surveillance program. The objective was to identify infections early for proper follow-on verification testing, contact tracing, and quarantine/isolation as needed. Program success depended on frequent and voluntary sample collection from over 40,000 students, faculty, and staff personnel. At that time, the nasopharyngeal (NP) swab, not saliva, was the main accepted sample type for COVID-19 testing. However, due to collection discomfort and the inability to be self-collected, the NP swab was not feasible for voluntary and frequent self-collection. Therefore, saliva was selected as the clinical sample type and validated. A saliva collection kit and a sample processing and analysis workflow were developed. The results of a clinical sample-type comparison study between co-collected and matched NP swabs and saliva samples showed 96.7% positive agreement and 100% negative agreement. During the Fall 2020 and Spring 2021 semesters, 319,988 samples were collected and tested. The program resulted in maintaining a low overall mean positivity rate of 0.78% and 0.54% for the Fall 2020 and Spring 2021 semesters, respectively. For this high-throughput asymptomatic COVID-19 screening application, saliva was an exceptionally good sample type.

Assuntos

COVID-19 , Nasofaringe , SARS-CoV-2 , Saliva , Manejo de Espécimes , Humanos , COVID-19/diagnóstico , COVID-19/epidemiologia , Saliva/virologia , Manejo de Espécimes/métodos , SARS-CoV-2/isolamento & purificação , Universidades , Nasofaringe/virologia , Teste para COVID-19/métodos , Georgia/epidemiologia

RESUMO

Immune responses of the epithelia of the upper respiratory tract are likely crucial in early inhibition of the viral replication and finally clearance of SARS-CoV-2. We aimed to compare the expression profiles of antimicrobial peptides/proteins (AMPs) and related cytokines observed in the nasopharynx of SARS-CoV-2-infected patients and non-infected controls and to assess the associations between these parameters and COVID-19 patients' outcomes. We included 45 subjects who had tested positive for SARS-CoV-2 and 22 control subjects who had tested negative for SARS-CoV-2. Biomaterial for SARS-CoV-2 detection, as well as gene and protein expression studies, was obtained from all subjects using nasopharyngeal swabs which were performed a maximum of 7 days before inclusion in the study. Univariable and multivariable statistics were performed. When compared to the controls, the mRNA expression levels of human ß-defensin 1 (hBD-1), LL-37, and trappin-2 were significantly higher in specimens of nasopharyngeal swabs from COVID-19 patients. Protein expression of hBD-1 was also increased in the COVID-19 group. mRNA expression levels of interferon-É£ (IFN-É£), tumor necrosis factor- ɑ (TNF-ɑ), and interleukin-6 (IL-6) measured in SARS-CoV-2-infected patients were significantly higher than those observed in the controls, which could also be confirmed in the protein levels of IFN-É£ and IL-6. A significant correlation between mRNA and protein levels could be observed only for IL-6. Univariable analysis revealed that low IFN-É£ mRNA levels were associated with severe/fatal outcomes. The occurrence of COVID-19 pneumonia was significantly associated with lower expression levels of IL-6 mRNA, IFN-É£ mRNA, and TNF-ɑ mRNA. Concerning the severe/fatal outcomes, the multivariable logistic regression model revealed that none of the aforementioned parameters remained significant in the model. However, the logistic regression model revealed that higher TNF-ɑ mRNA expression was a significant independent predictor of absence of pneumonia [odds ratio: 0.35 (95% CI 0.14 to 0.88, p = 0.024)]. In conclusion, nasopharyngeal expression of AMPs (hBD-1, LL-37, and trappin-2) and cytokines (IL-6, IFN-É£, and TNF-ɑ) is upregulated in response to early SARS-CoV-2 infection, indicating that these AMPs and cytokines play a role in the local host defense against the virus. Upregulated nasopharyngeal TNF-ɑ mRNA expression during the early phase of SARS-CoV-2 infection was a significant independent predictor of the absence of COVID-19 pneumonia. Hence, high TNF-ɑ mRNA expression in the nasopharynx appears to be a protective factor for lung complications in COVID-19 patients.

