Comprehensive investigation of HBV-related hepatocellular carcinoma and choice of anti-HBV therapy

Chronic hepatitis B virus(HBV)infection is a global public health problem,which can cause chronic hepatitis,liver cirrhosis,hepatocellular carcinoma(HCC),and other diseases.Antiviral therapy is the most critical measure to slow down the progression of chronic hepatitis B,prevent or delay cirrhosis,HCC,and other kinds of liver decompensation events.At present,the anti-hepatitis B virus drugs are mainly nucleoside(acid)analogues(NAs)and interferon.Each kind of antiviral drug has different effects on the clinical outcome of hepatitis B patients(such as HCC).In this paper,we discussed the biological characteristics,natural course and prognosis of HBV infection,the mechanism of HBV-related HCC,the effect of different antiviral drugs on patients’outcome,predictive biomarkers and model for HBV clinical outcome,predictors of sustained response and recurrence after withdrawal of antiviral therapy,consideration of expanding therapeutic indications and antiviral therapy,hoping to give a hand to the clinical diagnosis and treatment of HBV.

Biosafety and biobanking: Current understanding and knowledge gaps

Infectious disease outbreaks,such as'Coronavirus disease 20190(COVID-19),can constitute major global health threats with far-reaching consequences.As outbreaks develop,the international scientific community must provide high-quality scientific research-ready biological samples to solve the existing clinical and epidemiological questions to better combat the pandemic.Such examples are provided by dedicated biobank facilities,the latter collecting increasingly high volumes of biological samples.However,the more significant concentrations of infectious or potentially infectious biological materials can create a safety risk.The current short report describes the first attempt to identify the published scientific works on biobanking and safety.Three broad thematic areas have been identified:the physical security relevant to staff and sample integrity,the data safety aspects,and the governance parameters relating to the previous two.While the current publications reflect a broad alignment with existing standards and best practices in the biobanking field,they also demonstrate an opportunity for further in-depth work on this field in the post-COVID-19 era.

Caspase-8 activation regulates enterovirus D68 infection-induced inflammatory response and cell death

Enterovirus D68 (EV-D68) infection causes severe acute respiratory infection and severe neurological complications, such as acute flaccid myelitis (AFM), in children. However, although EV-D68 has pandemic potential, no effective drugs or vaccines are currently clinically available. Furthermore, EV-D68 infection-induced inflammatory response and cell death are not fully understood. In this study, we demonstrated that several inflammatory cytokines were upregulated in a multiplicity of infection (MOI) dependent manner in EV-D68-infected human rhabdomyosarcoma (RD) cells. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) confirmed that tumor necrosis factor-α (TNF-α), interleukin 6 (IL-6), C-C motif chemokine ligand-5 (CCL-5), and CXC motif chemokine ligand-5 (CXCL-5) mRNA levels were highly upregulated after EV-D68 infection. IL-1β processing and maturation mediated by caspase-8 was inhibited by the caspase-8 inhibitor Z-IETD-FMK. EV-D68 infection activates caspase-8 to mediate IL-1β maturation and secretion. Additionally, EV-D68 activated cell death-related proteins such as caspase-3, poly (ADP-ribose) polymerase 1 (PARP-1), phosphorylation of Mixed Lineage Kinase domain-like protein (pMLKL), and gasdermin E (GSDME). Thus, EV-D68 infection activates caspase-8, which triggers the necroptosis and apoptosis pathways. Overall, our data suggest that caspase-8 activation is associated with the inflammatory response and cell death in EV-D68-infected RD cells. This mechanism represents a novel target for the treatment of EV-D68 infection by inhibiting caspase-8 activation.

Review of pharmacologic and immunologic agents in the management of COVID-19

The novel coronavirus disease 2019(COVID-19)is the third coronavirus outbreak in the last two decades.Emerging and re-emerging infections like COVID-19 pose serious challenges of the paucity of information and lack of specific cure or vaccines.This leaves utilisation of existing scientific data on related viral infections and repurposing relevant aetiologic and supportive therapies as the best control approach while novel strategies are developed and trialled.Many promising antiviral agents including lopinavir,ritonavir,remdesivir,umifenovir,darunavir,and oseltamivir have been repurposed and are currently trialled for the care for COVID-19 patients.Adjunct therapies for the management of symptoms and to provide support especially in severe and critically ill patients have also been identified.This review provides an appraisal of the current evidence for the rational use of frontline therapeutics in the management of COVID-19.It also includes updates regarding COVID-19 immunotherapy and vaccine development.

