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Advances in genotypic antimicrobialresistance testing: a comprehensive review

Science China Life Sciences 2025年第01期 130-143页

作者： Boheng Duan Xianjun Zeng Junping Peng Huan Kui College of Nanchang University Department of Imaging The Second Affiliated Hospital of Nanchang University nhc key laboratory of systems biology of pathogens National Institute of Pathogen BiologyChinese Academy of Medical Sciences & Peking Union Medical College key laboratory of Respiratory Disease Pathogenomics Chinese Academy of Medical Sciences & Peking Union Medical College key laboratory of Pathogen Infection Prevention and Control (Ministry of Education) State Key Laboratory of Respiratory Health and MultimorbidityNational Institute of Pathogen BiologyChinese Academy of Medical Sciences & Peking Union Medical College

Antimicrobial resistance(AMR) represents a substantial threat to global public health, complicating the treatment of common infections and leading to prolonged illness and escalated healthcare expenses. To effectively combat AMR, timely and accurate detection is crucial for AMR surveillance and individual-based therapy. Phenotypic antibiotic resistance testing(AST) has long been considered the gold standard in clinical applications, serving as the foundation for clinical AMR diagnosis and optimized therapy. It has significantly contributed to ensuring patients′ health and the development of novel antimicrobials. Despite advancements in automated culture-based AST technologies,inherent limitations impede the widespread use of phenotypic AST in AMR surveillance. Genotypic AST technologies offer a promising alternative option, exhibiting advantages of rapidity, high sensitivity, and specificity. With the continuous advancement and expanding applications of genotypic AST technologies, such as microfluidics, mass spectrometry, and high-resolution melting curve analysis, new vigor has been injected into the development and clinical implementation of genotypic AST technologies. In this narrative review, we discuss the principles, applications, and advancements of emerging genotypic AST methods in clinical settings. The comprehensive review aims to highlight the significant scientific potential of emerging genotypic AST technologies in clinical AMR diagnosis, providing insights to enhance existing methods and explore novel approaches.

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Construction of Non-infectious SARS-CoV-2 Replicons and Their Application in Drug Evaluation

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Virologica Sinica 2021年第5期36卷 890-900页

作者： Bei Wang Chongyang Zhang Xiaobo Lei Lili Ren He Huang Jianwei Wang Zhendong Zhao nhc key laboratory of systems biology of pathogens Institute of Pathogen BiologyChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijing 100176China nhc key laboratory of systems biology of pathogens and christophe me´rieux laboratory Institute of Pathogen BiologyChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijing 100176China

Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)has caused a devastating pandemic *** and antiviral drugs are the most promising candidates for combating this global epidemic,and scientists all over the world have made great efforts to this ***,manipulation of the SARS-CoV-2 should be performed in the biosafety level3 *** makes experiments complicated and ***,a safer system for working with this virus is urgently ***,we report the construction of plasmid-based,non-infectious SARS-CoV-2 replicons with turbo-green fluorescent protein and/or firefly luciferase reporters by reverse genetics using transformation-associated recombination cloning in Saccharomyces *** of these replicons was achieved simply by direct transfection of cells with the replicon plasmids as evident by the expression of reporter *** SARS-CoV-2 replicons,the inhibitory effects of E64-D and remdesivir on SARS-CoV-2 replication were confirmed,and the halfmaximal effective concentration(EC50)value of remdesivir and E64-D was estimated by different quantification methods respectively,indicating that these SARS-CoV-2 replicons are useful tools for antiviral drug evaluation.

关键词： SARS-CoV-2 Reverse genetics Replicon Antiviral drugs Drug evaluation

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SARS-CoV-2-derived fusion inhibitor lipopeptides exhibit highly potent and broad-spectrum activity against divergent human coronaviruses

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Signal Transduction and Targeted Therapy 2021年第9期6卷 2615-2617页

作者： Yuanmei Zhu Danwei Yu Yue Hu Tong Wu Huihui Chong Yuxian He nhc key laboratory of systems biology of pathogens Institute of Pathogen BiologyChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijingChina

Dear Editor,Most members of coronaviruses(CoVs),including four annually circulating human CoVs(HCov-229E,HCoV-OC43,HCoV-NL63;and CoV-HKU1),infect the respiratory tract of mammals and cause mild illness;however,zoonotic CoVs can cross the species barrier from animal reservoirs and lead to epidemics with high morbidity and mortality in humans.