Assuntos

Peptídeos Antimicrobianos , COVID-19 , Citocinas , Nasofaringe , SARS-CoV-2 , Humanos , COVID-19/imunologia , COVID-19/virologia , COVID-19/metabolismo , Nasofaringe/virologia , Nasofaringe/metabolismo , Feminino , Masculino , Pessoa de Meia-Idade , SARS-CoV-2/imunologia , Citocinas/metabolismo , Citocinas/genética , Adulto , Peptídeos Antimicrobianos/genética , Peptídeos Antimicrobianos/metabolismo , Idoso

RESUMO

INTRODUCTION: In the fight against virus-caused pandemics like COVID-19, diagnostic tests based on RT-qPCR are essential, but they are sometimes limited by their dependence on expensive, specialized equipment and skilled personnel. Consequently, an alternative nucleic acid detection technique that overcomes these restrictions, called loop-mediated isothermal amplification following reverse transcription (RT-LAMP), has been broadly investigated. Nevertheless, the developed RT-LAMP assays for SARS-CoV-2 detection still require laboratory devices and electrical power, limiting their widespread use as rapid home tests. This work developed a flexible RT-LAMP assay that gets beyond the drawbacks of the available isothermal LAMP-based SARS-CoV-2 detection, establishing a simple and effective at-home diagnostic tool for COVID-19. METHODOLOGY: A multiplex direct RT-LAMP assay, modified from the previously developed test was applied to simultaneously identify the two genes of SARS-CoV-2. We used a colorimetric readout, lyophilized reagents, and benchmarked an electro-free and micropipette-free method that enables sensitive and specific detection of SARS-CoV-2 in home settings. RESULTS: Forty-one nasopharyngeal swab samples were tested using the developed home-testing RT-LAMP (HT-LAMP) assay, showing 100% agreement with the RT-qPCR results. CONCLUSIONS: This is the first electrically independent RT-LAMP assay successfully developed for SARS-CoV-2 detection in a home setting. Our HT-LAMP assay is thus an important development for diagnosing COVID-19 or any other infectious pandemic on a population scale.

Assuntos

COVID-19 , Colorimetria , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico , SARS-CoV-2 , Sensibilidade e Especificidade , Humanos , COVID-19/diagnóstico , Técnicas de Amplificação de Ácido Nucleico/métodos , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Colorimetria/métodos , Técnicas de Diagnóstico Molecular/métodos , Teste de Ácido Nucleico para COVID-19/métodos , RNA Viral/análise , RNA Viral/genética , RNA Viral/isolamento & purificação , Nasofaringe/virologia

RESUMO

INTRODUCTION: COVID-19, an emerging infectious disease caused by SARS-CoV-2, continues to be a global public health threat. The development of a colorimetric reverse transcription loop-mediated isothermal amplification (RT-LAMP) can extend the availability of simple, reliable molecular tests for the rapid detection of COVID-19. METHODOLOGY: The RT-LAMP assay was developed using a new primer set targeting a portion of SARS-CoV-2 orf8. The method was validated at 63 ºC for 60 minutes with naked-eye visualization of the color change. The clinical performance was compared to a real-time reverse transcription-polymerase chain reaction (rtRT-PCR) using 273 RNA samples extracted from nasopharyngeal swab specimens. RESULTS: The developed RT-LAMP was specific to SARS-CoV-2 with a limit of detection at 15 RNA copies per reaction. The assay demonstrated diagnostic accuracy, sensitivity, specificity, positive predictive value, and negative predictive value of 90.48% (95% CI: 86.36-93.68%), 87.00% (95% CI: 81.53-91.33%), 100% (95% CI: 95.07-100%), 100% (95% CI: not available), and 73.74% (95% CI: 66.22-80.07%), respectively, compared to the rtRT-PCR. The greatest sensitivity of 98.03% (95% CI: 94.34-99.59%) was demonstrated in samples with the cycle threshold (Ct) values < 30 cycles. CONCLUSIONS: The RT-LAMP method in this study showed good performance. The assay can increase the scope of laboratory testing for rapidly detecting SARS-CoV-2 in Thailand. Due to a decrease in COVID-19 cases, its application is beneficial when commercial alternatives are unavailable.