Temporal dynamics of SARS-CoV-2 genome mutations that occurred in vivo on an aircraft

We analyzed variations in the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)genome during a flight-related cluster outbreak of coronavirus disease 2019(COVID-19)in Shenzhen,China,to explore the characteristics of SARS-CoV-2 transmission and intra-host single nucleotide variations(iSNVs)in a confined space.Thirty-three patients with COVID-19 were sampled,and 14 were resampled 3-31 days later.All 47 nasopharyngeal swabs were deep-sequenced.iSNVs and similarities in the consensus genome sequence were analyzed.Three SARS-CoV-2 variants of concern,Delta(n=31),Beta(n=1),and C.1.2(n=1),were detected among the 33 patients.The viral genome sequences from 30 Delta-positive patients had similar SNVs;14 of these patients provided two successive samples.Overall,the 47 sequenced genomes contained 164 iSNVs.Of the 14 paired(successive)samples,the second samples(T2)contained more iSNVs(median:3;95%confidence interval[95%CI]:2.77-10.22)than did the first samples(T1;median:2;95%CI:1.63-3.74;Wilcoxon test,P=0.021).38 iSNVs were detected in T1 samples,and only seven were also detectable in T2 samples.Notably,T2 samples from two of the 14 paired samples had additional mutations than the T1 samples.The iSNVs of the SARS-CoV-2 genome exhibited rapid dynamic changes during a flight-related cluster outbreak event.Intra-host diversity increased gradually with time,and new site mutations occurred in vivo without a population transmission bottleneck.Therefore,we could not determine the generational relationship from the mutation site changes alone.

HLA-E-restricted Hantaan virus-specific CD8^(+)T cell responses enhance the control of infection in hemorrhagic fever with renal syndrome

Infection with the Hantaan virus(HTNV)may result in severe hemorrhagic fever with renal syndrome(HFRS).The functions of HLA-E-restricted CD8^(+)T lymphocytes in virus control and vaccine development have recently received increased attention.The purpose of this research is to discover HLA-E-restricted CD8^(+)T cell epitopes on HTNV as well as the features of these epitope-specific CD8^(+)T cells in HFRS patients.To anticipate HLA-Erestricted HTNV epitopes,the NetMHCpan servers were utilized.The K562/HLA-E cell binding test and the enzyme-linked immunospot assay were used to confirm epitope binding to HLA-E.The number and features of HLA-E-restricted epitope-specific CD8^(+)T lymphocytes in HFRS patients were investigated using tetramer staining,intracellular cytokine labeling,proliferation,and cytotoxicity assays.Six HTNV-derived HLA-Erestricted CD8^(+)T cell epitopes were found in this study.In mild/moderate HFRS patients,the frequency of HLA-E-restricted epitope-specific CD8^(+)T cells was greater than in severe/critical patients.CD38+HLA-DR+HLA-E-restricted CD8^(+)T cells were identified.Meanwhile,CD45RA^(+)CCR7^(-)effector memory-re-expressing CD45RA T cells with early and intermediate maturation and differentiation characteristics predominated.Notably,CD8^(+)T cells from milder HFRS patients produced more interferon-γ,interleukin-2,and granzyme B,had a stronger proliferative potential,and were inversely linked with the amount of plasma HTNV virus load.Furthermore,HLA-E-restricted epitope-specific CD8^(+)T cells demonstrated improved cytotoxic activity in vitro during the acute stage of HFRS.Taken together,the findings demonstrate the protective effects of HLA-E-restricted CD8^(+)T cells during HTNV infection,suggesting that HLA-E-targeted vaccines against HTNV might be developed for HLA-diverse populations.

