Objective:To characterize the infection patterns and growth characteristics of the Zika virus(ZIKV)strain JMB-185 from Indonesia in various mammalian cell lines.Methods:ZIKV was grown in human(A549,HEK293,HepG2,Huh7,J...Objective:To characterize the infection patterns and growth characteristics of the Zika virus(ZIKV)strain JMB-185 from Indonesia in various mammalian cell lines.Methods:ZIKV was grown in human(A549,HEK293,HepG2,Huh7,Jurkat,and THP-1)and non-human mammalian(RAW264.7,Vero,and Vero76)cell lines.Viral replication kinetics were measured using plaque assay,while intra-and extracellular viral RNA concentrations were assessed using RT-PCR.Flow cytometry was used to quantify the infected cells and cell viability was measured using an MTT assay.The ability of ZIKV to infect cell lines was visualized using a fluorescence immunostaining assay.Results:This ZIKV strain preferentially infected the lung,kidney,and liver cell lines A549,HEK293,Huh7,Vero,and Vero76,but not the immune cells Jurkat,RAW264.7,and THP-1.By contrast,the ZIKV showed no sign of infection in HepG2 cells,while maintaining viral titer over 3 days post-infection,with no infection recorded in immunostaining,no increase in viral RNA,and no indication of cell deterioration.Conclusions:The Indonesian ZIKV strain has a similar infection profile as other strains,except for its poor infectivity on HepG2 cells.Information on the growth characteristics of Indonesia ZIKV will help expand our understanding of the biology of ZIKV which will be useful for various applications including antiviral discovery.展开更多
This study explores the antiviral properties of high-voltage low-frequency electric field exposure on the replication of human viruses, including Herpes Simplex Virus type 1 (HSV-1), Human Coronavirus OC43 (HCoV OC43)...This study explores the antiviral properties of high-voltage low-frequency electric field exposure on the replication of human viruses, including Herpes Simplex Virus type 1 (HSV-1), Human Coronavirus OC43 (HCoV OC43), and Influenza A virus (A H1N1). Using the HealectricsTM device (model S02), which operates by applying high-voltage direct current (30 - 50 kV) with a polarity change frequency of ~0.2 Hz, we investigated the impact on viral infectivity and host cell viability. Virus cultures were exposed to electric fields during different stages: virion adsorption (0 - 1 hour), intracellular replication (1 - 8 hours), and both stages. Viral infectivity was assessed through titration, and cytotoxic effects were evaluated using MTT assays. Electric field exposure significantly reduced viral infectivity, particularly during the combined sorption and replication stages, with up to a 90% decrease in viral activity. Among the viruses tested, HCoV OC43 showed the least sensitivity, with a reduction in viral activity by a factor of 5. Comparisons revealed statistically significant reductions for influenza and herpes viruses, and a trend towards significance for HCoV OC43. The electric field treatment did not significantly affect the viability of Vero and MDCK cells, indicating the method’s safety. Our findings suggest that high-voltage low-frequency electric fields can effectively reduce viral infectivity and may serve as a potential therapeutic and preventive measure against a wide range of membrane-bound viruses, including SARS-CoV-2.展开更多
Effective oral infection is set off by interaction of a group of conserved per os infectivity factors(PIFs) with larval midgut columnar epithelial cells. We constructed pseudotyped viruses by substituting pif1, pif2 o...Effective oral infection is set off by interaction of a group of conserved per os infectivity factors(PIFs) with larval midgut columnar epithelial cells. We constructed pseudotyped viruses by substituting pif1, pif2 or pif3 genes of Helicoverpa armigera nucleopolyhedrovirus(Hear NPV) with their homologs from Mamestra bracissae multiple nucleopolyhedrovirus and tested their infectivity to tissue culture cells and to larvae. Transfection and infection assays revealed that all recombinant viruses generated infectious budded virus in both cell culture and in larvae. Electron microscopy showed synthesized occlusion body and occlusion derived virus(ODV) were morphologically indistinguishable from those of the parental virus. By contrast, feeding assays revealed that pseudotyped viruses could not rescue oral infectivity except for pif3 pseudotyped virus that only partially rescued oral infectivity but at a mortality rate much lower than that of the parental Hear NPV. Consistent with the bioassay result, PIF complex was detected in ODVs of pif3 pseudotyped virus only but not in pif1 or pif2 pseudotyped viruses. Our results suggest that PIF complex is essential for oral infectivity, and in the formation of the PIF complex, PIF1, 2 are virus-specific while PIF3 does not appear to be as specific and can function in heterologous environment, albeit to a much more limited extent.展开更多
The phasing out of protective measures by governments and public health agencies, despite continued seriousness of the coronavirus pandemic, leaves individuals who are concerned for their health with two basic options...The phasing out of protective measures by governments and public health agencies, despite continued seriousness of the coronavirus pandemic, leaves individuals who are concerned for their health with two basic options over which they have control: 1) minimize risk of infection by being vaccinated and by wearing a face mask when appropriate, and 2) minimize risk of transmission upon infection by self-isolating. For the latter to be effective, it is essential to have an accurate sense of the probability of infectivity as a function of time following the onset of symptoms. Epidemiological considerations suggest that the period of infectivity follows a lognormal distribution. This proposition is tested empirically by construction of the lognormal probability density function and cumulative distribution function based on quantiles of infectivity reported by several independent investigations. A comprehensive examination of a prototypical ideal clinical study, based on general statistical principles (the Principle of Maximum Entropy and the Central Limit Theorem) reveals that the probability of infectivity is a lognormal random variable. Subsequent evolution of new variants may change the parameters of the distribution, which can be updated by the methods in this paper, but the form of the probability function is expected to remain lognormal as this is the most probable distribution consistent with mathematical requirements and available information.展开更多
The appearance of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)variant Omicron(B.1.1.529)has caused panic responses around the world because of its high transmission rate and number of mutations.This rev...The appearance of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)variant Omicron(B.1.1.529)has caused panic responses around the world because of its high transmission rate and number of mutations.This review summarizes the highly mutated regions,the essential infectivity,transmission,vaccine breakthrough and antibody resistance of the Omicron variant of SARSCoV-2.The Omicron is highly transmissible and is spreading faster than any previous variant,but may cause less severe symptoms than previous variants.The Omicron is able to escape the immune system’s defenses and coronavirus disease 2019 vaccines are less effective against the Omicron variant.Early careful preventive steps including vaccination will always be key for the suppression of the Omicron variant.展开更多
