The effect of pneumonia induced in mice with Mycoplasma pulmonis on resistance to subsequent bacterial infection and the effect of a respiratory infection with Sendai virus on the resistance of mice to Mycoplasma pulmonis.

The effect of pneumonia induced by Mycoplasma pulmonis in mice on the resistance of the lung to additional bacterial infection was examined. The effect of pneumonia induced by Sendai virus on the resistance of mice to M. pulmonis was also investigated and compared with the effect of Sendai virus on resistance to Staphylococcus aureus. Sendai virus infection decreased subsequent resistance to M. pulmonis in proportion to the virus dose. Decreased resistance to subsequent S. aureus and M. pulmonis infection was greatest at about the same time after inoculation of virus and was related to virus-induced lesions. Besides affecting the resistance of mice to subsequent mycoplasma infection, Sendai virus could enhance an existing mycoplasma infection. Pneumonia induced by M. pulmonis did not decrease resistance to subsequent bacterial infection. The mechanism whereby Sendai virus decreases host resistance is therefore similar for bacteria and mycoplasmas, but pneumonia induced by mycoplasmas does not have the same effect.     

Opsonization of HIV with complement enhances infection of dendritic cells and viral transfer to CD4 T cells in a CR3 and DC-SIGN-dependent manner

In the present study, we demonstrated that opsonization of primary HIV-1 with human complement enhances infection of immature monocyte-derived dendritic cells (iDC) and transmission in trans of HIV to autologous CD4(+) T lymphocytes. Infection of iDC by opsonized primary R5- and X4-tropic HIV was increased 3- to 5-fold as compared with infection by the corresponding unopsonized HIV. Enhancement of infection was dependent on CR3 as demonstrated by inhibition induced by blocking Abs. The interaction of HIV with CCR5 and CXCR4 on iDC was affected by opsonization. Indeed, stromal-derived factor-1 was more efficient in inhibiting infection of iDC with opsonized R5-tropic HIV-1(BaL) (45%) than with heat-inactivated complement opsonized virus and similarly RANTES inhibited more efficiently infection of iDC with opsonized X4-tropic HIV-1(NDK) (42%) than with heat-inactivated complement opsonized virus. We also showed that attachment of complement-opsonized virus to DC-specific ICAM-grabbing nonintegrin (DC-SIGN) molecule on iDC and HeLa DC-SIGN(+) CR3(-) cells was 46% and 50% higher compared with heat-inactivated complement opsonized virus, respectively. Hence, Abs to DC-SIGN suppressed up to 80% and 60% the binding of opsonized virus to HeLa cells and iDC, respectively. Furthermore, Abs to DC-SIGN inhibited up to 70% of the infection of iDC and up to 65% of infection in trans of autologous lymphocytes with opsonized virus. These results further demonstrated the role of DC-SIGN in complement opsonized virus uptake and infection. Thus, the virus uses complement to its advantage to facilitate early steps leading to infection following mucosal transmission of HIV.     

异型流感病毒感染小鼠肺细胞因子水平变化

为了制备能够抵御不同型别流感病毒感染的疫苗,揭示机体对异型流感病毒感染交叉免疫保护作用的主要机制,用流感病毒疫苗免疫小鼠后分别感染同型、异型流感病毒,另设使用免疫增强剂IL-2后感染异型流感病毒组,观察小鼠的一般状况和肺指数,并用ELISA方法测定肺匀浆中细胞因子IFN-γ、IL-2、IL-4及IL-10的含量。结果显示,异型免疫组和异型免疫加强组病毒感染后细胞因子IFN-γ含量明显高于感染前（P〈0.05）。研究表明,异型病毒感染后IFN-γ水平明显增高,此细胞因子可能在流感病毒异型间交叉保护免疫反应中起重要作用,其机制有待进一步研究确定。     

仙台病毒RT-PCR检测方法的建立及灰仓鼠中流行情况调查

目的建立仙台病毒（SV）RT-PCR检测方法,并对灰仓鼠仙台病毒感染情况进行调查。方法根据NCBI发表的SV（gi：9627219）基因组序列设计引物,建立RT-PCR方法,对方法的特异性和灵敏性进行验证,并用该方法检测60份灰仓鼠的肺脏样本。结果建立的SV RT-PCR方法显示有较好的敏感性和特异性：以仙台病毒为模板扩增产生197 bp的单一目的条带,经测序比对与NCBI数据库中SV相关序列的一致率为98%,而以猴副流感病毒（SV5）、犬瘟热病毒、小鼠肺炎病毒、呼肠孤病毒III型及腮腺炎病毒为对照无任何条带产生;能检出的SVcDNA最低浓度是96.8 ng/mL;用该方法检测60份灰仓鼠,SV的感染率为3.33%（2/60）。结论建立的SV RT-PCR方法可用于实验类啮齿动物动物SV的常规检测,自然条件下灰仓鼠感染SV的问题不容忽视。     