Surveillance of enteric pathogens in imported seafood and environmental surfaces in five seafood markets before the outbreak of COVID-19

To monitor the presence of enteric pathogens in imported seafood,a total of 140 seafood samples imported from eight overseas countries were collected from Beijing,Dalian,Shanghai,Guangzhou,and Wuhan seafood markets from June to November 2019.Additionally,116 viral,environmental swab samples were also collected from the Wuhan and Guangzhou seafood markets.Five typical enteric bacterial pathogens(Aeromonas spp.,Shigella spp.,Salmonella spp.,Vibrio spp.,and Listeria monocytogenes)and four viruses(Rotavirus,Norovirus,Astrovirus,and Sapovirus)were detected positive.Results showed that eight Vibrio parahaemolyticus isolates appeared in seafood imported to Dalian,Wuhan,Shanghai,Guangzhou,and Beijing.In contrast,Vibrio fluvialis and Aeromonas were isolated in another two samples.Norovirus was detected in one oyster sample imported from France and environmental surface in Guangzhou.The remaining pathogens were negative in all the samples being tested.With 120 V.parahaemolyticus isolates from the above countries,the genomic analysis revealed that sequence type ST1152 isolates imported from Canada were clustered with two V.parahaemolyticus isolates from Canada.This study presented the first microbiological analysis of the Wuhan seafood market before the outbreak of COVID-19,which demonstrated that supervision should be strengthened to prevent enteric pathogens via imported seafood.

Protein amplification technology:New advances in human prion disease diagnosis

Early and accurate diagnosis of human prion diseases is a long-standing difficulty.Currently,the definitive diagnosis of human prion diseases relies on pathognomonic histological features or PrPSc detection of patients'brain tissue biopsy or autopsy samples,which is not feasible in most cases.Therefore,clinical diagnosis mainly relies on the combinations of the patient’s clinical symptoms.MRI and EEG are used to check for brain damage and detect surrogate markers such as the 14-3-3 protein in Cerebrospinal fluid(CSF),but this is often challenging.In recent years,the development of in vitro cell-free conversion techniques,such as technologies protein misfolding cyclic amplification(PMCA)and real-time quaking-induced conversion(RT-QuIC),have extensively promoted the diagnosis of human prion diseases.PMCA has high diagnostic accuracy in the blood,CSF,and urine samples of variant creutzfeldt–jakob disease(vCJD)patients.Again,RT-QuIC has high diagnostic accuracy for cerebrospinal fluid,olfactory mucosa,and skin samples of sporadic Creutzfeldt Jakob Disease(sCJD)patients.Applying these two technologies is of great significance to the early clinical diagnosis of human prion diseases and the reduction of blood-borne and iatrogenic transmission of prion.

A novel method to assess antibody-dependent cell-mediated cytotoxicity against influenza A virus M2 in immunized murine models

The matrix protein 2 (M2) is a preferred target for developing a universal vaccine against the influenza A virus (IAV). This study aimed to develop a method for assessing antibody-dependent cell-mediated cytotoxicity (ADCC) associated with M2-based immunization in mice. We first established a stable cell line derived from mouse lymphoma cells (YAC-1) expressing M2 of H3N2. This cell line, designated as YAC-1-M2, was generated using a second-generation lentiviral tricistronic plasmid system to transduce the M2 gene into YAC-1 cells. The ADCC effect induced by polyclonal antibodies targeting matrix protein 2 ectodomain (M2e) was demonstrated by YAC-1-M2 cell lysis by natural killer cells (NK) derived from mice, in the presence of anti-M2 antibodies obtained from mice immunized with an mRNA vaccine based on M2e. This ADCC effect was found to be stronger compared to the effect induced by monoclonal antibodies (14C2) against M2. Moreover, the ADCC effect was enhanced as the effector-to-target ratio of NK to YAC-1-M2 cells increased. In conclusion, we established a novel method to detect ADCC of M2 of IAV, which paves the way for the development of an M2-based universal vaccine against IAV and an in-depth analysis of its mechanism of broad-spectrum immune protection in mice.