Papaya leaf curl China virus (PaLCuCNV) was previously reported as a distinct begomovirus infecting papaya in southern China. Based on molecular diagnostic survey, 13 PaLCuCNV isolates were obtained from tomato plants...Papaya leaf curl China virus (PaLCuCNV) was previously reported as a distinct begomovirus infecting papaya in southern China. Based on molecular diagnostic survey, 13 PaLCuCNV isolates were obtained from tomato plants showing leaf curl symptoms in Henan and Guangxi Provinces of China. Complete nucleotide sequences of 5 representative isolates (AJ558116, AJ558117, AJ704604, FN256260, and FN297834) were determined to be 2738–2751 nucleotides, which share 91.7%–97.9% sequence identities with PaLCuCNV isolate G2 (AJ558123). DNA-β was not found to be associated with PaLCuCNV isolates. To investigate the infectivity of PaLCuCNV, an in-fectious clone of PaLCuCNV-[CN:HeNZM1] was constructed and agro-inoculated into Nicotiana benthamiana, N. tabacum Samsun, N. glutinosa, Solanum lycopersicum and Petunia hybrida plants, which induced severe leaf curling and crinkling symptoms in these plants. Southern blot analysis and polymerase chain reaction (PCR) indicated a systemic infection of test plants by the agro-infectious clone.展开更多
Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)is a highly contagious virus that can transmit through respiratory droplets,aerosols,or contacts.Frequent touching of contaminated surfaces in public areas is...Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)is a highly contagious virus that can transmit through respiratory droplets,aerosols,or contacts.Frequent touching of contaminated surfaces in public areas is therefore a potential route of SARS-CoV-2 transmission.The inanimate surfaces have often been described as a source of nosocomial infections.However,summaries on the transmissibility of coronaviruses from contaminated surfaces to induce the coronavirus disease 2019 are rare at present.This review aims to summarize data on the persistence of different coronaviruses on inanimate surfaces.The literature was systematically searched on Medline without language restrictions.All reports with experimental evidence on the duration persistence of coronaviruses on any type of surface were included.Most viruses from the respiratory tract,such as coronaviruses,influenza,SARS-CoV,or rhinovirus,can persist on surfaces for a few days.Persistence time on inanimate surfaces varied from minutes to up to one month,depending on the environmental conditions.SARSCoV-2 can be sustained in air in closed unventilated buses for at least 30 min without losing infectivity.The most common coronaviruses may well survive or persist on surfaces for up to one month.Viruses in respiratory or fecal specimens can maintain infectivity for quite a long time at room temperature.Absorbent materials like cotton are safer than unabsorbent materials for protection from virus infection.The risk of transmission via touching contaminated paper is low.Preventive strategies such as washing hands and wearing masks are critical to the control of coronavirus disease 2019.展开更多
A new morphological variant strain of microsporidium was produced by infecting the mulberry looper, Hemerophilaatrilineata [Phthonandria atrilineata], with Nosema bombycis successively for 24 times, and named 24Nbh. C...A new morphological variant strain of microsporidium was produced by infecting the mulberry looper, Hemerophilaatrilineata [Phthonandria atrilineata], with Nosema bombycis successively for 24 times, and named 24Nbh. Comparativestudies on morphology, infectivity and spore surface protein were conducted. 24Nbh was short and wide, and had asignificant difference (P<0.01) over the Nb spores. The infectivity tests conducted on second instar silkworm larvaeshowed that IC50 of 24Nbh was 1.98104 spores mL-1 and of Nb was 1.72103 spores mL-1, thus indicating that the infectivityof Nb decreased 11.5 times after multiplying in mulberry looper for 24 times. The IC50 of spores from silkworm infected with24 Nbh was 6.9 times less than Nb, showing that the infectivity of 24Nbh spores rejuvenated very fast when reinfected tosilkworms, further more, the length and width of such spore was larger than 24Nbh (P<0.01) and smaller than Nb (P<0.05).The SDS-PAGE profiles of Nb and 24Nbh were generally the same, 4 distinct proteins of 12, 17, 30, 33 kDa were obtainedwith difference in quantity. When 120 g of protein was applied for 2D-PAGE, five suspected different proteins withdifference in quantity were observed. These results demonstrate that these differential proteins maybe associated withvariation in infectivity of the spores.展开更多
The ultrastructure of porcine kidney(PK)-15 cells was examined after lipofectamine-aided transfection of the molecular clone of the P1 agent.PK-15 cells transfected with the tandem dimer of the P1molecular DNA clone h...The ultrastructure of porcine kidney(PK)-15 cells was examined after lipofectamine-aided transfection of the molecular clone of the P1 agent.PK-15 cells transfected with the tandem dimer of the P1molecular DNA clone had numbers of intracytoplasmic inclusions,and a few cells had intranuclear inclusions.Intracytoplasmic inclusions were round to oval and 0.1-0.3μm in diameter,and intranuclear inclusions,which were more electron dense,were of two general types:the first were round and small(0.1μm approximately)and the second were hexagonal and larger(0.4-0.8μm in diameter).Cells transfected with the tandem dimer of the P1 molecular DNA clone tested positive for P1 DNA at passage 5.This is the first report that the P1 molecular clone has infectivity in vitro and it will provide fundamental materials for further study of the biological characterization of P1.展开更多
Hepatitis B virus(HBV)genotype B and C are two major genotypes that are prevalent in Asia and differ in natural history and disease progression.The impact of HBV genotypes on viral replication and protein expression h...Hepatitis B virus(HBV)genotype B and C are two major genotypes that are prevalent in Asia and differ in natural history and disease progression.The impact of HBV genotypes on viral replication and protein expression has been explored by the transfection of hepatoma cells with replication-competent HBV DNA,which mimics the later stages of the viral life cycle.However,the influence of HBV genotypes on the early events of viral infection remains undetermined,mainly due to the difficulties in obtaining sufficient infectious viral particles for infection assays.Here,we report that a high-titer HBV inoculum can be generated from the transient transfection-based cell model after optimizing transfection conditions and modifying the HBV-expressing construct.By performing in vitro infection assays using transiently transfected derived viruses,we found that clinical genotype C isolates possessed higher infectivity than genotype B isolates.Moreover,we identified a naturally occurring mutation sL21S in small hepatitis B surface protein,which markedly decreased the infectivity of HBV genotype C isolates,but not that of genotype B isolates.In summary,using infectious viral particles provided by the optimized transient transfection-based cell model,we have been able to investigate a wide range of HBV variants on viral infectivity,which may contribute to our understanding of the reasons for different clinical outcomes in HBV infections and the development of therapeutic drugs targeting the early stages of HBV life cycle.展开更多
