Respiratory viruses augment the adhesion of bacterial pathogens to respiratory epithelium in a viral species- and cell type-dependent manner

Secondary bacterial infections often complicate respiratory viral infections, but the mechanisms whereby viruses predispose to bacterial disease are not completely understood. We determined the effects of infection with respiratory syncytial virus (RSV), human parainfluenza virus 3 (HPIV-3), and influenza virus on the abilities of nontypeable Haemophilus influenzae and Streptococcus pneumoniae to adhere to respiratory epithelial cells and how these viruses alter the expression of known receptors for these bacteria. All viruses enhanced bacterial adhesion to primary and immortalized cell lines. RSV and HPIV-3 infection increased the expression of several known receptors for pathogenic bacteria by primary bronchial epithelial cells and A549 cells but not by primary small airway epithelial cells. Influenza virus infection did not alter receptor expression. Paramyxoviruses augmented bacterial adherence to primary bronchial epithelial cells and immortalized cell lines by up-regulating eukaryotic cell receptors for these pathogens, whereas this mechanism was less significant in primary small airway epithelial cells and in influenza virus infections. Respiratory viruses promote bacterial adhesion to respiratory epithelial cells, a process that may increase bacterial colonization and contribute to disease. These studies highlight the distinct responses of different cell types to viral infection and the need to consider this variation when interpreting studies of the interactions between respiratory cells and viral pathogens.     

Quantitative proteome profiling of respiratory virus-infected lung epithelial cells

Respiratory virus infections are among the primary causes of morbidity and mortality in humans. Influenza virus, respiratory syncytial virus (RSV), parainfluenza (PIV) and human metapneumovirus (hMPV) are major causes of respiratory illness in humans. Especially young children and the elderly are susceptible to infections with these viruses. In this study we aim to gain detailed insight into the molecular pathogenesis of respiratory virus infections by studying the protein expression profiles of infected lung epithelial cells.A549 cells were exposed to a set of respiratory viruses [RSV, hMPV, PIV and Measles virus (MV)] using both live and UV-inactivated virus preparations. Cells were harvested at different time points after infection and processed for proteomics analysis by 2-dimensional difference gel electrophoresis. Samples derived from infected cells were compared to mock-infected cells to identify proteins that are differentially expressed due to infection.We show that RSV, hMPV, PIV3, and MV induced similar core host responses and that mainly proteins involved in defense against ER stress and apoptosis were affected which points towards an induction of apoptosis upon infection. By 2-D DIGE analyses we have gathered information on the induction of apoptosis by respiratory viruses in A549 cells.     

280例呼吸道感染儿童呼吸道病毒病原学检出情况及流行规律分析

目的:了解呼吸道感染儿童呼吸道病毒病原学检出情况及其流行规律,为儿童呼吸道感染的预防、诊断及治疗提供病原学依据。方法:选取2016年1月-2017年12月期间中国人民解放军中部战区总医院收治的280例呼吸道感染患儿为研究对象,分析患儿呼吸道分泌物中呼吸道病毒的检出情况,并分析呼吸道感染儿童呼吸道病毒感染与年龄、季节、疾病类型的关系。结果:280例呼吸道感染患儿中共检出98份阳性标本,阳性率为35.00%,其中有2份标本中检出2种病毒感染,混合感染阳性率为0.71%;在所有病毒类型中,呼吸道合胞病毒(RSV)病毒感染阳性率最高。1岁患儿的病毒感染阳性率最高,与其他年龄段病毒感染阳性率比较差异有统计学意义(P0.05)。呼吸道感染患儿春季、冬季的病毒感染阳性率明显高于夏季、秋季(P0.05)。不同呼吸道感染疾病类型患儿病毒感染阳性率比较差异有统计学意义(P0.05),以喘息性肺炎、毛细支气管炎、肺炎患儿病毒感染阳性率较高。结论:RSV是呼吸道感染儿童呼吸道病毒感染的主要致病病原体,1岁的婴幼儿较易感染,春季、冬季为其高发季节,且以肺炎、毛细支气管炎、喘息性肺炎患儿的病毒感染阳性率较高。     