Analysis of the distribution characteristics of enterovirus types based on environmental surveillance from 2013 to 2021 in Fujian Province, China

Environmental surveillance (ES) is a useful approach for monitoring circulating viruses, including polioviruses (PVs) and non-polio enteroviruses (NPEVs). In this study, the results of nine years of ES from 2013 to 2021 at six sampling sites in three cities in Fujian Province, China, were summarized. It showed that the sewage samples contained abundant viruses, but the positive rate was affected by different sampling sites. From the 520 samples, 431 PVs, 1,713 NPEVs, and 281 human adenoviruses (HAdVs) were isolated. PV isolates had been markedly affected following the adjustment of the immunization strategy. All but one PV isolate were Sabin-like strains without wild PVs. One isolate was vaccine-derived PV type 3 with 10 variation points in theVP1 region. After May 2016, PV type 2 was no longer detected, and PV type 3 became a superior serotype. Of 1,713 NPEVs, 24 serotypes were identified, including echovirus11 (E11), E6, coxsackievirus B3 (CVB3), CVB5, E7, and E3 were the predominant serotypes (37.65%, 20.96%, 11.50%, 8.87%, 8.23%, and 7.06%, respectively). The temporal dynamic of the six common serotypes was inconsistent. E3 was frequently isolated, but the number of isolates was low, with no obvious peaks. E6, E7, and CVB3 exhibited periodic changes with a high peak every three to four years, and E11 only had one high peak lasting four years. Summer-fall peaks of the echoviruses and spring-winter peaks of CVB were observed in the monthly distribution of virus isolation. The infectious isolates of various serotypes of different species identified from the sewage samples showed that ES is an essential part of pathogen surveillance.

Assessment of biosafety and biorisk management practices among medical laboratory students in two institutions in Uganda

Medical laboratory workers handle clinical specimens,which are a threat of exposure to infectious agents.Notably,medical laboratory science students report for internships with only theoretical knowledge of biosafety and biorisk management practices,predisposing them to a higher risk of laboratory hazards.In this study,we assessed the influence of entry-level students'adherence to practices and attitudes towards biosafety and biorisk management during the Internship.An online survey tool was used to explore the practices and attitudes towards laboratory biosafety and risk management.Of the 96 students,60(62.5%)anonymous responses were received,and of these,60.3%were direct entrants,and 32.8%were diploma entrants.Most(91.7%)of the students attended hospital internships,with 60.2%in Biosafety Level(BSL)-2 laboratories and 70.2%rotating in all the core areas of laboratory medicine.The 8.3%who did not attend any internship were under the direct entry category.Exposure to biohazards was not significantly associated with laboratory safety level and student entry category(P>0.05).Recommended laboratory biosafety practices were not significantly associated with the safety level of the laboratory and student entry category(P>0.05).Poor attitudes towards certain laboratory biosafety practices were not significantly associated with the biosafety level of the training laboratory(P>0.05),whereas training(P=0.021)and clean-up procedures(P=0.048)were associated with laboratory safety levels,respectively.The direct entrants had no access to BSL-3 laboratories,and this category of students had a negative attitude towards internship attendance.Therefore,there is a need to create a multi-channel full range laboratory biosafety and biorisk management teaching reforms based on practical application,real case studies,and laboratory simulation to be incorporated into the curriculum to benefit the direct entrant.

The impact of sample processing on the rapid antigen detection test for SARS-CoV-2: Virus inactivation, VTM selection, and sample preservation

Many factors have been identified as having the ability to affect the sensitivity of rapid antigen detection(RAD)tests for severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).This study aimed to identify the impact of sample processing on the sensitivity of the RAD tests.We explored the effect of different inactivation methods,viral transport media(VTM)solutions,and sample preservation on the sensitivity of four RAD kits based on two SARS-CoV-2 strains.Compared with non-inactivation,heat inactivation significantly impacted the sensitivity of most RAD kits;however,β-propiolactone inactivation only had a minor effect.Some of the VTM solutions(VTM2,MANTACC)had a significant influence on the sensitivity of the RAD kits,especially for low viral-loads samples.The detection value of RAD kits was slightly decreased,while most of them were still in the detection range with the extension of preservation time and the increase of freeze–thaw cycles.Our results showed that selecting the appropriate inactivation methods and VTM solutions is necessary during reagent development,performance evaluation,and clinical application。