The Enterovirus 71(EV71)VP4 is co-translationally linked to myristic acid at its amino-terminal glycine residue.However,the role of this myristoylation in the EV71 life cycle remains largely unknown.To investigate thi...The Enterovirus 71(EV71)VP4 is co-translationally linked to myristic acid at its amino-terminal glycine residue.However,the role of this myristoylation in the EV71 life cycle remains largely unknown.To investigate this issue,we developed a myristoylation-deficient virus and reporter(luciferase)pseudovirus with a Gly-to-Ala mutation(G2A)on EV71 VP4.When transfecting the EV71-G2 A genome encoding plasmid in cells,the loss of myristoylation on VP4 did not affect the expression of viral proteins and the virus morphology,however,it did significantly influence viral infectivity.Further,in myristoylation-deficient reporter pseudovirus-infected cells,the luciferase activity and viral genome RNA decreased significantly as compared to that of wild type virus;however,cytopathic effect and viral capsid proteins were not detected in myristoylation-deficient virus-infected cells.Also,although myristoylation-deficient viral RNA and proteins were detected in the second blind passage of infection,they were much fewer in number compared to that of the wild type virus.The replication of genomic RNA and negative-strand viral RNA were both blocked in myristoylation-deficient viruses,suggesting that myristoylation affects viral genome RNA release from capsid to cytoplasm.Besides,loss of myristoylation on VP4 altered the distribution of VP4-green fluorescent protein protein,which disappeared from the membrane structure fraction.Finally,a liposome leakage assay showed that EV71 myristoylation mediates the permeability of the model membrane.Hence,the amino-terminal myristoylation of VP4 is pivotal to EV71 infection and capsidmembrane structure interaction.This study provides novel molecular mechanisms regarding EV71 infection and potential molecular targets for antiviral drug design.展开更多
The effects of free chlorine disinfection of tap water and wastewater effluents on the infectivity, gene integrity and surface antigens of rotaviruses were evaluated by a bench-scale chlorine disinfection experiments....The effects of free chlorine disinfection of tap water and wastewater effluents on the infectivity, gene integrity and surface antigens of rotaviruses were evaluated by a bench-scale chlorine disinfection experiments. Plaque assays, integrated cell culture-quantitative RT- PCR （ICC-RT-qPCR）, RT-qPCR, and enzyme-linked immunosorbent assays （ELISA）, respectively, were used to assess the influence of the disinfectant on virus infectivity as well as genetic and antigenic integrity of simian rotavirus SA11 as a surrogate for human rotaviruses. The ICC-RT-qPCR was able to detect rotaviruses survival from chlorine disinfection at chlorine dose up to 20 mg/L （60 min contact）, which suggested a required chlorine dose of 5 folds （from 1 to 5 mg/L） higher than that indicated by the plaque assay to achieve 1.8 log10 reductions in tap water with 60 rain exposing. The VP7 gene was more resistant than the infectivity and existed at chlorine dose up to 20 mg/L （60 min contact）, while the antigencity was undetectable with chlorine dose more than 5 mg/L （60 min contact）. The water quality also impacted the inactivation efficiencies, and rotaviruses have a relatively higher resistant in secondary effluents than in the tap water under the same chlorine disinfection treatments. This study indicated that rotaviruses have a higher infectivity, gene and antigencity resistance to chlorine than that previously indicated by plaque assay only, which seemed to underestimate the resistance of rotaviruses to chlorine and the risk of rotaviruses in environments. Present results also suggested that re-evaluation of resistance of other waterborne viruses after disinfections by more sensitive infectivity detection method （such as ICC-RT-qPCR） may be necessary, to determine the adequate disinfectant doses required for the inactivation of waterborne viruses.展开更多
Objective To evaluate the infectivity of severe acute respiratory syndrome （SARS） during its incubation period by investigating chains of transmission and individuals isolated for medical observation with a view to ...Objective To evaluate the infectivity of severe acute respiratory syndrome （SARS） during its incubation period by investigating chains of transmission and individuals isolated for medical observation with a view to providing scientific evidence for updating protocols of medical isolation. Methods Individuals related with the two SARS chains of transmission in Beijing in 2003 and a group of individuals isolated for medical observation in Haidian district of Beijing during the SARS outbreak were selected as subjects of study. Contactors with SARS patients and those with symptom development following the contacts were investigated via questionnaire. Serum samples were collected from super transmitters and tested for SARS-CoV antibody by neutralization test and enzyme linked immunosorbent assay （ELISA）. Results A total of 1 112 contactors were investigated in three surveys. Of them, 669 had a history of close contact with symptomatic SARS patients, 101 developed symptoms with a rate of 15.1%, 363 had a history of close contact with patients in their incubation period, none of whom developed symptoms （0%）. Serum samples were collected fi＇om 32 highly-exposed individuals, of whom 13 developing SARS symptoms atter contact had serum samples positive for SARS-CoV antibody. Samples collected from the asymptomatic contactors were all negative for SARS-CoV antibody. Conclusion SARS cases are infectious only during their symptomatic period and are non-infectious during the incubation period. Isolation for medical observation should be placed for individuals who are in close contact with symptomatic SARS patients. The results of our study are of decisive significance for the Ministry of Health to the definition of SARS close contactor.展开更多
Importance of urease activity on pathogenic differences among Brucella species was evaluated. In cell-free extracts, the B. suis urease showed 12 times greater specific activity than the B. melitensis urease. When Fis...Importance of urease activity on pathogenic differences among Brucella species was evaluated. In cell-free extracts, the B. suis urease showed 12 times greater specific activity than the B. melitensis urease. When Fisher-344 rats were inoculated intraperitoneally (IP), at 1 week post-inoculation (PI), B. melitensis wild type 16 M was recovered from spleens and livers in greater numbers than B. suis wild type 1330. At 8 weeks PI, spleens were clear of B. melitensis, whereas B. suis remained. The wild type and the urease deficient strains of B. suis did not differ from each other in terms of recovery from spleen or liver. Our observations suggest that B. melitensis induces greater acute infectivity in Fisher-344 rats, whereas B. suis causes chronic infectivity;and urease activity has no influence on Brucella infection using an IP route.展开更多