Acute effects of parainfluenza virus on epithelial electrolyte transport

Parainfluenza viruses are important causes of respiratory disease in both children and adults. In particular, they are the major cause of the serious childhood illness croup (laryngotracheobronchitis). The infections produced by parainfluenza viruses are associated with the accumulation of ions and fluid in the respiratory tract. It is not known, however, whether this accumulation is because of a direct effect of the viruses on ion and fluid transport by the respiratory epithelium. Here we show that a model parainfluenza virus (the Sendai virus), in concentrations observed during respiratory infections, activates Cl- secretion and inhibits Na+ absorption across the tracheal epithelium. It does so by binding to a neuraminidase-insensitive glycolipid, possibly asialo-GM1, triggering the release of ATP, which then acts in an autocrine fashion on apical P2Y receptors to produce the observed changes in ion transport. These findings indicate that fluid accumulation in the respiratory tract associated with parainfluenza virus infection is attributable, at least in part, to direct effects of the virus on ion transport by the respiratory epithelium.     

Observations on Clinical and Immunofluorescent Diagnosis of Parainfluenza Virus Infections

Immunofluorescent techniques have been applied to nasopharyngeal secretions for the rapid diagnosis of parainfluenza virus types 1, 2, and 3 infections. Seventy-five infections were found by isolation techniques; 55 of these had nasopharyngeal secretions taken and 53 were positive by direct examination. A comparison of the results of 60 neutralization tests with immunofluorescence applied to monkey kidney isolations showed complete agreement. Immunofluorescence appeared to be a satisfactory method for differentiating the various haemadsorption viruses. The importance of parainfluenza viruses and respiratory syncytial virus in croup was noted and the association of the parainfluenza viruses with acute respiratory virus infection was confirmed. The clinical relationship between respiratory syncytial virus and parainfluenza virus type 3 is discussed.     

Reliability of the serological method of examination as a basis for diagnosis of associated respiratory virus infections.

The reliability of the results of serological examination in diagnostics of associated infections was studied on a model of artificially provoked vaccinal infections in humans and in laboratory animals. The effect of administered monopreparations on changes in the level of both homologous and heterologous antibodies was tested. The character of immunological changes following simultaneous administration of two or more respiratory viruses was analysed and the effect of these viruses in diseases of divers etiology was studied. According to the results of experiments on laboratory animals, repeated administration of any of the earlier used respiratory viruses stimulated the accumulation of only homologous antibodies while heterologous antibodies did not increase at all. The results revealed the possibility of simultaneous immunological reorganization of a child's organism in response to the influence of several different antigens from the group of respiratory viruses acting synchronously or in succession. Results of the analysis demonstrated the reliability of the employed serological methods of diagnosis of respiratory virus infections.     

季节性流感病毒与其他病原混合或继发感染的研究进展与思考

流感病毒包括甲(A)、乙(B)、丙(C)和丁(D)四种型。人流行性感冒是由甲型和乙型季节性流感病毒引起的一种急性呼吸道传染病。流感病毒感染患者主要表现出呼吸道症状,严重时可以导致肺炎。此外,与其他病毒、细菌和支原体等病原体混合或继发感染时,会增加流感患者的重症率和死亡率。近几年,流感病毒与其他病原协同感染的病例有增加趋势。本文归纳总结了流感病毒与其他病原混合及继发感染的研究现状,希望为流感病毒复杂感染情况的临床诊断和治疗方案的制定提供线索。     

569例呼吸道感染住院儿童7种常见呼吸道病毒病原学分析

目的了解呼吸道感染住院患儿呼吸道病毒的分布情况。方法选取2015年7月至2016年6月呼吸道感染的住院患儿病例,抽取鼻咽分泌物,采用直接免疫荧光法检测常见的7种病毒,即呼吸道合胞病毒(RSV)、甲型流感病毒(IVA)、乙型流感病毒(IVB)、副流感病毒1型(PIV1)、副流感病毒2型(PIV2)、副流感病毒3型(PIV3)、腺病毒(IVD)。结果共有569例样本送检,193例病毒检测阳性(33.92%),其中有3例为2种病毒的混合感染。男性共369例,女性共200例。其中RSV、PIV3、ADV的感染率居前三位。呼吸道病毒的感染率随着患儿年龄的增长逐渐下降,除新生儿外,6个月内的婴幼儿呼吸道病毒检出率最高。RSV检测阳性率自11月开始呈增高趋势,12月最高,达55.86%。PIV3的检出高峰出现在7月,达13.64%。结论 RSV是本地区儿童呼吸道感染最常见的病毒。呼吸道病毒的检出率与年龄、季节等因素有关。     