Assuntos

COVID-19 , Colorimetria , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico , SARS-CoV-2 , Sensibilidade e Especificidade , Humanos , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico/métodos , Colorimetria/métodos , Tailândia , COVID-19/diagnóstico , Técnicas de Diagnóstico Molecular/métodos , RNA Viral/genética , RNA Viral/análise , RNA Viral/isolamento & purificação , Teste de Ácido Nucleico para COVID-19/métodos , Nasofaringe/virologia

RESUMO

Real-time reverse transcription polymerase chain reaction (RT-PCR), a standard method recommended for the diagnosis of coronavirus disease 2019 (COVID-19) requires 2-4 h to get the result. Although antigen test kit (ATK) is used for COVID-19 screening within 15-30 min, the drawback is its limited sensitivity. Hence, a rapid one-step quadruplex real-time RT-PCR assay: termed Æ©S COVID-19 targeting ORF1ab, ORF3a, and N genes of SARS-CoV-2; and Avocado sunblotch viroid (ASBVd) as an internal control was developed. Based on strategies including designing high melting temperature primers with short amplicons, applying a fast ramp rate, minimizing hold time, and reducing the range between denaturation and annealing/extension temperatures; the assay could be accomplished within 25 min. The limit of detection of ORF1ab, ORF3a, and N genes were 1.835, 1.310, and 1 copy/reaction, respectively. Validation was performed in 205 combined nasopharyngeal and oropharyngeal swabs. The sensitivity, specificity, positive predictive value, and negative predictive value were 92.8%, 100%, 100%, and 97.1%, respectively with 96.7% accuracy. Cohen's Kappa was 0.93. The newly developed rapid real-time RT-PCR assay was highly sensitive, specific, and fast, making it suitable for use as an alternative method to support laboratory diagnosis of COVID-19 in outpatient and emergency departments.

Assuntos

COVID-19 , SARS-CoV-2 , Sensibilidade e Especificidade , COVID-19/diagnóstico , COVID-19/virologia , Humanos , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/métodos , Teste de Ácido Nucleico para COVID-19/métodos , Feminino , Masculino , Pessoa de Meia-Idade , RNA Viral/genética , Adulto , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Nasofaringe/virologia , Proteínas Virais , Poliproteínas

RESUMO

BACKGROUND: Human parainfluenza virus-1 (HPIV-1) is a notable pathogen instigating acute respiratory tract infections in children. The article is to elucidate the epidemiological and genetic characteristics of HPIV-1 circulating in Hangzhou during the period of 2021-2022. METHODS: A cohort of 2360 nasopharyngeal swabs were amassed and subsequently examined via RT-PCR, with HPIV-1 positive samples undergoing P gene sequencing. RESULTS: The highest HPIV-1 infection rates were found in children aged between 3 and 6 years. A pronounced positive rate persisted through the latter half of 2021, with a notable decline observed in the initial half of 2022. All HPIV-1 strains could be clustered into 2 groups: Cluster 1, with strains similar to those found in Japan (LC764865, LC764864), and Cluster 2, with strains similar to the Beijing strain (MW575643). CONCLUSION: In conclusion, our study contributes to the comprehensive data on the epidemiological and genetic characteristics of HPIV-1 in pediatric patients from Hangzhou, post the COVID-19 peak.