关键词： mortality respiratory annually

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Application of Nanopore Sequencing Technology in the Clinical Diagnosis of Infectious Diseases

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Biomedical and Environmental Sciences 2022年第5期35卷 381-392页

作者： ZHANG Lu Lu ZHANG Chi PENG Jun Ping nhc key laboratory of systems biology of pathogens Institute of Pathogen BiologyChinese Academy of Medical Sciences&Peking Union Medical CollegeBeijing 100176China key laboratory of Respiratory Disease Pathogenomics Chinese Academy of Medical SciencesBeijing 100176China

Infectious diseases are an enormous public health burden and a growing threat to human health *** or classic recurrent pathogens,or pathogens with resistant traits,challenge our ability to diagnose and control infectious *** sequencing technology has the potential to enhance our ability to diagnose,interrogate,and track infectious diseases due to the unrestricted read length and system *** review focuses on the application of nanopore sequencing technology in the clinical diagnosis of infectious diseases and includes the following:(i)a brief introduction to nanopore sequencing technology and Oxford Nanopore Technologies(ONT)sequencing platforms;(ii)strategies for nanopore-based sequencing technologies;and(iii)applications of nanopore sequencing technology in monitoring emerging pathogenic microorganisms,molecular detection of clinically relevant drug-resistance genes,and characterization of disease-related microbial ***,we discuss the current challenges,potential opportunities,and future outlook for applying nanopore sequencing technology in the diagnosis of infectious diseases.

关键词： Nanopore sequencing Infectious diseases Pathogen Oxford Nanopore Technologies

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A comprehensive survey of bat sarbecoviruses across China in relation to the origins of SARS-CoV and SARS-CoV-2

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National Science Review 2023年第6期10卷 117-130页

作者： Zhiqiang Wu Yelin Han Yuyang Wang Bo Liu Lamei Zhao Junpeng Zhang Haoxiang Su Wenliang Zhao Liguo Liu Shibin Bai Jie Dong Lilian Sun Yafang Zhu Siyu Zhou Yiping Song Hongtao Sui Jian Yang Jianwei Wang Shuyi Zhang Zhaohui Qian Qi Jin nhc key laboratory of systems biology of pathogens Institute of Pathogen Biology Chinese Academy of Medical Sciences and Peking Union Medical College key laboratory of Respiratory Disease Pathogenomics Chinese Academy of Medical Sciences and Peking Union Medical College College of Animal Science and Veterinary medicine Shenyang Agricultural University

SARS-C oV and SARS-C oV-2 have been thought to originate from *** this study,we screened pharyngeal and anal swabs from 13 064 bats collected between 2016 and 2021 at 703 locations across China for sarbecoviruses,covering almost all known southern hotspots,and found 146 new bat *** analyses of all available sarbecoviruses show that there are three different lineages—L1 as SARS-CoV-related CoVs(SARSr-CoVs),L2 as SARS-CoV-2-related CoVs(SC2r-CoVs) and novel L-R(recombinants of L1 and L2)—present in Rhinolophus pusillus bats,in the mainland of *** the146 sequences,only four are ***,none belong in the L2 lineage,indicating that circulation of SC2r-C oVs in China might be very *** remaining 142 sequences belong in the L1 lineage,of which YN2020B-G shares the highest overall se quence identity with SARS-C oV(95.8 %).The observation suggests endemic circulations of SARSr-CoVs,but not SC2r-CoVs,in bats in *** analysis of the collection sites in this study,together with all published reports,indicates that SC2r-C oVs maybe mainly present in bats of Southeast Asia,including the southern border of Yunnan province,but absent in all other regions within *** contrast,SARSr-CoVs appear to have broader geographic distribution,with the highest genetic diversity and sequence identity to human sarbecoviruses along the southwest border of *** data provide the rationale for further extensive surveys in broader geographical regions within,and beyond,Southeast Asia in order to find the most recent ancestors of human sarbecoviruses.

关键词： SARS-CoV SARS-CoV-2 bats China origin tracing

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An ultrapotent RBD-targeted biparatopic nanobody neutralizes broad SARS-CoY-2 variants

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Signal Transduction and Targeted Therapy 2022年第3期7卷 780-789页

作者： Xiaojing Chi Xinhui Zhang Shengnan Pan Yanying Yu Yujin Shi Tianli Lin Huarui Duan Xiuying Liu Wenfang Chen Xuehua Yang Lan Chen Xiaoqian Dong Lili Ren Qiang Ding Jianwei Wang Wei Yang nhc key laboratory of systems biology of pathogens Institute of Pathogen BiologyChinese Academy of Medical Sciences&Peking Union Medical College100176 BeijingChina School of medicine Tsinghua University100084 BeijingChina

The wide transmission and host adaptation of SARS-CoV-2 have led to the rapid accumulation of mutations,posing significant challenges to the effectiveness of vaccines and therapeutic *** several neutralizing antibodies were authorized for emergency clinical use,convalescent patients derived natural antibodies are vulnerable to SARS-CoV-2 Spike mutation.