Advanced researches on the inhibition of influenza virus by Favipiravir and Baloxavir

Anti-influenza drugs are one of themost critical pathways for control of influenza virus infection.Drugs that have been developed or are developing may function via different mechanisms,and so far,inhibitors of influenza virus polymerase are among the most promising types of drugs.Favipiravir and Baloxavir,also named T-705 and Xofluza respectively,have been approved for influenza treatment in Japan and the United States.Favipiravir effectively and selectively inhibits the RNA-dependent RNA polymerase(RdRp)of RNA viruses while Baloxavir specifically targets the cap-dependent endonuclease PA of influenza viruses.These two drugs have been suggested as the first candidate drugs for influenza infection treatment,especially for strains resistant to other anti-influenza drugs.This review will focus on the pharmaceutical mechanisms and anti-influenza activity of these two drugs.

Laboratory biosafety guide for 2019-nCoV (Second Edition)

According to the biological characteristics,epidemiological features,pathogenicity,clinical manifestation and other related information of the novel coronavirus(2019-nCoV)currently available,the pathogen should be provisionally managed as the Risk Group 2 pathogenic microorganisms in the classification of pathogenic microorganisms.

Use of antimicrobials in food animals and impact of transmission of antimicrobial resistance on humans

Antimicrobial resistance leads to failure of clinical antimicrobial therapy,and has raised urgent global public health concern.Humans can acquire antimicrobial resistance fromdrugs through the food chain or the environment(contaminated water,air,soil,or manure).While antimicrobials have been regular supplements in animal feed that maintain health and improve productivity of livestock,their over-use in feeding forage has led to a rise in antibacterial resistance.This review summarizes the current use of antimicrobials in livestock,the harmful effects of antimicrobial resistance,and the comprehensive combat measures.

Corrigendum to "Case report for human infection with a highly pathogenic avian influenza A(H5N6) virus in Beijing, China 2019" [Biosafety and Health 2 (2020) 49-52]

The authors regret that an ethics statement was missing from the above article.The Ethics Statement section should be added as follows:Ethics statement As a public health response to an emerging infectious disease outbreak,written informed consent could be waived.Data and sample collection of the human case were determined by the National Health Commission of the People's Republic of China to be part of a continuing public health outbreak investigation and were exempt from institutional review board approval.The authors would like to apologize for any inconvenience caused.

Development and effectiveness of pseudotyped SARS-CoV-2 system as determined by neutralizing efficiency and entry inhibition test in vitro

With the development of the COVID-19 epidemic,there is an urgent need to establish a system for determining the effectiveness and neutralizing activity of vaccine candidates in biosafety level 2(BSL-2)facilities.Previously,researchers had developed a pseudotyped virus systemfor SARS-CoV andMERS-CoV,based onHIV-1 core,bearing virus spike protein.During the development of a pseudotyped SARS-CoV-2 system,a eukaryotic expression plasmid expressing SARSCoV-2 spike(S)protein was constructed and then co-transfectedwith HIV-1 based plasmid which containing the firefly luciferase reporter gene,into HEK293T cells to prepare the pseudotyped SARS-CoV-2 virus(ppSARS-2).We have successfully established the pseudotyped SARS-CoV-2 system for neutralization and entry inhibition assays.Huh7.5 cell line was found to be the most susceptible to our pseudotyped virus model.Different levels of neutralizing antibodies were detected in convalescent serum samples of COVID-19 patients using ppSARS-2.The recombinant,soluble,angiotensin-converting enzyme 2 protein was found to inhibit the entry of ppSARS-2 in Huh7.5 cells effectively.Furthermore,the neutralization results for ppSARS-2 were consistent with those of live SARS-CoV-2 and determined using the serum samples fromconvalescent patients.In conclusion,we have developed an easily accessible and reliable tool for studying the neutralizing efficiency of antibodies against SARS-CoV-2 and the entry process of the virus in a BSL-2 laboratory.