Live bivalent vaccine for parainfluenza and influenza virus infections

Influenza and human parainfluenza virus infections are of both medical and economical importance. Currently, inactivated vaccines provide suboptimal protection against influenza, and vaccines for human parainfluenza virus infection are not available, underscoring the need for new vaccines against these respiratory diseases. Furthermore, to reduce the burden of vaccination, the development of multivalent vaccines is highly desirable. Thus, to devise a single vaccine that would elicit immune responses against both influenza and parainfluenza viruses, we used reverse genetics to generate an influenza A virus that possesses the coding region for the hemagglutinin/neuraminidase ectodomain of parainfluenza virus instead of the influenza virus neuraminidase. The recombinant virus grew efficiently in eggs but was attenuated in mice. When intranasally immunized with the recombinant vaccine, all mice developed antibodies against both influenza and parainfluenza viruses and survived an otherwise lethal challenge with either of these viruses. This live bivalent vaccine has obvious advantages over combination vaccines, and its method of generation could, in principle, be applied in the development of a "cocktail" vaccine with efficacy against several different infectious diseases.     

Pre-existing heterosubtypic immunity provides a barrier to airborne transmission of influenza viruses

Human-to-human transmission of influenza viruses is a serious public health threat, yet the precise role of immunity from previous infections on the susceptibility to airborne infection is still unknown. Using the ferret model, we examined the roles of exposure duration and heterosubtypic immunity on influenza transmission. We demonstrate that a 48 hour exposure is sufficient for efficient transmission of H1N1 and H3N2 viruses. To test pre-existing immunity, a gap of 8–12 weeks between primary and secondary infections was imposed to reduce innate responses and ensure robust infection of donor animals with heterosubtypic viruses. We found that pre-existing H3N2 immunity did not significantly block transmission of the 2009 H1N1pandemic (H1N1pdm09) virus to immune animals. Surprisingly, airborne transmission of seasonal H3N2 influenza strains was abrogated in recipient animals with H1N1pdm09 pre-existing immunity. This protection from natural infection with H3N2 virus was independent of neutralizing antibodies. Pre-existing immunity with influenza B virus did not block H3N2 virus transmission, indicating that the protection was likely driven by the adaptive immune response. We demonstrate that pre-existing immunity can impact susceptibility to heterologous influenza virus strains, and implicate a novel correlate of protection that can limit the spread of respiratory pathogens through the air.     

The 22,000-kilodalton protein of respiratory syncytial virus is a major target for Kd-restricted cytotoxic T lymphocytes from mice primed by infection.

Recombinant vaccinia viruses containing the 22-kilodalton protein (matrixlike or 22K protein) or phosphoprotein gene from respiratory syncytial virus were constructed. These recombinant viruses expressed proteins which were immunoprecipitated by appropriate respiratory syncytial virus antibodies and comigrated with authentic proteins produced by respiratory syncytial virus infection. The new recombinant viruses (and others previously described containing the attachment glycoprotein, fusion, or nucleoprotein genes of respiratory syncytial virus) were used to infect target cells for cultured polyclonal cytotoxic T lymphocytes generated from the spleens of BALB/c or DBA/2 mice primed by intranasal infection with respiratory syncytial virus. Respiratory syncytial virus-specific cytotoxic T lymphocytes (CTL) showed strong Kd (but not Dd)-restricted recognition of the 22K protein. As previously reported, the fusion protein and nucleoprotein were both seen by CTL, but recognition of these proteins was comparatively weak. There was no detectable recognition of other respiratory syncytial virus proteins tested (including phosphoprotein). 22K protein-specific splenic memory CTL persisted for at least 11 months after infection of BALB/c mice. Priming BALB/c mice with recombinant vaccinia virus containing the 22K protein gene induced respiratory syncytial virus-specific memory CTL at lower levels than that previously reported following infection with a similar recombinant containing the fusion protein gene. These data identify the 22K protein as a major target antigen for respiratory syncytial virus-specific CTL from H-2d mice primed by respiratory syncytial virus infection.     