关键词： neutral antibodies clinical

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Discovery and characterization of novel paramyxoviruses from bat samples in China

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Virologica Sinica 2023年第2期38卷 198-207页

作者： Haoxiang Su Yuyang Wang Yelin Han Qi Jin Fan Yang Zhiqiang Wu nhc key laboratory of systems biology of pathogens Institute of Pathogen BiologyChinese Academy of Medical Sciences&Peking Union Medical CollegeBeijing100730China key laboratory of Respiratory Disease Pathogenomics Chinese Academy of Medical Sciences&Peking Union Medical CollegeBeijing100730China

Many paramyxoviruses are responsible for a variety of mild to severe human and animal *** on the novel discoveries over the past several decades,the family Paramyxoviridae infecting various hosts across the world includes 4 subfamilies,17 classified genera and 78 species ***,no systematic surveys of bat paramyxoviruses are available from the Chinese *** this study,13,064 samples from 54 bat species were collected and a comprehensive paramyxovirus survey was *** obtained 94 new genome sequences distributed across paramyxoviruses from 22 bat species in seven *** phylodynamic and phylogenetic analyses showed that there were four different lineages in the Jeilongvirus *** on available data,results of host and region switches showed that the bat colony was partial to interior,whereas the rodent colony was exported,and the felines and hedgehogs were most likely the intermediate hosts from Scotophilus *** than *** on the evolutionary trend,genus Jeilongvirus may have originated from Mus *** Australia,then transmitted to bats and rodents in Africa,Asia and Europe,and finally to bats and rodents in America.

关键词： Paramyxovirus Jeilongvirus Bat Host and region switches China

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Crystal structure of SARS-CoV-2 papain-like protease

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Acta Pharmaceutica Sinica B 2021年第1期11卷 237-245页

作者： Xiaopan Gao Bo Qin Pu Chen Kaixiang Zhu Pengjiao Hou Justyna Aleksandra Wojdyla meitian Wang Sheng Cui nhc key laboratory of systems biology of pathogens Institute of Pathogen BiologyChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijing 100730China The Swiss Light Source(SLS)at the Paul Scherrer Institut Villigen 5232Switzerland

The pandemic of coronavirus disease 2019(COVID-19)is changing the world like never *** crisis is unlikely contained in the absence of effective therapeutics or *** papain-like protease(PLpro)of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)plays essential roles in virus replication and immune evasion,presenting a charming drug *** the PLpro proteases of SARS-CoV-2 and SARS-CoV share significant homology,inhibitor developed for SARS-CoV PLpro is a promising starting point of therapeutic *** this study,we sought to provide structural frameworks for PLpro inhibitor *** determined the unliganded structure of SARS-CoV-2 PLpro mutant C111 S,which shares many structural features of SARS-CoV *** crystal form has unique packing,high solvent content and reasonable resolution 2.5 A°,hence provides a good possibility for fragment-based screening using crystallographic *** characterized the protease activity of PLpro in cleaving synthetic peptide harboring nsp2/nsp3 *** demonstrate that a potent SARS-CoV PLpro inhibitor GRL0617 is highly effective in inhibiting protease activity of SARSCoV-2 with the IC50 of 2.2?0.3 mmol/*** then determined the structure of SARS-CoV-2 PLpro complexed by GRL0617 to 2.6 A°,showing the inhibitor accommodates the S3 e S4 pockets of the substrate binding *** binding of GRL0617 induces closure of the BL2 loop and narrows the substrate binding cleft,whereas the binding of a tetrapeptide substrate enlarges the ***,our results suggest a mechanism of GRL0617 inhibition,that GRL0617 not only occupies the substrate pockets,but also seals the entrance to the substrate binding cleft hence prevents the binding of the LXGG motif of the substrate.