Immunofluorescence for detection of viral antigen in mice infected with Sendai virus

Sendai virus infection transmitted by contact from cagemates was followed by virus titration and immunofluorescence. The virus grew in the respiratory tract and caused macroscopic lesions in all contact mice. The virus grew to a higher titer in the lung than in the trachea. Tracheal smears, however, were found to be the most suitable for the diagnosis of Sendai virus infection by immunofluorescence, since they contained a large number of cells with intense fluorescence. Diagnosis of Sendai virus infection was made by immunofluorescence within a few hours after autopsy made at early stages of infection.     

Differential effect of prior influenza infection on alveolar macrophage phagocytosis of Staphylococcus aureus and Escherichia coli: involvement of interferon-gamma production

The influenza A virus is one of the main causes of respiratory infection. Although influenza virus infection alone can result in pneumonia, secondary bacterial infection combined with the virus is the major cause of morbidity and mortality. Interestingly, while influenza infection increases susceptibility to some bacteria, including Streptococcus pneumoniae, Staphylococcus aureus (S. aureus), and Haemophilus influenzae, other bacteria such as Escherichia coli (E. coli) and Klebsiella pneumoniae are not associated with influenza infection. The reason for this discrepancy is not known. In this study, it was found that prior influenza virus infection inhibits murine alveolar macrophage phagocytosis of S. aureus but not of E. coli. Here, the mechanism for this inhibition is elucidated: prior influenza virus infection strongly increases interferon gamma (IFN-γ) production. Furthermore, it was shown that IFN-γ differentially affects alveolar macrophage phagocytosis of S. aureus and E. coli. The findings of the present study explain how influenza virus infection increases susceptibility to some bacteria, such as S. aureus, but not others, and provides evidence that IFN-γ might be a promising target for protecting the human population from secondary bacterial infection by influenza.     

DNA distribution and respiratory activity of Spodoptera frugiperda populations infected with wild-type and recombinant Autographa californica nuclear polyhedrosis virus.

Spodoptera frugiperda cells were infected with a wild-type Autographa californica nuclear polyhedrosis virus and with a recombinant Autographa californica nuclear polyhedrosis virus. The recombinant virus was derived from the wild-type virus and produced beta-galactosidase instead of polyhedrin. The changes in cell size, cell growth, viability, DNA distribution, and respiratory activity were followed through the time course of the infection. The DNA content as measured by flow cytometry of infected cells increased to approximately 1.8 times the value of uninfected cells and the distributions of single-cell DNA content of the infected cells were strongly deformed. Early in the infection the respiratory activity passed through a maximum. The mitochondrial activity based on Rhodamine 123 labelling of cells infected with the recombinant virus, as determined by flow cytometry, also passed through a maximum at 24 h post infection while the mitochondrial activity of cells infected with the wild-type virus continued to increase. Evolution of single-cell mitochondrial activity was different in uninfected populations and in populations infected with wild-type and with recombinant virus. In all experiments performed, the recombinant virus influenced cell behavior and the measured parameters earlier than the wild-type virus. The influence of the multiplicity of infection was stronger for the wild-type virus than for the recombinant virus.     

The role of cellular and humoral factors of body resistance in the development of experimental influenza-staphylococcal infections in mice

The clinico-experimental studies of mixed influenza-staphylococcal infection constantly point to the development of the aggravation of the infectious process due to the synergic action of the bacterial and viral infective agents. But, as shown by the authors of the present work, in those cases when the experimental infection with the virus was preceded by staphylococcal infection by 72 hours no synergism was observed. In cases of infection with adaptogenic virus the mortality rate of mice resulting from meningococcal infection was twice as low. The possible explanation of this fact is discussed.     

Immunization with replication-defective mutants of herpes simplex virus type 1: sites of immune intervention in pathogenesis of challenge virus infection.