Assuntos

Vírus da Parainfluenza 1 Humana , Filogenia , Humanos , China/epidemiologia , Pré-Escolar , Criança , Vírus da Parainfluenza 1 Humana/genética , Vírus da Parainfluenza 1 Humana/isolamento & purificação , Masculino , Feminino , Lactente , Adolescente , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/virologia , Nasofaringe/virologia , Infecções por Respirovirus/epidemiologia , Infecções por Respirovirus/virologia , Recém-Nascido

RESUMO

BACKGROUND: SARS-CoV-2 is responsible for the ongoing global pandemic, and the continuous emergence of novel variants threatens fragile populations, such as immunocompromised patients. This subgroup of patients seems to be seriously affected by intrahost viral changes, as the pathogens, which are keen to cause replication inefficiency, affect the impaired immune system, preventing efficient clearance of the virus. Therefore, these patients may represent an optimal reservoir for the development of new circulating SARS-CoV-2 variants. The following study aimed to investigate genomic changes in SARS-CoV-2-positive immunocompromised patients over time. METHODS: SARS-CoV-2-positive nasopharyngeal swabs were collected at different time points for each patient (patient A and patient B), extracted and then analyzed through next-generation sequencing (NGS). The resulting sequences were examined to determine mutation frequencies, describing viral evolution over time. CASE PRESENTATION: Patient A was a 53-year-old patient with onco-hematological disease with prolonged infection lasting for 51 days from May 28th to July 18th, 2022. Three confirmed SARS-CoV-2-positive samples were collected on May 28th, June 15th and July 4th. Patient B was 75 years old and had onco-hematological disease with prolonged infection lasting for 146 days. Two confirmed positive SARS-CoV-2 samples were collected at the following time points: May 21st and August 18th. CONCLUSION: Heat map construction provided evidence of gain and/or loss of mutations over time for both patients, suggesting within-host genomic evolution of the virus. In addition, mutation polymorphisms and changes in the SARS-CoV-2 lineage were observed in Patient B. Sequence analysis revealed high mutational pattern variability, reflecting the high complexity of viral replication dynamics in fragile patients.

Assuntos

COVID-19 , Sequenciamento de Nucleotídeos em Larga Escala , Hospedeiro Imunocomprometido , SARS-CoV-2 , Humanos , COVID-19/virologia , SARS-CoV-2/genética , Pessoa de Meia-Idade , Genoma Viral/genética , Masculino , Mutação , Evolução Molecular , Nasofaringe/virologia

RESUMO

The molecular reverse transcription-polymerase chain reaction (RT-PCR) testing of respiratory tract swabs has become mandatory to confirm the diagnosis of coronavirus disease 2019 (COVID-19). However, RT-PCR tests are expensive, require standardized equipment, and relatively long testing times, and the sample pooling method has been introduced to solve this issue. The aim of this study was to compare the cycle threshold (Ct) values of the individual sample and pooled sample methods to assess how accurate the pooling method was. Repeat RT-PCR examinations were initially performed to confirm the Ct values for each sample before running the pooled test procedure. Sample extraction and amplification were performed in both assays to detect ORF1ab, N, and E genes with a cut-off point value of Ct <38. Overall, there was no difference in Ct values between individual sample and pooled sample groups at all concentrations (p=0.259) and for all pooled sizes. Only pooled size of five could detect the Ct value in the pooled samples for all concentration samples, including low-concentration sample (Ct values 36 to 38). This study highlighted that pooled RT-PCR testing strategy did not reduce the quality of individually measured RT-PCR Ct values. A pool size of five could provide a practical technique to expand the screening capacity of RT-PCR.

Assuntos

COVID-19 , Nasofaringe , Reação em Cadeia da Polimerase Via Transcriptase Reversa , SARS-CoV-2 , Manejo de Espécimes , Humanos , Nasofaringe/virologia , SARS-CoV-2/genética , COVID-19/diagnóstico , COVID-19/virologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Manejo de Espécimes/métodos , Teste de Ácido Nucleico para COVID-19/métodos