This paper considers two differential infectivity（DI） epidemic models with a nonlinear incidence rate and constant or varying population size. The models exhibits two equilibria, namely., a disease-free equilibrium ...This paper considers two differential infectivity（DI） epidemic models with a nonlinear incidence rate and constant or varying population size. The models exhibits two equilibria, namely., a disease-free equilibrium O and a unique endemic equilibrium. If the basic reproductive number σ is below unity,O is globally stable and the disease always dies out. If σ〉1, O is unstable and the sufficient conditions for global stability of endemic equilibrium are derived. Moreover,when σ〈 1 ,the local or global asymptotical stability of endemic equilibrium for DI model with constant population size in n-dimensional or two-dimensional space is obtained.展开更多
The aim of our study was to develop a scoring system to predict whether diarrhea is of a bacterial origin and whether the diarrheal patients constitute a potential source of infection to others. Adults with acute diar...The aim of our study was to develop a scoring system to predict whether diarrhea is of a bacterial origin and whether the diarrheal patients constitute a potential source of infection to others. Adults with acute diarrhea （n=424） were enrolled in the study. Logistic regression and standard regression coefficients were used to formulate the Early Warning Infectivity Score System for Adults with Acute Bacterial Diarrhea （EWIS-ABD）. Four risk factors were identified by logistic regression, including body temperature （P〈0.01）, abdominal pain （P〈0.01）, leukocyte count in stool （P〈0.01）, and unclean dietary history （P〈0.01）. EWIS-ABD was thus developed, in which the value 〉5 points was set as an indicator of bacterial diarrhea. The incidence of bacterial diarrhea increased along with the elevated score. EWIS-ABD was more specific for bacterial diarrhea than for viral diarrhea. The accuracy and reliability of EWIS-ABD was high by prospective validation in 478 patients with acute diarrhea.展开更多
AIM To investigate the effect of a single amino acid mutation in human class B scavenger receptor I(SR-BI) on the infectivity of cell culture-derived hepatitis C virus(HCVcc) in SR-BI knock-down Huh7-si SR-BI cells.ME...AIM To investigate the effect of a single amino acid mutation in human class B scavenger receptor I(SR-BI) on the infectivity of cell culture-derived hepatitis C virus(HCVcc) in SR-BI knock-down Huh7-si SR-BI cells.METHODS Site-directed mutagenesis was used to construct the SR-BI S112 F mutation,and the mutation was confirmed by nucleotide sequencing. SR-BI knock-down Huh7-si SR-BI cells were transfected with SR-BI S112 F,SR-BI wild type(WT) and control plasmids,and then infected with HCVpp(HCV pseudoparticles) and hepatitis C virus derived from cell culture(HCVcc). A fluorescence assay was performed to analyze the effect of the S112 F mutation on HCV entry; quantitative real-time PCR,immunofluorescence,and Western blot assays were used to analyze the effect of the S112 F mutation on HCV infectivity. CHO cells expressing WT and SRBI S112 F were incubated with the HCV E2 protein expressed in HEK 293 T cells,and flow cytometry was performed to examine the ability of SR-BI S112 F to bind to the HCV E2 protein. Huh7-si SR-BI cells were transfected with SR-BI WT and the S112 F mutant,andthen Di I-HDL was added and images captured under the microscope to assess the ability of SR-BI S112 F to take up HDL.RESULTS The SR-BI S112 F mutation was successfully constructed. The S112 F mutation decreased the expression of the SR-BI m RNA and protein. SR-BI S112 F decreased HCV entry and HCVcc infectivity in Huh7-si SR-BI cells. The S112 F mutation impaired the binding of SR-BI to HCV E2 protein and decreased the HDL uptake of SR-BI.CONCLUSION The S112 F single amino acid mutation in SR-BI decreased the levels of the SR-BI m RNA and protein,as well as the ability of SR-BI to bind to the HCV E2 protein. Amino acid 112 in SR-BI plays important roles in HCV entry and the infectivity of HCVcc in vitro.展开更多
Dear Editor The most important property of a virus is its infectivity.To measure infectivity,one can assay viral replication in cells to obtain a titer for a given virus stock.A titer is defined as a given number of i...Dear Editor The most important property of a virus is its infectivity.To measure infectivity,one can assay viral replication in cells to obtain a titer for a given virus stock.A titer is defined as a given number of infectious viral units per unit volume,and an infectious unit is the smallest amount of virus that produces recognizable effects[e.g.,cytopathic effect(CPE),dot blot immunoreactivity].The median tissue culture infectious dose(TCID50)is defined as the dilution of a virus required to infect 50%of a given cell culture.展开更多
DEAR EDITOR,The severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) pandemic remains an important global public health issue. In this study, we unexpectedly found that wildtype Sprague-Dawley(SD) rats can be i...DEAR EDITOR,The severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) pandemic remains an important global public health issue. In this study, we unexpectedly found that wildtype Sprague-Dawley(SD) rats can be infected with the SARS-CoV-2 prototype. Our results showed direct experimental evidence of the infectivity of SARS-CoV-2 infection, subsequent pathogenicity, and protection against reinfection in rats.展开更多
Theileria luwenshuni and Theileria uilenbergi are important tick-borne pathogens and cause substantial losses to the sheep industry in China. The improvement in detection techniques has allowed the identification of m...Theileria luwenshuni and Theileria uilenbergi are important tick-borne pathogens and cause substantial losses to the sheep industry in China. The improvement in detection techniques has allowed the identification of multi-homing parasitism in Theileria parasites. Herein we evaluated the experimental infectivity of T. luwenshuni and T. uilenbergi in Chinese Kunming mice by screening blood samples of experimentally inoculated mice by microscopic examination（ME） and PCR. T. luwenshuni infected Chinese Kunming mice and 20 mice inoculated with this parasite were positive by ME and PCR. In addition, T. uilenbergi infected mice and 20 mice inoculated with this species were positive by ME and PCR. However, the number of red blood cells and the levels of hemoglobin of 40 infected mice had no obvious changes in the course of infection. Our results demonstrated the multi-homing parasitism of T. luwenshuni and T. uilenbergi, which were believed to be parasites of sheep and goats. This study was the first to demonstrate the infection of T. luwenshuni and T. uilenbergi in Kunming mice.展开更多
基金supported by a research grant from the Ministry of Education,Culture,Research and Technology(KEMENDIKBUD RISTEK)number NKB-022/UN2.RST/HKP.05.00/2021 awarded to AB.