Development and evaluation of a quadruple real-time fluorescence-based quantitative reverse transcription polymerase chain reaction assay for detecting Langya, Mojiang, Nipah, and Cedar viruses

The emerging viruses within the genus Henipavirus in the family Paramyxoviridae pose a great threat to public biosafety.To develop a quadruple real-time fluorescence-based quantitative reverse transcription polymerase chain reaction(qRT-PCR)assay is pivotal for the early warning of the potential of zoonotic infectious diseases.Specific primers and probes were designed for the relatively conserved regions based on whole genome sequences of Langya virus(LayV),Mojiang virus(MojV),Nipah virus(NiV),and Cedar virus(CedV),followed by the establishment of a quadruple real-time fluorescence-based qRT-PCR detection method.No cross-reactivity was observed with other viral nucleic acids.The optimal linear detection range for LayV,MojV,NiV,and CedV was 10^(1)-10^(8)copies/μL,and the lower limit of detection was 10 copies/μL.Three different DNA concentrations of LayV,MojV,NiV,and CedV(10^(4),10^(5),and 10^(6)copies/μL)were tested 14 times,achieving good repeatability.The standard deviation of the cycle threshold values for each concentration was<0.5 and the coefficient of variation was<3%.Furthermore,the amplification efficiency of quadruple real-time fluorescence-based qRT-PCR was>90%,and the correlation coefficient was>0.99.The established quadru-ple real-time fluorescence-based qRT-PCR assay for the detection of LayV,MojV,NiV,and CedV exhibits good sensitivity,specificity,and repeatability.Therefore,it can be used to detect Henipavirus and other related clinical specimens.

Comprehensive interactome analysis of the spike protein of swine acute diarrhea syndrome coronavirus

Swine acute diarrhea syndrome coronavirus(SADS‐CoV)is a recently discovered coronavirus that causes severe and acute diarrhea and rapid weight loss in piglets.SADS‐CoV was reported to be capable of infecting cell lines derived from diverse species,including bats,mice,hamsters,rats,chickens,pigs,nonhuman primates,and humans,implying its high risk of cross‐species infection.However,its receptor is still unknown.In this study,the receptor‐binding domain of the SADS‐CoV spike(S)protein was purified and then subjected to affinity purification(AP)‐coupled mass spectrometry(MS)‐based proteomic analysis to identify the interactors of the SADS‐CoV S protein.Forty‐three host proteins were identified,and a Gene Ontology analysis indicated that these interactors can be grouped into categories such as“cell‐cell adhesion”,“translation”“viral transcription”,suggesting that these processes may participate in the SADS‐CoV life cycles.RNA interference‐based screening of these interactors indicated that PPIB and vimentin can affect SADS‐CoV replication.Our study provides an overarching view into the host interactome of the SADS‐CoV S protein and highlights potential targets for the development of therapeutics against SADS‐CoV.

Host protein ABCE1 interacts with the viral phosphoprotein and promotes rabies virus replication

Rabies virus(RABV)phosphoprotein(P)plays an important role in disrupting host interferon(IFN)-mediated antiviral immune response.ABCE1 is known as an RNase L inhibitor that negatively regulates the 2′,5′-oligoadenylate(2-5A)/RNase L antiviral system related to the IFN signaling pathway.In this study,we screened the host factors associated with RABV P protein,while focusing on ABCE1 because of its role in the life cycle of several viruses.Our results showed that knockout of ABCE1 in HEK293T cells inhibited RABV replication.In contrast,the overexpression of ABCE1 in HEK293T cells enhanced RABV replication.Notably,the co-immunoprecipitation assay showed that ABCE1 interacted with RABV P protein.Since ABCE1 and RABV P proteins are related to the IFN signaling pathway,we propose that the interaction of viral P protein with ABCE1 leads to the disruption of host antiviral immune response.In summary,our research showed that ABCE1 is an important host protein that interacts with viral P protein and regulates RABV replication.Moreover,the interaction between ABCE1 and RABV P protein may facilitate the viral P protein-mediated disruption of host antiviral immune response.