关键词： SARS-CoV-2 PLpro Proteinase inhibitor Crystal structure Antiviral drug Drug design

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On the origin and continuing evolution of SARS-CoV-2

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National Science Review 2020年第6期7卷 1012-1023页

作者： Xiaolu Tang Changcheng Wu Xiang Li Yuhe Song Xinmin Yao Xinkai Wu Yuange Duan Hong Zhang Yirong Wang Zhaohui Qian Jie Cui Jian Lu State key laboratory of Protein and Plant Gene Research Center for Bioinformatics School of Life Sciences Peking University CAS key laboratory of Molecular Virology & Immunology Institut Pasteur of Shanghai Chinese Academy of Sciences Center for Biosafety mega-Science Chinese Academy of Sciences University of Chinese Academy of Sciences School of Life Sciences Shanghai University nhc key laboratory of systems biology of pathogens Institute of Pathogen Biology Chinese Academy of Medical Sciences and Peking Union Medical College

The SARS-Co V-2 epidemic started in late December 2019 in Wuhan, China, and has since impacted a large portion of China and raised major global concern. Herein, we investigated the extent of molecular divergence between SARS-CoV-2 and other related coronaviruses. Although we found only 4% variability in genomic nucleotides between SARS-CoV-2 and a bat SARS-related coronavirus(SARSr-CoV;Ra TG13), the difference at neutral sites was 17%, suggesting the divergence between the two viruses is much larger than previously estimated. Our results suggest that the development of new variations in functional sites in the receptor-binding domain(RBD) of the spike seen in SARS-Co V-2 and viruses from pangolin SARSr-Co Vs are likely caused by natural selection besides recombination. Population genetic analyses of 103 SARS-CoV-2 genomes indicated that these viruses had two major lineages(designated L and S), that are well defined by two different SNPs that show nearly complete linkage across the viral strains sequenced to date. We found that L lineage was more prevalent than the S lineage within the limited patient samples we examined. The implication of these evolutionary changes on disease etiology remains *** findings strongly underscores the urgent need for further comprehensive studies that combine viral genomic data, with epidemiological studies of coronavirus disease 2019(COVID-19).

关键词： SARS-CoV-2 virus molecular evolution population genetics

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Determining antimicrobial resistance profiles and identifying novel mutations of Neisseria gonorrhoeae genomes obtained by multiplexed MinION sequencing

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Science China(Life Sciences) 2020年第7期63卷 1063-1070页

作者： Chi Zhang Feng Wang Cansheng Zhu Leshan Xiu Yamei Li Li Li Bo Liu Yizhun Li Yaling Zeng Boyang Guo Junping Peng nhc key laboratory'of systems biology of pathogens Institute of Pathogen BiologyChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijing 100176China Shenzhen Center for Chronic Disease Control Shenzhen 518020China Department of STD laboratory Shaanxi Provincial Institute for Skin Disease and STD ControlXi'an 710003China

Gonorrhea is one of the most common sexually transmitted diseases worldwide. To cure infection and prevent transmission,timely and appropriate antimicrobial therapy is necessary. Unfortunately, Neisseria gonorrhoeae, the etiological agent of gonorrhea, has acquired nearly all known mechanisms of antimicrobial resistance(AMR), thereby compromising the efficacy of antimicrobial therapy. Treatment failure resulting from AMR has become a global public health concern. Whole-genome sequencing is an effective method to determine the AMR characteristics of N. gonorrhoeae. Compared with next-generation sequencing, the MinION sequencer(Oxford Nanopore Technologies(ONT)) has the advantages of long read length and portability. Based on a pilot study using MinION to sequence the genome of N. gonorrhoeae, we optimized the workflow of sequencing and data analysis in the current study. Here we sequenced nine isolates within one flow cell using a multiplexed sequencing strategy. After hybrid assembly with Illumina reads, nine integral circular chromosomes were obtained. By using the online tool Pathogenwatch and a BLAST-based workflow, we acquired complete AMR profiles related to seven classes of antibiotics. We also evaluated the performance of ONT-only assemblies. Most AMR determinants identified by ONT-only assemblies were the same as those identified by hybrid assemblies. Moreover, one of the nine assemblies indicated a potentially novel antimicrobial-related mutation located in mtrR which results in a frame-shift, premature stop codon, and truncated *** addition, this is the first study using the MinION sequencer to obtain complete genome sequences of N. gonorrhoeae strains which are epidemic in China. This study shows that complete genome sequences and antimicrobial characteristics of *** can be obtained using the MinION sequencer in a simple and cost-effective manner, with hardly any knowledge of bioinformatics required. More importantly, this strategy provid

关键词： nanopore sequencing MinION sequencer Neisseria gonorrhoeae whole-genome sequencing antimicrobial resistance

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