RESUMO

Epidemiological studies report the impact of co-infection with pneumococcus and respiratory viruses upon disease rates and outcomes, but their effect on pneumococcal carriage acquisition and bacterial load is scarcely described. Here, we assess this by combining natural viral infection with controlled human pneumococcal infection in 581 healthy adults screened for upper respiratory tract viral infection before intranasal pneumococcal challenge. Across all adults, respiratory syncytial virus (RSV) and rhinovirus asymptomatic infection confer a substantial increase in secondary infection with pneumococcus. RSV also has a major impact on pneumococcal density up to 9 days post challenge. We also study rates and kinetics of bacterial shedding through the nose and oral route in a subset. High levels of pneumococcal colonization density and nasal inflammation are strongly correlated with increased odds of nasal shedding as opposed to cough shedding. Protection against respiratory viral infections and control of pneumococcal density may contribute to preventing pneumococcal disease and reducing bacterial spread.

Assuntos

Derrame de Bactérias , Portador Sadio , Coinfecção , Infecções por Picornaviridae , Infecções Pneumocócicas , Infecções por Vírus Respiratório Sincicial , Rhinovirus , Streptococcus pneumoniae , Humanos , Rhinovirus/fisiologia , Adulto , Infecções Pneumocócicas/microbiologia , Infecções por Picornaviridae/virologia , Infecções por Picornaviridae/microbiologia , Portador Sadio/microbiologia , Masculino , Feminino , Infecções por Vírus Respiratório Sincicial/virologia , Coinfecção/microbiologia , Coinfecção/virologia , Adulto Jovem , Carga Bacteriana , Pessoa de Meia-Idade , Inflamação , Vírus Sinciciais Respiratórios/fisiologia , Infecções Respiratórias/microbiologia , Infecções Respiratórias/virologia , Adolescente , Nasofaringe/microbiologia , Nasofaringe/virologia

RESUMO

Objective: To investigate the epidemiological characteristics of human respiratory syncytial virus (HRSV) in patients with Severe Acute Respiratory Infection (SARI) in Tianjin from 2015 to 2020. Methods: The study data were obtained from the Third Center Hospital of Tianjin, a designated sentinel hospital, from 2015 to 2020, with 1 597 SARI patients enrolled in this study. The clinical specimens of the research participants were subjected to respiratory multi-pathogen testing. HRSV-positive specimens were subtyped to analyze the differences in HRSV detection rates among cases of different age groups and periods and their mixed infection situations. Results: A total of 1 597 nasopharyngeal swabs were collected, with an HRSV detection rate of 4.20%. Among 67 HRSV-positive specimens, there were 19 pure HRSV-A nucleic acid-positive specimens, 19 pure HRSV-B nucleic acid-positive specimens and 29 mixed HRSV-A and HRSV-B nucleic acid-positive specimens. The difference in HRSV detection rate among different age groups was statistically significant (P<0.05), and the HRSV detection rate in children under five years old was higher than that in other age groups. From 2016 to 2020, the detection rate of HRSV showed an increasing trend year by year. The HRSV detection rate of SARI cases was highest in the winter season, at 7.15%. There were 10 (14.93%) mixed positive cases for HRSV and other viruses, of which four were mixed positive for HRSV and influenza A. Conclusion: The incidence of HRSV in Tianjin exhibits an increasing trend from 2016 to 2020, peaking during the winter season, with children under five years old constituting a high-risk demographic for HRSV infection.

Assuntos

Infecções por Vírus Respiratório Sincicial , Vírus Sincicial Respiratório Humano , Infecções Respiratórias , Humanos , Vírus Sincicial Respiratório Humano/isolamento & purificação , Infecções por Vírus Respiratório Sincicial/epidemiologia , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/virologia , China/epidemiologia , Lactente , Criança , Pré-Escolar , Nasofaringe/virologia , Estações do Ano , Doença Aguda , Feminino , Masculino , Adulto , Adolescente , Pessoa de Meia-Idade