文摘Objective:To characterize the infection patterns and growth characteristics of the Zika virus(ZIKV)strain JMB-185 from Indonesia in various mammalian cell lines.Methods:ZIKV was grown in human(A549,HEK293,HepG2,Huh7,Jurkat,and THP-1)and non-human mammalian(RAW264.7,Vero,and Vero76)cell lines.Viral replication kinetics were measured using plaque assay,while intra-and extracellular viral RNA concentrations were assessed using RT-PCR.Flow cytometry was used to quantify the infected cells and cell viability was measured using an MTT assay.The ability of ZIKV to infect cell lines was visualized using a fluorescence immunostaining assay.Results:This ZIKV strain preferentially infected the lung,kidney,and liver cell lines A549,HEK293,Huh7,Vero,and Vero76,but not the immune cells Jurkat,RAW264.7,and THP-1.By contrast,the ZIKV showed no sign of infection in HepG2 cells,while maintaining viral titer over 3 days post-infection,with no infection recorded in immunostaining,no increase in viral RNA,and no indication of cell deterioration.Conclusions:The Indonesian ZIKV strain has a similar infection profile as other strains,except for its poor infectivity on HepG2 cells.Information on the growth characteristics of Indonesia ZIKV will help expand our understanding of the biology of ZIKV which will be useful for various applications including antiviral discovery.
文摘This study explores the antiviral properties of high-voltage low-frequency electric field exposure on the replication of human viruses, including Herpes Simplex Virus type 1 (HSV-1), Human Coronavirus OC43 (HCoV OC43), and Influenza A virus (A H1N1). Using the HealectricsTM device (model S02), which operates by applying high-voltage direct current (30 - 50 kV) with a polarity change frequency of ~0.2 Hz, we investigated the impact on viral infectivity and host cell viability. Virus cultures were exposed to electric fields during different stages: virion adsorption (0 - 1 hour), intracellular replication (1 - 8 hours), and both stages. Viral infectivity was assessed through titration, and cytotoxic effects were evaluated using MTT assays. Electric field exposure significantly reduced viral infectivity, particularly during the combined sorption and replication stages, with up to a 90% decrease in viral activity. Among the viruses tested, HCoV OC43 showed the least sensitivity, with a reduction in viral activity by a factor of 5. Comparisons revealed statistically significant reductions for influenza and herpes viruses, and a trend towards significance for HCoV OC43. The electric field treatment did not significantly affect the viability of Vero and MDCK cells, indicating the method’s safety. Our findings suggest that high-voltage low-frequency electric fields can effectively reduce viral infectivity and may serve as a potential therapeutic and preventive measure against a wide range of membrane-bound viruses, including SARS-CoV-2.
基金supported by grants from the Key Research Program of Frontier Sciences of the Chinese Academy of Sciences (grant No.QYZDJ-SSW-SMC021)the National Natural Science Foundation of China (grants No.31621061 and 31130058)+2 种基金the Virology Key Frontier Science Program of State Key Laboratory of Virology (grant No.klv-2016-03)the National Key R&D Program of China (2017YFD0200400)sponsored by CAS-TWAS President’s Fellowship for International PhD Students
文摘Effective oral infection is set off by interaction of a group of conserved per os infectivity factors(PIFs) with larval midgut columnar epithelial cells. We constructed pseudotyped viruses by substituting pif1, pif2 or pif3 genes of Helicoverpa armigera nucleopolyhedrovirus(Hear NPV) with their homologs from Mamestra bracissae multiple nucleopolyhedrovirus and tested their infectivity to tissue culture cells and to larvae. Transfection and infection assays revealed that all recombinant viruses generated infectious budded virus in both cell culture and in larvae. Electron microscopy showed synthesized occlusion body and occlusion derived virus(ODV) were morphologically indistinguishable from those of the parental virus. By contrast, feeding assays revealed that pseudotyped viruses could not rescue oral infectivity except for pif3 pseudotyped virus that only partially rescued oral infectivity but at a mortality rate much lower than that of the parental Hear NPV. Consistent with the bioassay result, PIF complex was detected in ODVs of pif3 pseudotyped virus only but not in pif1 or pif2 pseudotyped viruses. Our results suggest that PIF complex is essential for oral infectivity, and in the formation of the PIF complex, PIF1, 2 are virus-specific while PIF3 does not appear to be as specific and can function in heterologous environment, albeit to a much more limited extent.
文摘The phasing out of protective measures by governments and public health agencies, despite continued seriousness of the coronavirus pandemic, leaves individuals who are concerned for their health with two basic options over which they have control: 1) minimize risk of infection by being vaccinated and by wearing a face mask when appropriate, and 2) minimize risk of transmission upon infection by self-isolating. For the latter to be effective, it is essential to have an accurate sense of the probability of infectivity as a function of time following the onset of symptoms. Epidemiological considerations suggest that the period of infectivity follows a lognormal distribution. This proposition is tested empirically by construction of the lognormal probability density function and cumulative distribution function based on quantiles of infectivity reported by several independent investigations. A comprehensive examination of a prototypical ideal clinical study, based on general statistical principles (the Principle of Maximum Entropy and the Central Limit Theorem) reveals that the probability of infectivity is a lognormal random variable. Subsequent evolution of new variants may change the parameters of the distribution, which can be updated by the methods in this paper, but the form of the probability function is expected to remain lognormal as this is the most probable distribution consistent with mathematical requirements and available information.
文摘The appearance of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)variant Omicron(B.1.1.529)has caused panic responses around the world because of its high transmission rate and number of mutations.This review summarizes the highly mutated regions,the essential infectivity,transmission,vaccine breakthrough and antibody resistance of the Omicron variant of SARSCoV-2.The Omicron is highly transmissible and is spreading faster than any previous variant,but may cause less severe symptoms than previous variants.The Omicron is able to escape the immune system’s defenses and coronavirus disease 2019 vaccines are less effective against the Omicron variant.Early careful preventive steps including vaccination will always be key for the suppression of the Omicron variant.