Replication-defective mutants of herpes simplex virus type 1 (HSV-1) were used as a new means to immunize mice against HSV-1-mediated ocular infection and disease. The effects of the induced immune responses on pathogenesis of acute and latent infection by challenge virus were investigated after corneal inoculation of immunized mice with virulent HSV-1. A single subcutaneous injection of replication-defective mutant virus protected mice against development of encephalitis and keratitis. Replication of the challenge virus at the initial site of infection was lower in mice immunized with attenuated, wild-type parental virus (KOS1.1) or replication-defective mutant virus than in mice immunized with uninfected cell extract or UV-inactivated wild-type virus. Significantly, latent infection in the trigeminal ganglia was reduced in mice given one immunization with replication-defective mutant virus and was completely prevented by two immunizations. Acute replication in the trigeminal ganglia was also prevented in mice immunized twice with wild-type or mutant virus. The level of protection against infection and disease generated by immunization with replication-defective mutant viruses was comparable to that of infectious wild-type virus in all cases. In addition, T-cell proliferative and neutralizing antibody responses following immunization and corneal challenge were of similar strength in mice immunized with replication-defective mutant viruses or with wild-type virus. Thus, protein expression by forms of HSV-1 capable of only partially completing the replication cycle can induce an immune response in mice that efficiently decreases primary replication of virulent challenge virus, interferes with acute and latent infection of the nervous system, and inhibits the development of both keratitis and systemic neurologic disease.     

Replication and persistence of measles virus in defined subpopulations of human leukocytes.

Replication of Edmonston strain measles virus was studied in several human lymphoblast lines, as well as in defined subpopulations of circulating human leukocytes. It was found that measles virus can productively infect T cells, B cells, and monocytes from human blood. These conclusions were derived from infectious center studies on segregated cell populations, as well as from ultrastructural analyses on cells labeled with specific markers. In contrast, mature polymorphonuclear cells failed to synthesize measles virus nucleocapsids even after infection at a relatively high multiplicity of infection. Measles virus replicated more efficiently in lymphocytes stimulated with mitogens than in unstimulated cells. However, both phytohemagglutinin and pokeweed mitogen had a negligible stimulatory effect on viral synthesis in purified populations of monocytes. In all instances the efficiency of measles virus replication by monocytes was appreciably less than that of mitogenically stimulated lymphocytes or of continuously culture lymphoblasts. Under standard conditions of infection, all of the surveyed lymphoblast lines produced equivalent amounts of measles virus regardless of the major histocompatibility (HL-A) haplotype. Hence, no evidence was found that the HL-A3,7 haplotype conferred either an advantage or disadvantage with respect to measles virus synthesis in an immunologically neutral environment. A persistent infection with measles virus could be established in both T and B lymphoblasts. The release of infectious virus from such persistently infected cells was stable over a period of several weeks and was approximately 100-fold less than peak viral titers obtained in each respective line after acute infection.     

Human lung carcinoma (A-549) continuing cell line and human endothelial (ECV-304) continuing cell line responses to the influenza virus at different multiplicities of infection

The course of infection upon virus entry into the cell depends not only on the biological characteristics of the cells and of the virus itself, but also on the intensity of the cell infection by the virus, i.e., on the multiplicity of infection. The purpose of our work was to perform a comparative study of the responses of two human cell lines, the lung carcinoma cell line A-549 and the endothelium cell line ECV-304, to the infection with the influenza virus A at different multiplicities of infection. At the first passage, both cell lines responded by enhancement of proliferation and apoptosis induction only to the low doses of influenza virus (ID 1–10). In A-49 cells, the stimulatory effect of the low virus doses was observed 1–2 days earlier than in ECV-304 cells. Enhanced proliferation was observed in both cell lines from the second to the fourth passages, when cells were infected with higher virus doses (ID 100 and 1000). In addition, the response of the A-549 cells to low doses of the H3N2 strain of the influenza virus A depended on the virus propagation conditions—namely, no enhancement of cell proliferation was observed in response to the infection with the virus propagating in chicken embryonated eggs, in contrast to infection with the virus that propagated in cell culture. Immunocytochemistry of A-549 cells has demonstrated that, on the third day after infection, there could be observed a change (in the dose-dependent manner) in the intracellular localization of p53 and cyclin A, proteins involved in the cell cycle progression. At the low virus dose, cyclin A was predominantly detected in the nuclei (63%), while at the high virus dose it was p53 (54%), which was predominantly detected in this cellular compartment, this observation confirming that stimulation of cell proliferation in the case of very low multiplicity of infection and cell division arrest takes place in the case of high multiplicity of influenza virus infection. The study of the influenza virus A reproduction in A-549 and ECV-304 cells using a whole number of virology techniques showed low sensitivity of these cells to the influenza virus, which manifested in the gradual decrease in the viral RNA expression and the impairment of mature viral particles assembly during several passages. Therefore, the decrease in the multiplicity of infection is associated in the A-549 and ECV-304 cells with impairment of production of mature virus particles or certain virus protein synthesis, which is accompanied by cell proliferation enhancement and apoptosis induction. As a result of the comparative study of the two cell lines (A-549 and ECV-304) upon infection with different doses of influenza virus A, we have revealed common principles and specific features indicating the effects of the biological properties of the viruses and cells, as well as of the multiplicity of infection on the course of virus infection.     