RESUMO

BACKGROUND: Acute respiratory infections are a leading cause of morbidity and mortality in children. However, studies on the prevalence of respiratory viruses among children with acute respiratory infections in Kunming, China, are lacking. Therefore, we aimed to investigate the epidemiological characteristics of respiratory pathogens among children with acute respiratory infections in Kunming during the coronavirus disease 2019 pandemic. METHODS: Nasopharyngeal swab samples were collected from 4956 children with acute respiratory infections at Yunnan Provincial First People's Hospital between January 2020 and December 2022, patients with COVID-19 were excluded from the study. Multiplex reverse transcription polymerase chain reaction was used to detect respiratory pathogens. RESULTS: The frequency of respiratory pathogens among children was significantly lower in 2020 than in 2021 and 2022. The following pathogens had the highest prevalence rates (in descending order) from 2020 to 2022: HRV > RSV > PIV > ADV > MP; HRV > RSV > HADV > PIV > MP and HRV > Mp > HADV > H3N2 > HMPV. The overall frequency of respiratory pathogens exhibited an inverted U-shape with increasing age among the children. Human bocavirus, human parainfluenza virus, and human respiratory syncytial virus were the dominant respiratory viruses in children aged ≤ 3 years, whereas Mycoplasma pneumoniae was the dominant respiratory pathogen in children aged > 3 years. HRV has the highest prevalence and is the main pathogen of mixed infection. The prevalence of the influenza A virus has decreased significantly, whereas HRSV and Mp are found to be seasonal. CONCLUSIONS: Our findings offer an objective evaluation of transmission dynamics and epidemiological shifts in respiratory pathogens during the coronavirus disease 2019 pandemic in Kunming, serving as a basis for informed decision-making, prevention, and treatment strategies.

Assuntos

COVID-19 , Infecções Respiratórias , SARS-CoV-2 , Humanos , COVID-19/epidemiologia , COVID-19/transmissão , China/epidemiologia , Pré-Escolar , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/virologia , Lactente , Criança , Feminino , Masculino , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Prevalência , Adolescente , Nasofaringe/virologia , Recém-Nascido

RESUMO

BACKGROUND: The COVID-19 pandemic has underscored the critical role of sequencing technology in disease control and outbreak response. However, resource limitations and challenging environments often impede such efforts in low and middle-income countries. This study aimed to investigate the spectrum of viral co-infections, particularly with human viral pathogens, in SARS-CoV-2 positive individuals in Sierra Leone using metagenomic sequencing, evaluating the feasibility of utilizing this technology for epidemiological and evolutionary surveillance of pathogens related to public health in low-income environments. METHODS: We retrospectively collected and analyzed 98 nasopharyngeal swab specimens from SARS-CoV-2 positive individuals in Sierra Leone. Samples were pre-processed locally and transferred to China via FTA cards for metagenomic sequencing, which was performed using the Novaseq platform. The study focused on the identification of nasopharyngeal viruses co-infecting with SARS-CoV-2, with a deeper analysis of significant human viral pathogens such as HPV. RESULTS: The study identified 22 viral taxa from 20 families, including 4 human viruses. Notably, 19.4% of samples showed HPV co-infection with 34 distinct types, predominantly beta and gamma HPVs. Multiple HPV types were found in individual samples, indicating a high complexity of viral co-infections. CONCLUSIONS: The identification of a wide range of co-infecting viruses, particularly multiple HPV genotypes, highlights the complexity of viral interactions and their potential implications for public health. These findings enhance our understanding of viral co-infections and provide valuable insights for public health interventions in Sierra Leone. Further research is needed to explore the clinical significance of these findings and their impact on disease outcomes.

Assuntos

COVID-19 , Coinfecção , Infecções por Papillomavirus , SARS-CoV-2 , Humanos , Serra Leoa/epidemiologia , Estudos Retrospectivos , COVID-19/virologia , COVID-19/epidemiologia , Infecções por Papillomavirus/virologia , Infecções por Papillomavirus/epidemiologia , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/classificação , Coinfecção/virologia , Coinfecção/epidemiologia , Feminino , Adulto , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Nasofaringe/virologia , Adolescente , Metagenômica , Filogenia , Papillomaviridae/genética , Papillomaviridae/classificação , Papillomaviridae/isolamento & purificação , Idoso , Criança