Selective infection of lower respiratory tract by respiratory viruses in children with recurrent respiratory tract infections.

Thirty preschool children presenting with recurrent respiratory infections and their unaffected siblings were observed prospectively for a year. The index children experienced more episodes of acute respiratory infection than their siblings. Respiratory viruses were the major cause of respiratory infections. The index children had lower respiratory tract disease, predominantly wheeze, during 34% of proved respiratory virus infections compared with 11% of such infections experienced by the control children (p less than 0.02). Atopic children had an increased tendency to wheeze that did not reach significance, but atopy was not associated with increased susceptibility to respiratory infections.     

Dynamics of human respiratory virus-specific CD8+ T cell responses in blood and airways during episodes of common cold

We determined the dynamics of CD8(+) T cells specific for influenza virus and respiratory syncytial virus in blood and tracheostoma aspirates of children during the course of respiratory infections. We showed that during localized respiratory infections the ratio of activated effector CD8(+) T cells to resting memory/naive CD8(+) T cells in peripheral blood increased significantly. Furthermore, the number of effector/memory T cells specific for respiratory viruses declined in blood and increased in the airways, suggesting that these T cells redistributed from blood to airways. T cells specific for the infecting virus were present in the airways for longer periods at increased levels than nonspecifically recruited bystander T cells. After clearance of the infection, the ratio of resting memory and naive CD8(+) T cells normalized in peripheral blood and also memory T cell numbers specific for unrelated viruses that declined during the infection due to bystander recruitment were restored. Taken together, these results showed a significant systemic T cell response during relatively mild secondary infections and extensive dynamics of virus-specific and nonspecific Ag-experienced T cells.     

Coxsackie virus infection in association with heart affection in man

The frequency of actual infections by selected types of coxsackie virus (B1-B6, A7, A9) has been followed up in adult patients hospitalized for reason of heart disease. Criteria of actual coxsackie virus infection have been defined, including a significant rise of titer of virus neutralizing antibodies and/or presence of virus-specific IgM antibodies. An actual coxsackie virus infection has been found in 71 (46.3%) out of 153 patients. Most frequently coxsackie B1 virus infection (24 times) and B4 (17 times) could be proved. In sera of patients with actual coxsackie virus infection, a higher proportion of infection with some types of coxsackie viruses, along with the presence of antibodies against a larger number of coxsackie virus types has been found, as compared to sera from patients without actual coxsackie virus infection, and/or, in sera of healthy blood donors (control group). The role of repeated infections with different types of coxsackie virus in the etiology of heart disease has been discussed.     

Vaccine-induced immunopathology during bovine respiratory syncytial virus infection: exploring the parameters of pathogenesis

The bovine and human respiratory syncytial viruses cause severe lower respiratory tract infections. Effective vaccines against the respiratory syncytial viruses have been lacking since vaccine failures in the 1960s and 1970s. In this report, we describe a bovine respiratory syncytial virus (bRSV) challenge model in which both classical bRSV respiratory infection and vaccine-enhanced immune pathology were reproduced. The classical, formalin-inactivated (FI) bRSV vaccine that has been associated with vaccine failure was efficient in inducing high antibody titers and reducing viral loads but also primed calves for a far more serious enhanced respiratory disease after a bRSV challenge, thereby mimicking the enhanced clinical situation in FI human RSV (hRSV)-immunized and hRSV-infected infants in the 1960s. We show that immunization with FI-bRSV mainly primes a Th2-like inflammatory response that is characterized by a significant eosinophilic influx in the bronchial alveolar lung fluid and lung tissues and high levels of immunoglobulin E serum antibodies. The current model may be useful in the evaluation of new bRSV candidate vaccines for potency and safety.     