Salivary Excretion of Coxsackie B-1 Virus in Rabbits

Coxsackie B-1 virus was injected into the ear vein of albino doe rabbits. Saliva and blood samples were taken before the injection of virus and at specific times thereafter. Virus was recovered in the whole saliva when the blood titer was approximately 10(4) TCID(50) per 0.1 ml or greater. The virus could be detected in the saliva as early as 2 min after the initiation of the viremia. The recovered virus was shown to be the same as the injected virus by serological identification of the recovered virus with neutralizing antibody for Coxsackie B-1 virus. These results suggest that virus may be transmitted to other animals in the saliva of animals who are in the viremic phase of infection without infection of the oropharyngeal tissues.     

PCT和CRP在感染性疾病诊断中的临床意义

目的:探讨PCT、CRP在感染性疾病诊断中的价值,评价两者与感染类型的相关性。方法:回顾性统计分析420例发热待查的患者血清PCT,血浆CRP浓度与感染的关系,采用胶体金法检测血清中PCT的浓度,免疫荧光比色法检测血浆中CRP浓度,血培养及血清学的方法检测感染是否存在及感染类型。结果:通过血培养或血清学鉴定出132例存在细菌感染(细菌组),病毒感染103例(病毒组),细菌病毒混合感染76例(混合感染组),109例阴性(非感染组)。PCT在细菌组水平最高(20.6±6.7)μg/L,非感染组水平最低(8.7±2.3)μg/L,四组间差异有统计学意义(F=34.69,P0.05)。CRP在混合组中水平最高(18.1±3.7)mg/L,在非感染组水平最低(5.8±1.7)mg/L,四组间差异有统计学意义(F=23.55,P0.05)。PCT对细菌感染检测的敏感度为92.7%,特异度为63.2%。PCT对病毒感染检测的敏感度为78.7%,特异性为46.0%。细菌感染组、病毒感染组和混合感染组血清CRP和PCT均呈正相关(P=0.000.05)。结论:细菌感染的PCT、CRP血清浓度明显高于病毒感染,两者对细菌感染患者的特异性均明显高于病毒感染,细菌感染PCT血清浓度较CRP变化敏感。     

侵染天南星科植物病毒的分子鉴定及其生态学研究

通过病毒粒子部分提纯和形态学观察,发现侵染我国南方天南星科植物的病毒主要有线状和球状两种形态．经病毒基因组序列分析确定线状病毒为芋花叶病毒(DsMV);经血清学反应和序列分析确定球状病毒为黄瓜花叶病毒(CMV)．CMV CP基因序列同源性分析的结果表明,侵染天南星科的CMV是相对独立的种内变异类型,归属于亚组L同时,CMV存在对天南星科植物的适应性变异．对采自我国海南、湖南、浙江、上海等地的126个天南星科植物样品进行RNA核酸斑点杂交检测,获得病毒检测结果。海南省样品DsMV的检出率为73．3％,CMV的检出率为46．7％;湖南省样品DsMV的检出率为100％,CMV的检出率为38.5％;浙江省样品DsMV的检出率为93．0％,CMV的检出率为7．0％;上海市样品DsMV的检出率为100％,尚没有检测到CMV,首次证实了自然条件下CMV作为天南星科植物主要病毒的存在,在我国南方地区,该病毒对天南星科植物的自然侵染受到气候、季节和寄主等生态因子的影响。DsMV则在天南星科植物上普遍存在。     

Roles of neuraminidase in the initial stage of influenza virus infection

We propose a concept that neuraminidase (NA) promotes virus entry into target cells during the initial stage of viral infection, in addition to the generally accepted concept that influenza virus NA promotes the release of progeny virus from a host cell at the final stage of viral replication. When NA activity was inhibited with specific inhibitors such as zanamivir and oseltamivir carboxylate, infection efficiency of the virus to MDCK and A549 cells was reduced to approximately 1/4 and 1/8, respectively. NA inhibitors did not significantly affect virus binding and envelope fusion activities, when assessed using an erythrocyte and virus system. Since the initial stage of viral infection involves binding of the virus to the target cell, virus entry into an endosome and envelope fusion with the endosomal membrane, our results indicated that NA inhibitors interfered with the virus entry step. Thus, NA is thought to promote virus entry, and thereby enhances infection efficiency.     

