Clinical features of patients infected with 2019 novel coronavirus in Wuhan, China

Background Since December 2019, when coronavirus disease 2019 (Covid-19) emerged in Wuhan city and rapidly spread throughout China, data have been needed on the clinical characteristics of...

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Clinical Characteristics of Coronavirus Disease 2019 in China.

In December 2019, a cluster of patients with pneumonia of unknown cause was linked to a seafood wholesale market in Wuhan, China. A previously unknown betacoronavirus was discovered through the use of unbiased sequencing in samples from patients with pneumonia. Human airway epithelial cells were used to isolate a novel coronavirus, named 2019-nCoV, which formed a clade within the subgenus sarbecovirus, Orthocoronavirinae subfamily. Different from both MERS-CoV and SARS-CoV, 2019-nCoV is the seventh member of the family of coronaviruses that infect humans. Enhanced surveillance and further investigation are ongoing. (Funded by the National Key Research and Development Program of China and the National Major Project for Control and Prevention of Infectious Disease in China.).

https://www.nejm.org/doi/pdf/10.1056/NEJMoa2001017?articleTools=true

A Novel Coronavirus from Patients with Pneumonia in China, 2019.

Importance In December 2019, novel coronavirus (2019-nCoV)–infected pneumonia (NCIP) occurred in Wuhan, China. The number of cases has increased rapidly but information on the clinical characteristics of affected patients is limited. Objective To describe the epidemiological and clinical characteristics of NCIP. Design, Setting, and Participants Retrospective, single-center case series of the 138 consecutive hospitalized patients with confirmed NCIP at Zhongnan Hospital of Wuhan University in Wuhan, China, from January 1 to January 28, 2020; final date of follow-up was February 3, 2020. Exposures Documented NCIP. Main Outcomes and Measures Epidemiological, demographic, clinical, laboratory, radiological, and treatment data were collected and analyzed. Outcomes of critically ill patients and noncritically ill patients were compared. Presumed hospital-related transmission was suspected if a cluster of health professionals or hospitalized patients in the same wards became infected and a possible source of infection could be tracked. Results Of 138 hospitalized patients with NCIP, the median age was 56 years (interquartile range, 42-68; range, 22-92 years) and 75 (54.3%) were men. Hospital-associated transmission was suspected as the presumed mechanism of infection for affected health professionals (40 [29%]) and hospitalized patients (17 [12.3%]). Common symptoms included fever (136 [98.6%]), fatigue (96 [69.6%]), and dry cough (82 [59.4%]). Lymphopenia (lymphocyte count, 0.8 × 109/L [interquartile range {IQR}, 0.6-1.1]) occurred in 97 patients (70.3%), prolonged prothrombin time (13.0 seconds [IQR, 12.3-13.7]) in 80 patients (58%), and elevated lactate dehydrogenase (261 U/L [IQR, 182-403]) in 55 patients (39.9%). Chest computed tomographic scans showed bilateral patchy shadows or ground glass opacity in the lungs of all patients. Most patients received antiviral therapy (oseltamivir, 124 [89.9%]), and many received antibacterial therapy (moxifloxacin, 89 [64.4%]; ceftriaxone, 34 [24.6%]; azithromycin, 25 [18.1%]) and glucocorticoid therapy (62 [44.9%]). Thirty-six patients (26.1%) were transferred to the intensive care unit (ICU) because of complications, including acute respiratory distress syndrome (22 [61.1%]), arrhythmia (16 [44.4%]), and shock (11 [30.6%]). The median time from first symptom to dyspnea was 5.0 days, to hospital admission was 7.0 days, and to ARDS was 8.0 days. Patients treated in the ICU (n = 36), compared with patients not treated in the ICU (n = 102), were older (median age, 66 years vs 51 years), were more likely to have underlying comorbidities (26 [72.2%] vs 38 [37.3%]), and were more likely to have dyspnea (23 [63.9%] vs 20 [19.6%]), and anorexia (24 [66.7%] vs 31 [30.4%]). Of the 36 cases in the ICU, 4 (11.1%) received high-flow oxygen therapy, 15 (41.7%) received noninvasive ventilation, and 17 (47.2%) received invasive ventilation (4 were switched to extracorporeal membrane oxygenation). As of February 3, 47 patients (34.1%) were discharged and 6 died (overall mortality, 4.3%), but the remaining patients are still hospitalized. Among those discharged alive (n = 47), the median hospital stay was 10 days (IQR, 7.0-14.0). Conclusions and Relevance In this single-center case series of 138 hospitalized patients with confirmed NCIP in Wuhan, China, presumed hospital-related transmission of 2019-nCoV was suspected in 41% of patients, 26% of patients received ICU care, and mortality was 4.3%.

https://jamanetwork.com/journals/jama/articlepdf/2761044/jama_wang_2020_oi_200019.pdf

Clinical Characteristics of 138 Hospitalized Patients With 2019 Novel Coronavirus-Infected Pneumonia in Wuhan, China.