基金Project supported by the National Natural Science Foundation of China (No. 30530520)the Zhejiang Agricultural Science and Tech-nology Key Research Projects (No. 2007C12054)the Natural Science Foundation of Zhejiang Province, China (No. Y307397)
文摘Papaya leaf curl China virus (PaLCuCNV) was previously reported as a distinct begomovirus infecting papaya in southern China. Based on molecular diagnostic survey, 13 PaLCuCNV isolates were obtained from tomato plants showing leaf curl symptoms in Henan and Guangxi Provinces of China. Complete nucleotide sequences of 5 representative isolates (AJ558116, AJ558117, AJ704604, FN256260, and FN297834) were determined to be 2738–2751 nucleotides, which share 91.7%–97.9% sequence identities with PaLCuCNV isolate G2 (AJ558123). DNA-β was not found to be associated with PaLCuCNV isolates. To investigate the infectivity of PaLCuCNV, an in-fectious clone of PaLCuCNV-[CN:HeNZM1] was constructed and agro-inoculated into Nicotiana benthamiana, N. tabacum Samsun, N. glutinosa, Solanum lycopersicum and Petunia hybrida plants, which induced severe leaf curling and crinkling symptoms in these plants. Southern blot analysis and polymerase chain reaction (PCR) indicated a systemic infection of test plants by the agro-infectious clone.
文摘Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)is a highly contagious virus that can transmit through respiratory droplets,aerosols,or contacts.Frequent touching of contaminated surfaces in public areas is therefore a potential route of SARS-CoV-2 transmission.The inanimate surfaces have often been described as a source of nosocomial infections.However,summaries on the transmissibility of coronaviruses from contaminated surfaces to induce the coronavirus disease 2019 are rare at present.This review aims to summarize data on the persistence of different coronaviruses on inanimate surfaces.The literature was systematically searched on Medline without language restrictions.All reports with experimental evidence on the duration persistence of coronaviruses on any type of surface were included.Most viruses from the respiratory tract,such as coronaviruses,influenza,SARS-CoV,or rhinovirus,can persist on surfaces for a few days.Persistence time on inanimate surfaces varied from minutes to up to one month,depending on the environmental conditions.SARSCoV-2 can be sustained in air in closed unventilated buses for at least 30 min without losing infectivity.The most common coronaviruses may well survive or persist on surfaces for up to one month.Viruses in respiratory or fecal specimens can maintain infectivity for quite a long time at room temperature.Absorbent materials like cotton are safer than unabsorbent materials for protection from virus infection.The risk of transmission via touching contaminated paper is low.Preventive strategies such as washing hands and wearing masks are critical to the control of coronavirus disease 2019.
基金funded by the National Natural Science Foundation of China(30270898).
文摘A new morphological variant strain of microsporidium was produced by infecting the mulberry looper, Hemerophilaatrilineata [Phthonandria atrilineata], with Nosema bombycis successively for 24 times, and named 24Nbh. Comparativestudies on morphology, infectivity and spore surface protein were conducted. 24Nbh was short and wide, and had asignificant difference (P<0.01) over the Nb spores. The infectivity tests conducted on second instar silkworm larvaeshowed that IC50 of 24Nbh was 1.98104 spores mL-1 and of Nb was 1.72103 spores mL-1, thus indicating that the infectivityof Nb decreased 11.5 times after multiplying in mulberry looper for 24 times. The IC50 of spores from silkworm infected with24 Nbh was 6.9 times less than Nb, showing that the infectivity of 24Nbh spores rejuvenated very fast when reinfected tosilkworms, further more, the length and width of such spore was larger than 24Nbh (P<0.01) and smaller than Nb (P<0.05).The SDS-PAGE profiles of Nb and 24Nbh were generally the same, 4 distinct proteins of 12, 17, 30, 33 kDa were obtainedwith difference in quantity. When 120 g of protein was applied for 2D-PAGE, five suspected different proteins withdifference in quantity were observed. These results demonstrate that these differential proteins maybe associated withvariation in infectivity of the spores.
基金supported by the State Key Basic Research Project(973 project)of China(Grant No.2007CB116308)Planned Projects for Postdoctoral Research Funds of Jiangsu Province,China(Grant No.5910602)Postdoctoral Funds of Jiangsu Academy of Agricultural Sciences,China(Grant No.6510501)
文摘The ultrastructure of porcine kidney(PK)-15 cells was examined after lipofectamine-aided transfection of the molecular clone of the P1 agent.PK-15 cells transfected with the tandem dimer of the P1molecular DNA clone had numbers of intracytoplasmic inclusions,and a few cells had intranuclear inclusions.Intracytoplasmic inclusions were round to oval and 0.1-0.3μm in diameter,and intranuclear inclusions,which were more electron dense,were of two general types:the first were round and small(0.1μm approximately)and the second were hexagonal and larger(0.4-0.8μm in diameter).Cells transfected with the tandem dimer of the P1 molecular DNA clone tested positive for P1 DNA at passage 5.This is the first report that the P1 molecular clone has infectivity in vitro and it will provide fundamental materials for further study of the biological characterization of P1.
基金supported by grants of the National Youth Natural Science Foundation of China,China(82102380)the Youth Natural Science Foundation of Jiangsu Province(BK20200126)the National Youth Natural Science Foundation of China,China(81602400)。
文摘Hepatitis B virus(HBV)genotype B and C are two major genotypes that are prevalent in Asia and differ in natural history and disease progression.The impact of HBV genotypes on viral replication and protein expression has been explored by the transfection of hepatoma cells with replication-competent HBV DNA,which mimics the later stages of the viral life cycle.However,the influence of HBV genotypes on the early events of viral infection remains undetermined,mainly due to the difficulties in obtaining sufficient infectious viral particles for infection assays.Here,we report that a high-titer HBV inoculum can be generated from the transient transfection-based cell model after optimizing transfection conditions and modifying the HBV-expressing construct.By performing in vitro infection assays using transiently transfected derived viruses,we found that clinical genotype C isolates possessed higher infectivity than genotype B isolates.Moreover,we identified a naturally occurring mutation sL21S in small hepatitis B surface protein,which markedly decreased the infectivity of HBV genotype C isolates,but not that of genotype B isolates.In summary,using infectious viral particles provided by the optimized transient transfection-based cell model,we have been able to investigate a wide range of HBV variants on viral infectivity,which may contribute to our understanding of the reasons for different clinical outcomes in HBV infections and the development of therapeutic drugs targeting the early stages of HBV life cycle.