Repeated passage of wild-type woodchuck hepatitis virus in lymphoid cells does not generate cell type-specific variants or alter virus infectivity

Woodchuck hepatitis virus (WHV), which is closely related to human hepatitis B virus, infects the liver but also invariably establishes persistent infection in the lymphatic system. Although the dose of invading virus appears to be the main factor in determining whether WHV infection is restricted to the lymphatic system or also engages the liver, the nature of WHV lymphotropism remains unclear and a role for a specific lymphotropic variant was not excluded. The availability of woodchuck lymphocyte and hepatocyte cultures susceptible to WHV infection allows investigation of this issue in vitro. We hypothesized that repeated passage of wild-type WHV in lymphoid cells should lead to enrichment of a lymphotropic virus variant, if in fact such a variant exists. For this purpose, wild-type WHV with a homogeneous sequence was used as the inoculum, while lymphoid cells from a single healthy woodchuck donor and a normal woodchuck WCM-260 hepatocyte line served as infection targets. The serial passage of the wild-type virus repeated up to 13 times for both cell types did not lead to the emergence of cell type-specific WHV variants, as revealed by sequence analysis of the virus envelope and the core and X gene sequences. Moreover, the virus passaged in both cell types remained infectious for naive woodchucks, produced infection profiles that depended upon virus dose but not on virus cellular origin, and retained its initial DNA sequence. These results imply that WHV lymphotropism is a natural propensity of the wild-type virus and is not a consequence of infection with a viral variant.     

Immunization of mice against West Nile virus with recombinant envelope protein.

West Nile (WN) virus is a mosquito-borne flavivirus that emerged in the United States in 1999 and can cause fatal encephalitis. Envelope (E) protein cDNA from a WN virus isolate recovered from Culex pipiens in Connecticut was expressed in Escherichia coli. The recombinant E protein was purified and used as Ag in immunoblot assays and immunization experiments. Patients with WN virus infection had Abs that recognized the recombinant E protein. C3H/HeN mice immunized with E protein developed E protein Abs and were protected from infection with WN virus. Passive administration of E protein antisera was also sufficient to afford immunity. E protein is a candidate vaccine to prevent WN virus infection.     

Appearanc of cytotoxic cells within the bronchus after local infection with herpes simplex virus.

Cells from rabbit spleens, bronchial washings (BW) and bronchus-associated lymphoid tissues (BALT) were examined for their ability to lyse cells infected with herpes simplex virus (HSV). Specific lysis of HSV-infected cells was mediated by BW cells as early as 4 days after intratracheal infection of the rabbits with the virus whereas lysis by spleen cells and BALT cells was not detected until 7 or more days after infection. Lysis by spleen cells was initially detected 7 days after intraperitoneal injection of the virus but lysis by BW and BALT cells was not observed until 14 days after infection. Although spleen, BW, and BALT cells could lyse antibody-coated target cells, antibodies detectable by antibody-dependent cellular cytotoxicity could not be detected in bronchial washings until 7 or more days after infection. The data suggest that cells capable of direct cytotoxicity of virus-infected cells appear within the bronchus after local infection by the virus.     

Binding of measles virus to membrane cofactor protein (CD46): importance of disulfide bonds and N-glycans for the receptor function.

Two cellular proteins, membrane cofactor protein (MCP) and moesin, were reported recently to be functionally associated with the initiation of a measles virus infection. We have analyzed the interaction of measles virus with cell surface proteins, using an overlay binding assay with cellular proteins immobilized on nitrocellulose. Among surface-biotinylated proteins from a human rectal tumor cell line (HRT), measles virus was able to bind only to a 67-kDa protein that was identified as MCP. The virus recognized different isoforms of MCP expressed from human (HRT and HeLa) and simian (Vero) cell lines. The binding of measles virus to MCP was abolished after cleavage of the disulfide bonds by reducing agents as well as after enzymatic release of N-linked oligosaccharides. By contrast, removal of sialic acid or O-linked oligosaccharides did not affect the recognition of MCP measles virus. These data indicate that the receptor determinant of MCP is dependent on a conformation of the protein that is maintained by disulfide bonds and N-glycans present in the complement binding domains. Our results are consistent with a role of MCP as primary attachment site for measles virus in the initial stage of an infection. The functional relationship between MCP and moesin in a measles virus infection is discussed.