SARS-CoV-2 Cell Entry Depends on ACE2 and TMPRSS2 and Is Blocked by a Clinically Proven Protease Inhibitor

Human immunodeficiency virus 1 (HIV-1) attacks the immune system, making people susceptible to various diseases, thus increasing their risk of death. Comprehensive detection of major HIV-1 strains circulating in China is vital for effective HIV-1 infection prevention and treatment. HIV-1 nucleic acid detection is considered effective for HIV-1 diagnosis since traditional immunological testing may fail to detect HIV-1 infection during the window period. This work demonstrates a one-pot two-stage amplification assay (RT-RAP), a combination of reverse transcription recombinase (RT- RAA), and quantitative real-time polymerase chain reaction (qRT-PCR). The turn-around time of the assay is only 50 min and can be performed with commonly available laboratory equipment, the qPCR devices. The RT-RAP assay could detect approximately 5 and 14 copies/reaction of HIV-1 DNA and RNA using recombinant plasmids and standard reference strains, respectively. Additionally, we found that the clinical performance of RT-RAP (detected 169 samples out of 170 specimens) was consistent with that of qRT-PCR. The sensitivity and specificity of RT-RAP were 100.00% (99/99) and 98.59% (70/71), respectively, while its positive and negative predictive values were 99.00% (99/100) and 100.00% (70/70), respectively. The total coincidence rate of the RT-RAP was 99.41% (169/170), with a kappa value of 0.988 (p < 0.05). We demonstrated that RT-RAP could rapidly detect the common HIV-1 subtypes commonly circulating in China with comparable sensitivity and specificity to qRT-PCR.

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A Rapid and Sensitive Detection of HIV-1 with a One-Pot Two-Stage Reverse Transcription Recombinase Aided Real-Time PCR Assay

In 2008, China launched a national surveillance system for hand-foot-and-mouth disease (HFMD). Several million cases of HFMD are reported every year, CVA16 was the leading cause of HFMD epidemic in Yantai city, China in recent years, but the information of epidemiology and molecular characterization of CVA16 in Yantai is limited. The aim of this study is to investigate the epidemiological characteristics and pathogenic spectrum of HFMD, and most importantly, the molecular characterization of CVA16 in Yantai from 2018 to 2021. A total of 2,000 clinical samples were collected in Yantai city from 2018 to 2021 and the enterovirus typing was performed using real-time reverse transcriptase–polymerase chain reaction (qRT-PCR). VP1 coding regions of 41 CVA16 isolates were amplified and Sanger sequenced, and phylogenetic analysis was performed. During the study period, HFMD became prevalent from May to August each year. It peaked in June and declined in September. The incidence was highest in children aged 1 to 5 years, while more common in males than females. 1,617 out of 2,000 clinical collection of samples were tested positive for enterovirus. Among them, 614 were identified as CVA16, 45 were EV-A71, and 958 were other enterovirus serotypes. All 41 CVA16 strains belonged to the Bla and B1b genotypes. Homology analysis showed that 41 CVA16 isolates shared 83.2%–100% nucleotide and 93.7%–100% amino acid similarity among themselves. The results of this study update molecular epidemiology of CVA16 and provide a reference for HFMD prevention and control. 