基金supported by the National Natural Science Foundation of China(Grant No.31770184)
文摘The Enterovirus 71(EV71)VP4 is co-translationally linked to myristic acid at its amino-terminal glycine residue.However,the role of this myristoylation in the EV71 life cycle remains largely unknown.To investigate this issue,we developed a myristoylation-deficient virus and reporter(luciferase)pseudovirus with a Gly-to-Ala mutation(G2A)on EV71 VP4.When transfecting the EV71-G2 A genome encoding plasmid in cells,the loss of myristoylation on VP4 did not affect the expression of viral proteins and the virus morphology,however,it did significantly influence viral infectivity.Further,in myristoylation-deficient reporter pseudovirus-infected cells,the luciferase activity and viral genome RNA decreased significantly as compared to that of wild type virus;however,cytopathic effect and viral capsid proteins were not detected in myristoylation-deficient virus-infected cells.Also,although myristoylation-deficient viral RNA and proteins were detected in the second blind passage of infection,they were much fewer in number compared to that of the wild type virus.The replication of genomic RNA and negative-strand viral RNA were both blocked in myristoylation-deficient viruses,suggesting that myristoylation affects viral genome RNA release from capsid to cytoplasm.Besides,loss of myristoylation on VP4 altered the distribution of VP4-green fluorescent protein protein,which disappeared from the membrane structure fraction.Finally,a liposome leakage assay showed that EV71 myristoylation mediates the permeability of the model membrane.Hence,the amino-terminal myristoylation of VP4 is pivotal to EV71 infection and capsidmembrane structure interaction.This study provides novel molecular mechanisms regarding EV71 infection and potential molecular targets for antiviral drug design.
基金supported by the Special Fund of State Key Joint Laboratory of Environment Simulation and Pollution Control of China (No. 10Y04ESPCT)the National Natural Science Fundation of China (No. 51178242)the Major Science and Technology Program for Water Pollution Control and Treatment of China (No. 2008ZX07313-007)
文摘The effects of free chlorine disinfection of tap water and wastewater effluents on the infectivity, gene integrity and surface antigens of rotaviruses were evaluated by a bench-scale chlorine disinfection experiments. Plaque assays, integrated cell culture-quantitative RT- PCR （ICC-RT-qPCR）, RT-qPCR, and enzyme-linked immunosorbent assays （ELISA）, respectively, were used to assess the influence of the disinfectant on virus infectivity as well as genetic and antigenic integrity of simian rotavirus SA11 as a surrogate for human rotaviruses. The ICC-RT-qPCR was able to detect rotaviruses survival from chlorine disinfection at chlorine dose up to 20 mg/L （60 min contact）, which suggested a required chlorine dose of 5 folds （from 1 to 5 mg/L） higher than that indicated by the plaque assay to achieve 1.8 log10 reductions in tap water with 60 rain exposing. The VP7 gene was more resistant than the infectivity and existed at chlorine dose up to 20 mg/L （60 min contact）, while the antigencity was undetectable with chlorine dose more than 5 mg/L （60 min contact）. The water quality also impacted the inactivation efficiencies, and rotaviruses have a relatively higher resistant in secondary effluents than in the tap water under the same chlorine disinfection treatments. This study indicated that rotaviruses have a higher infectivity, gene and antigencity resistance to chlorine than that previously indicated by plaque assay only, which seemed to underestimate the resistance of rotaviruses to chlorine and the risk of rotaviruses in environments. Present results also suggested that re-evaluation of resistance of other waterborne viruses after disinfections by more sensitive infectivity detection method （such as ICC-RT-qPCR） may be necessary, to determine the adequate disinfectant doses required for the inactivation of waterborne viruses.
文摘Objective To evaluate the infectivity of severe acute respiratory syndrome （SARS） during its incubation period by investigating chains of transmission and individuals isolated for medical observation with a view to providing scientific evidence for updating protocols of medical isolation. Methods Individuals related with the two SARS chains of transmission in Beijing in 2003 and a group of individuals isolated for medical observation in Haidian district of Beijing during the SARS outbreak were selected as subjects of study. Contactors with SARS patients and those with symptom development following the contacts were investigated via questionnaire. Serum samples were collected from super transmitters and tested for SARS-CoV antibody by neutralization test and enzyme linked immunosorbent assay （ELISA）. Results A total of 1 112 contactors were investigated in three surveys. Of them, 669 had a history of close contact with symptomatic SARS patients, 101 developed symptoms with a rate of 15.1%, 363 had a history of close contact with patients in their incubation period, none of whom developed symptoms （0%）. Serum samples were collected fi＇om 32 highly-exposed individuals, of whom 13 developing SARS symptoms atter contact had serum samples positive for SARS-CoV antibody. Samples collected from the asymptomatic contactors were all negative for SARS-CoV antibody. Conclusion SARS cases are infectious only during their symptomatic period and are non-infectious during the incubation period. Isolation for medical observation should be placed for individuals who are in close contact with symptomatic SARS patients. The results of our study are of decisive significance for the Ministry of Health to the definition of SARS close contactor.
文摘Importance of urease activity on pathogenic differences among Brucella species was evaluated. In cell-free extracts, the B. suis urease showed 12 times greater specific activity than the B. melitensis urease. When Fisher-344 rats were inoculated intraperitoneally (IP), at 1 week post-inoculation (PI), B. melitensis wild type 16 M was recovered from spleens and livers in greater numbers than B. suis wild type 1330. At 8 weeks PI, spleens were clear of B. melitensis, whereas B. suis remained. The wild type and the urease deficient strains of B. suis did not differ from each other in terms of recovery from spleen or liver. Our observations suggest that B. melitensis induces greater acute infectivity in Fisher-344 rats, whereas B. suis causes chronic infectivity;and urease activity has no influence on Brucella infection using an IP route.