RESUMO

BACKGROUND AND OBJECTIVE: Viral respiratory infections stand as a considerable global health concern, presenting significant risks to the health of both humans and animals. This study aims to conduct a preliminary analysis of the time series of viral load in the nasal cavity-nasopharynx (NC-NP) of the human and rhesus macaque (RM). METHODS: Taking into account the random uniform distribution of virus-laden droplets with a diameter of 10 µm in the mucus layer, this study applies the computational fluid dynamics-host cell dynamics (CFD-HCD) method to 3D-shell NC-NP models of human and RM, analyzing the impact of initial distribution of droplets on the viral dynamics of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), estimating parameters in the HCD model based on experimental data, integrating them into simulations to predict the time series of viral load and cell counts, and being visualized. The reproductive number (R0) are calculated to determine the occurrence of infection. The study also considers cross-parameter combinations and cross-experimental datasets to explore potential correlations between the human and RM. RESULTS: The research findings indicate that the uniform distribution of virus-laden droplets throughout the whole NC-NP models of human and RM is reasonable for simulating and predicting viral dynamics. The visualization results offer dynamic insights into virus infection over a period of 20 days. Studies involving parameter and dataset exchanges between the two species underscore certain similarities in predicting virus infections between the human and RM. CONCLUSIONS: This study lays the groundwork for further exploration into the parallels and distinctions in respiratory virus dynamics between humans and RMs, thus aiding in making more informed decisions in research and experimentation.

Assuntos

COVID-19 , Macaca mulatta , Cavidade Nasal , Nasofaringe , SARS-CoV-2 , Carga Viral , Humanos , Animais , Cavidade Nasal/virologia , Nasofaringe/virologia , COVID-19/virologia , Hidrodinâmica , Simulação por Computador , Muco/virologia , Modelos Biológicos

RESUMO

The novel coronavirus SARS-CoV-2 was first isolated in late 2019; it has spread to all continents, infected over 700 million people, and caused over 7 million deaths worldwide to date. The high transmissibility of the virus and the emergence of novel strains with altered pathogenicity and potential resistance to therapeutics and vaccines are major challenges in the study and treatment of the virus. Ongoing screening efforts aim to identify new cases to monitor the spread of the virus and help determine the danger connected to the emergence of new variants. Given its sensitivity and specificity, nucleic acid amplification tests (NAATs) such as RT-qPCR are the gold standard for SARS-CoV-2 detection. However, due to high costs, complexity, and unavailability in low-resource and point-of-care (POC) settings, the available RT-qPCR assays cannot match global testing demands. An alternative NAAT, RT-LAMP-based SARS-CoV-2 detection offers scalable, low-cost, and rapid testing capabilities. We have developed an automated RT-LAMP-based microfluidic chip that combines the RNA isolation, purification, and amplification steps on the same device and enables the visual detection of SARS-CoV-2 within 40 min from saliva and nasopharyngeal samples. The entire assay is executed inside a uniquely designed, inexpensive disposable microfluidic chip, where assay components and reagents have been optimized to provide precise and qualitative results and can be effectively deployed in POC settings. Furthermore, this technology could be easily adapted for other novel emerging viruses.

Assuntos

COVID-19 , Técnicas de Diagnóstico Molecular , Nasofaringe , Técnicas de Amplificação de Ácido Nucleico , RNA Viral , SARS-CoV-2 , Saliva , Sensibilidade e Especificidade , Humanos , Saliva/virologia , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/genética , Nasofaringe/virologia , COVID-19/diagnóstico , COVID-19/virologia , Técnicas de Amplificação de Ácido Nucleico/métodos , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Diagnóstico Molecular/instrumentação , RNA Viral/genética , RNA Viral/análise , RNA Viral/isolamento & purificação , Dispositivos Lab-On-A-Chip , Teste de Ácido Nucleico para COVID-19/métodos , Teste de Ácido Nucleico para COVID-19/instrumentação , Teste para COVID-19/métodos , Microfluídica/métodos , Microfluídica/instrumentação