Epidemiology and genetic characteristics of coxsackievirus A16 associated with hand-foot-and-mouth disease in Yantai city, China in 2018-2021

Introduction Diarrhea is a significant health problem in third world countries; identification of causative agents of diarrhea is essential to apply measures to prevent and control this disease. In addition, scant data are available regarding childhood diarrhea in Sudan. Our research aimed to determine the incidence of specific protozoan pathogens (Entameobia histolytica, Cryptosporidium spp., and Gardia lambelia) among the young (aged less than five years) in Khartoum, Sudan. Methods We conducted a parasitological cross-sectional survey, and stool samples from 437 patients were examined for E. histolytica, C.  parvum, and G. lambelia using a multiplex real-time PCR method. Results Of the 437 stool samples tested, infection with intestinal parasite was found in 155 (35.5%) cases, and co-infection was identified in 16 (3.7%) cases. G. lambelia (18.8%) and C. parvum (15.8 %) were the most frequently identified parasites, followed by E. histolytica (0.5%). The highest and lowest rates of parasitic infections were seen in the less than two years age group (32.7%), and in the 2–4-year-old group (2.7%), the male children showed higher rates of infections (23.7%) compared to females (11.7%). The incidence of protozoan infection was higher (37.7%) in the rainy season (August to December) (32.7%) in contrast with that (2.7%) in the dry season (April to June). Discussion Our present study demonstrated the high prevalence of G. lambelia and C. parvum in children with diarrhea in Khartoum State and the multiplex real-time technique's usefulness in disclosing pathogenic protozoal agents. Our result highlighted the necessity of developing intervention measurement and control strategies to deal with childhood parasitic diarrhea in this region.

Molecular survey of certain protozoan agents that cause diarrhea in children in Sudan

Nucleic acid amplification enhancers suitable for use in a recombinase-aided amplification (RAA) assay were studied for the first time, and amplification of a long-fragment (509 bp) was initially explored. Using recombinant plasmids and clinical samples, RAA fluorescence and basic methods were used to evaluate the efficacy. The fluorescence method was evaluated by threshold time and fluorescence value, and the basic method was characterized by 2% agarose gel electrophoresis. Taking a previously established RAA assay for HPV18 as an example, we demonstrated that the addition of 0.2 M, 0.4 M, and 0.6 M betaine and 10% pullulan could enhance the RAA. The new RAA assays with betaine and pullulan were named B-RAA and P-RAA, respectively. Using the B-RAA and P-RAA fluorescence methods, the threshold time values could be shortened by 1.72–2.32 minutes and 2.60 minutes, respectively, and the fluorescence values could be enhanced by 8847.25–9094.37 mv and 5250 mv, respectively. Using the basic method, the sensitivity could be increased 10-fold. We successfully amplified a long-fragment of 509 bp using a P-RAA assay with a sensitivity of 102 copies/µL (compared with 103 copies/µL in the RAA assay). Thus, we concluded that betaine and pullulan are effective additives to enhance the sensitivity of RAA assays. 

Enhancement of a recombinase-aided amplification assay using betaine and pullulan

Epstein-Barr virus (EBV), a common human γ-herpesvirus, infects more than 90% of adults worldwide. The purpose of this study was to establish a novel EBV detection method by combining the recombinase aided amplification (RAA) assay with an initial enrichment step that utilizes magnetic beads coated with a recombinant human mannan-binding lectin (rhMBL, M1 protein). An M1 protein-protein A magnetic bead complex (M1 beads) was prepared and used to achieve separation and enrichment of EBV from blood. After nucleic acid extraction, DNA was amplified by RAA. Using 388 whole blood samples and 1 serum sample, we explored the specificity, sensitivity and applicability of the newly developed detection method and compared it with commercial quantitative real-time polymerase chain reaction (qPCR) following M1 bead enrichment, traditional qPCR and traditional RAA. After enrichment, the positivity rate of EBV was increased from 15.94% to 17.74% by RAA (P < 0.05) and from 7.20% to 15.17% by qPCR (P < 0.05). The viral loads after enrichment were increased by 1.13 to 23.19-fold (P < 0.05). Our data demonstrates that an RAA assay incorporating M1 bead enrichment is a promising tool for detecting low EBV viral loads in blood samples that will facilitate an early response to EBV infection. 

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Xuejun Ma

Papers

A Rapid and Sensitive Detection of HIV-1 with a One-Pot Two-Stage Reverse Transcription Recombinase Aided Real-Time PCR Assay

Epidemiology and genetic characteristics of coxsackievirus A16 associated with hand-foot-and-mouth disease in Yantai city, China in 2018-2021

Molecular survey of certain protozoan agents that cause diarrhea in children in Sudan

Enhancement of a recombinase-aided amplification assay using betaine and pullulan

Detection of low-load Epstein-Barr virus in blood samples by enriched recombinase aided amplification assay