文摘This paper considers two differential infectivity（DI） epidemic models with a nonlinear incidence rate and constant or varying population size. The models exhibits two equilibria, namely., a disease-free equilibrium O and a unique endemic equilibrium. If the basic reproductive number σ is below unity,O is globally stable and the disease always dies out. If σ〉1, O is unstable and the sufficient conditions for global stability of endemic equilibrium are derived. Moreover,when σ〈 1 ,the local or global asymptotical stability of endemic equilibrium for DI model with constant population size in n-dimensional or two-dimensional space is obtained.
基金funded by the National Key Technology R&D Program during the Eleventh 5-year Plan Period[2007BAI24B06]
文摘The aim of our study was to develop a scoring system to predict whether diarrhea is of a bacterial origin and whether the diarrheal patients constitute a potential source of infection to others. Adults with acute diarrhea （n=424） were enrolled in the study. Logistic regression and standard regression coefficients were used to formulate the Early Warning Infectivity Score System for Adults with Acute Bacterial Diarrhea （EWIS-ABD）. Four risk factors were identified by logistic regression, including body temperature （P〈0.01）, abdominal pain （P〈0.01）, leukocyte count in stool （P〈0.01）, and unclean dietary history （P〈0.01）. EWIS-ABD was thus developed, in which the value 〉5 points was set as an indicator of bacterial diarrhea. The incidence of bacterial diarrhea increased along with the elevated score. EWIS-ABD was more specific for bacterial diarrhea than for viral diarrhea. The accuracy and reliability of EWIS-ABD was high by prospective validation in 478 patients with acute diarrhea.
基金Supported by National Natural Science Foundation of China,No.31370196the National 973 Program,No.2013CB531601
文摘AIM To investigate the effect of a single amino acid mutation in human class B scavenger receptor I(SR-BI) on the infectivity of cell culture-derived hepatitis C virus(HCVcc) in SR-BI knock-down Huh7-si SR-BI cells.METHODS Site-directed mutagenesis was used to construct the SR-BI S112 F mutation,and the mutation was confirmed by nucleotide sequencing. SR-BI knock-down Huh7-si SR-BI cells were transfected with SR-BI S112 F,SR-BI wild type(WT) and control plasmids,and then infected with HCVpp(HCV pseudoparticles) and hepatitis C virus derived from cell culture(HCVcc). A fluorescence assay was performed to analyze the effect of the S112 F mutation on HCV entry; quantitative real-time PCR,immunofluorescence,and Western blot assays were used to analyze the effect of the S112 F mutation on HCV infectivity. CHO cells expressing WT and SRBI S112 F were incubated with the HCV E2 protein expressed in HEK 293 T cells,and flow cytometry was performed to examine the ability of SR-BI S112 F to bind to the HCV E2 protein. Huh7-si SR-BI cells were transfected with SR-BI WT and the S112 F mutant,andthen Di I-HDL was added and images captured under the microscope to assess the ability of SR-BI S112 F to take up HDL.RESULTS The SR-BI S112 F mutation was successfully constructed. The S112 F mutation decreased the expression of the SR-BI m RNA and protein. SR-BI S112 F decreased HCV entry and HCVcc infectivity in Huh7-si SR-BI cells. The S112 F mutation impaired the binding of SR-BI to HCV E2 protein and decreased the HDL uptake of SR-BI.CONCLUSION The S112 F single amino acid mutation in SR-BI decreased the levels of the SR-BI m RNA and protein,as well as the ability of SR-BI to bind to the HCV E2 protein. Amino acid 112 in SR-BI plays important roles in HCV entry and the infectivity of HCVcc in vitro.
基金the support of the National Key Research and Development Program of China(2017YFD0201206)the WIV‘‘One-Three-Five’’strategic programs(Y602111SA1)
文摘Dear Editor The most important property of a virus is its infectivity.To measure infectivity,one can assay viral replication in cells to obtain a titer for a given virus stock.A titer is defined as a given number of infectious viral units per unit volume,and an infectious unit is the smallest amount of virus that produces recognizable effects[e.g.,cytopathic effect(CPE),dot blot immunoreactivity].The median tissue culture infectious dose(TCID50)is defined as the dilution of a virus required to infect 50%of a given cell culture.
基金supported by the National Natural Science Foundation of China(U1902215 to Y.G.Y.,32070569 to L.X.)Applied Basic Research Foundation of Yunnan Province(202001AS070023 to D.Y.)+1 种基金Key Project of the CAS“Light of West China”Program(to D.Y.)。
文摘DEAR EDITOR,The severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) pandemic remains an important global public health issue. In this study, we unexpectedly found that wildtype Sprague-Dawley(SD) rats can be infected with the SARS-CoV-2 prototype. Our results showed direct experimental evidence of the infectivity of SARS-CoV-2 infection, subsequent pathogenicity, and protection against reinfection in rats.
基金financially supported by the National Key Research and Development Program of China (2017YFD0501200, 2016YFC1202000, 2016YFC1202002)the earmarked fund for China Agriculture Research System (CARS-37)+5 种基金the National Natural Science Foundation of China (31272556, 31402189, 31372432)the Agricultural Science and Technology Innovation Program, China (2014ZL010)the National Basic Research Program of China (2015CB150300)the Special Funds for Agroscientific Research in the Public Research, China (201303035)the Gansu International Collaboration Special Project, China (1504WKCA056)the Jiangsu Co-Innovation Center Programme for Prevention and Control of Important Animal Infectious Diseases and Zoonoses,State Key Laboratory of Veterinary Etiological Biology Project, China
文摘Theileria luwenshuni and Theileria uilenbergi are important tick-borne pathogens and cause substantial losses to the sheep industry in China. The improvement in detection techniques has allowed the identification of multi-homing parasitism in Theileria parasites. Herein we evaluated the experimental infectivity of T. luwenshuni and T. uilenbergi in Chinese Kunming mice by screening blood samples of experimentally inoculated mice by microscopic examination（ME） and PCR. T. luwenshuni infected Chinese Kunming mice and 20 mice inoculated with this parasite were positive by ME and PCR. In addition, T. uilenbergi infected mice and 20 mice inoculated with this species were positive by ME and PCR. However, the number of red blood cells and the levels of hemoglobin of 40 infected mice had no obvious changes in the course of infection. Our results demonstrated the multi-homing parasitism of T. luwenshuni and T. uilenbergi, which were believed to be parasites of sheep and goats. This study was the first to demonstrate the infection of T. luwenshuni and T. uilenbergi in Kunming mice.