Studies on the epidemiology of child infections. 3. Parainfluenza viruses (types 1-4) and respiratory syncytial virus infections.

A seroepidemiological study on the rate of neutralizing antibodies for parainfluenza viruses (types 1-4) and for the respiratory syncytial virus in 2,514 infants and children between 0 and 15 years, residing in Bari and its hinterland was carried out. Positive results were very high for both the individual parainfluenza serotypes (81.7% for type 3, 78.0% for type 1, 76.6% for type 4, 71.9% for type 2) and for respiratory syncytial virus (88.5%). The pattern of infections due to respiratory syncytial virus differed from that of the parainfluenza viruses not only for the higher serologic positive rate, but also for the larger number of elevated titre responses in each age group.     

Reactivity of serum samples from patients with a flavivirus infection measured by immunofluorescence assay and ELISA

Flavivirus infections are a significant public health problem, since several members of the Flaviviridae family are highly pathogenic to humans. Accurate diagnosis and differentiation of the infecting virus is important, especially in areas where many flaviviruses are circulating. In this study we evaluated a newly developed commercially available immunofluorescence assay (IFA) (INDX, Baltimore, MD, USA) for the detection of IgM and IgG antibodies against dengue virus, yellow fever virus, Japanese encephalitis virus and West Nile virus. IFA was compared with standard diagnostic enzyme immunoassays (EIAs) specific for the detection of IgM and IgG antibodies against these viruses. Forty-seven serum samples from patients with a defined flavivirus infection were tested. As controls, serum samples from individuals with antibodies against tick-borne encephalitis virus and hepatitis C virus as well as healthy individuals were included. The results obtained from this study indicate that IFA showed a significantly better discrimination for flavivirus specific IgM antibodies than did the standard IgM specific EIAs (the overall cross-reactivity varied between 4 and 10% by IFA and 30-44% by EIA for the respective viruses). In contrast, the detection of flavivirus specific IgG antibodies showed high cross-reactions in both IFA and EIAs (overall cross-reactivity 16-71 and 62-84%, respectively). This study clearly stated the complexity of flavivirus diagnosis, showing that one cannot rely on one assay or search for one virus only. The flavivirus IFA is a useful tool for the identification of flavivirus infections during the acute stage of disease. In particular, IFA can be an important diagnostic tool for testing samples from travellers who have been accidentally exposed to these viruses.     

Limiting Respiratory Viral Infection by Targeting Antiviral and Immunological Functions of BST‐2/Tetherin: Knowledge and Gaps

Recent findings regarding the cellular biology and immunology of BST‐2 (also known as tetherin) indicate that its function could be exploited as a universal replication inhibitor of enveloped respiratory viruses (e.g., influenza, respiratory syncytial virus, etc.). BST‐2 inhibits viral replication by preventing virus budding from the plasma membrane and by inducing an antiviral state in cells adjacent to infection via unique inflammatory signaling mechanisms. This review presents the first comprehensive summary of what is currently known about BST‐2 anti‐viral function against respiratory viruses, how these viruses construct countermeasures to antagonize BST‐2, and how BST‐2 function might be targeted to develop therapies to treat respiratory virus infections. The authors address the current gaps in knowledge, including the need for mechanistic understanding of BST‐2 antagonism by respiratory viruses, that should be bridged to achieve that goal.

     

Persistent or Slow Viral Infections and Related Diseases

The discovery of persistent transmissible agents by veterinarians has led to striking advances in the infectious cause of neuropathies of human beings. There is evidence for persisting infection in congenital rubella and the herpes group of viruses including cytomegalovirus infections. Hepatitis types A and B are candidates for inclusion in the category of persisting viral infections.The rubeola or measles virus is established as a persistent virus which causes elevated antibodies in the serum and cerebrospinal fluid of many patients with severe demyelinating disease such as subacute sclerosing panencephalitis and multiple sclerosis. Elevated antibodies against vaccinia virus have been found in the cerebrospinal fluid of some patients with multiple sclerosis and neuromyelitis optica, a rare form of multiple sclerosis.