RESUMO

This study comprehensively evaluated the DNA/RNA Defend Pro (DRDP) sample collection buffer, designed to inactivate and stabilize patient samples. The primary objectives were to assess DRDP's efficacy in ensuring sample stability, facilitating extraction-free polymerase chain reaction (PCR), and ensuring compatibility with rapid antigen testing (RAT). Ninety-five diagnostic nasopharyngeal swab samples tested for influenza virus (influenza A), respiratory syncytial virus (RSV A), and/or severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) were 10-fold diluted with DRDP and anonymized. Initial characterization and retesting of these samples using cobas Liat confirmed 88 samples as positive, validating the presence of viral targets. Results from rapid antigen testing showed lower sensitivity compared to nucleic acid amplification testing (NAAT) but maintained perfect specificity, with 40 out of 88 positive samples by cobas Liat also testing positive for RAT. Direct RT-qPCR of DRDP-diluted samples demonstrated robust compatibility, with 72 out of 88 samples positive for cobas Liat also testing positive by direct RT-qPCR. Non-concordant results could be explained by the 200-fold lower input of extraction-free NAAT. Stability testing involved incubating 31 positive samples at 4 °C, 20 °C, and 37 °C for 7 days, with extraction-free NAAT. DRDP guaranteed viral RNA stability at all temperatures for influenza A, SARS-CoV-2, and RSV A, showing stability up to 7 days at 4 °C. In conclusion, DRDP is an effective stabilizing medium compatible with direct RT-qPCR and rapid antigen testing and shows great potential for optimizing diagnostic processes, particularly in resource-limited or time-sensitive scenarios.

Assuntos

Nasofaringe , SARS-CoV-2 , Manejo de Espécimes , Nasofaringe/virologia , Humanos , Manejo de Espécimes/métodos , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/imunologia , SARS-CoV-2/genética , RNA Viral/análise , RNA Viral/isolamento & purificação , RNA Viral/genética , Soluções Tampão , Vírus da Influenza A/isolamento & purificação , Vírus da Influenza A/imunologia , Vírus da Influenza A/genética , Sensibilidade e Especificidade , COVID-19/diagnóstico , COVID-19/virologia , Antígenos Virais/análise , Influenza Humana/diagnóstico , Influenza Humana/virologia

RESUMO

While severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is characterized by impaired induction of interferons (IFNs) and IFN-stimulated genes (ISGs), the IFNs and ISGs in upper airway is essential to restrict the spread of respiratory virus. Here, we identified the prominent IFN and ISG upregulation in the nasopharynx (NP) of mild and even severe coronavirus disease 2019 (COVID-19) patients (CoV2+) in Omicron era and to compare their clinical outcome depending on the level of IFNs and ISGs. Whereas the induction of IFNB was minimal, transcription of IFNA, IFNG, and IFNLs was significantly increased in the NP of CoV2 + patients. IFNs and ISGs may be more upregulated in the NP of CoV2 + patients at early phases of infection according to viral RNA levels and this is observed even in severe cases. IFN-related innate immune response might be characteristic in macrophages and monocytes at the NP and the CoV2 + patients with higher transcription of IFNs and ISGs in the NP showed a correlation with good prognosis of COVID-19. This study presents that IFNs and ISGs may be upregulated in the NP, even in severe CoV2 + patients depending on viral replication during Omicron-dominant period and the unique IFN-responsiveness in the NP links with COVID-19 clinical outcomes.

Assuntos

COVID-19 , Imunidade Inata , Interferons , Nasofaringe , SARS-CoV-2 , Humanos , COVID-19/imunologia , COVID-19/virologia , Nasofaringe/virologia , Nasofaringe/imunologia , SARS-CoV-2/imunologia , SARS-CoV-2/fisiologia , Interferons/metabolismo , Interferons/genética , Interferons/imunologia , Masculino , Feminino , Pessoa de Meia-Idade , Adulto , Idoso