Detection of multiple viral and bacterial infections in acute exacerbation of chronic obstructive pulmonary disease: A pilot prospective study

Few studies have evaluated the contribution of multiple virus and bacterial infections in acute exacerbation of chronic obstructive pulmonary disease. This study estimated the burden of multiple viral and bacterial respiratory infections in moderate to very severe chronic obstructive pulmonary disease patients that were prospectively followed‐up during a 12‐month pilot study. Clinical data were collected monthly and sputum was collected at the time of each acute exacerbation event. Classical culture techniques for bacteria and multiplex polymerase chain reaction (PCR) and microarray detection assays were performed to identify viral and atypical bacterial pathogens in the sputum. Overall, 51 patients were included and 45 acute exacerbation events were investigated clinically and microbiologically. Among the 45 acute exacerbation events, 44% had evidence of viral infection involving human rhinovirus (HRV) and metapneumovirus (hMPV) in 20% and 18%, respectively. Intracellular bacteria were not found in sputum by PCR. Common bacterial pathogens were identified in 42% of acute exacerbation patients, most frequently Branhamella catarrhalis, Streptococcus pneumoniae and Haemophilus influenzae. Viral or virus and bacteria co‐infections were detected in 27% of acute exacerbation events (n = 12) with HRV and hMPV involved in 92% of cases. Patients with co‐infections did not present greater clinical severity scores at exacerbation and more recurrence of acute exacerbation events at 3 and 6 months than those with single infections (P > 0.4). These results suggest that HRV and hMPV may be contributors or cofactors of AECOPD. These findings indicate that viral or virus and bacterial co‐infections do not impact significantly on the clinical severity of acute exacerbation of chronic obstructive pulmonary disease and recurrence at 3 and 6 months. J. Med. Virol. 85:866–873, 2013. © 2013 Wiley Periodicals, Inc.     

Ten years of human metapneumovirus research

Described for the first time in 2001, human metapneumovirus (hMPV) has become one of the main viral pathogens responsible for acute respiratory tract infections in children but also in the elderly and immuno-compromised patients. The pathogen most closely related to hMPV is human respiratory syncytial virus (hRSV), the most common cause of bronchiolitis and pneumonia in young children. hMPV has been classified into two main viral groups A and B and has a seasonal distribution in temperate countries with most cases occurring in winter and spring. Given the difficulties encountered in culturing hMPV in vitro, diagnosis is generally achieved using real-time polymerase chain reaction.Like other Paramyxoviridae, hMPV has a negative-sense single-stranded RNA genome that includes 8 genes coding for 9 different proteins. The genomic organization and functions of surface attachment and fusion glycoproteins are relatively similar to those of hRSV. Although many groups have studied the viral life cycle of hMPV, many questions remain unanswered concerning the exact roles of the viral proteins in the attachment, fusion and replication of hMPV.To date, there remains no approved modality to combat hMPV infections. The majority of treatments that have been tested on hMPV have already demonstrated activity against hRSV infections. Some innovative approaches based on RNA interference and on fusion inhibitors have shown efficacy in vitro and in animal studies and could be beneficial in treating human hMPV disease. Difficulties faced inducing a durable immune response represent the biggest challenge in the development of an effective hMPV vaccine. Several strategies, such as the use of live-attenuated viruses generated by reverse genetics or recombinant proteins, have been tested in animals with encouraging results.     

Two-year prospective study of single infections and co-infections by respiratory syncytial virus and viruses identified recently in infants with acute respiratory disease

A prospective 2-year analysis including 322 infant patients with acute respiratory disease (ARD) hospitalized in a pediatric department in northern Italy was carried out to evaluate the role as respiratory pathogens or co-pathogens of recently identified viruses. The presence of respiratory syncitial virus (RSV), human Metapneumoviruses (hMPVs), human Bocaviruses (hBoVs), and human Coronaviruses (hCoVs) was assayed by molecular detection and clinical symptoms evaluated. Nasopharyngeal aspirates from 150 of the 322 infants (46.6%) tested positive for at least one pathogen. Ninety samples (28.0%) tested positive for RSV RNA (61.5% genotype A and 38.5% genotype B), 46 (14.3%) for hMPV RNA (71.7% subtype A and 28.3% subtype B), 28 (8.7%) for hCoV RNA (39.3% hCoV-OC43, 35.7% hCoV-NL63, 21.4% hCoV-HKU1, and 3.6% hCoV-229E), and 7 (2.2%) for hBoV DNA (of the 6 typed, 50% subtype 1 and 50% subtype 2); 21/150 samples revealed the presence of 2 or more viruses. Co-infection rates were higher for hMPVs, hCoVs, and hBoV (38.3%, 46.4%, and 57.1%,) and lower for RSV (23.3%). RSV was associated with the presence of complications (P < 0.001) and hypoxia (P < 0.015). When the presence of RSV alone and the RSV-hMPV co-infections were considered, RSV mono-infected patients resulted to have longer hospitalization and higher hypoxia (P < 0.001). The data highlight that (i) RSV has a central role as a respiratory pathogen of infants, (ii) the wide circulation of recently identified viruses does not reduce the clinical and epidemiological importance of RSV, and that (iii) recently identified agents (hMPVs, hBoVs, and hCoVs) act as primary pathogens or co-pathogens.     

人偏肺病毒A与B基因型对小鼠致病性的观察

目的 观察人偏肺病毒( human metapneumovirus,hMPV)A和B亚型感染BALB/c小鼠后,小鼠在体重、病毒载量、病理及气道反应性等方面的改变,探讨两亚型病毒致病性上的差异,为hMPV的进一步研究提供基础资料.方法 hMPV感染BALB/c动物模型后,通过使用real-time RT-PCR方法检测鼠肺内病毒载量的变化,肺功能检测仪监测小鼠气道反应性的变化及组织系统评分判定病理改变情况,来观测两亚型致病性的差异.结果 hMPV两亚型感染组,在体重的动态监测、病毒载量、肺组织病理改变及气道反应性上差异均无统计学意义.结论 hMPV两基因型感染BALB/c小鼠的致病性无差异.     

Genetic heterogeneity of G and F protein genes from Argentinean human metapneumovirus strains

Human metapneumovirus (hMPV) is a newly identified paramixovirus, associated with respiratory illnesses in all age groups. Two genetic groups of hMPV have been described. The nucleotide sequences of the G and F genes from 11 Argentinean hMPV strains (1998-2003) were determined by RT-PCR and direct sequencing. Phylogenetic analysis showed that hMPV strains clustered into two main genetic lineages, A and B. Strains clustered into A group were split into two sublineages, A1 and A2. All strains belonging to group B clustered with representative strains from sublineage B1. No Argentinean strains belonged to sublineage B2. F sequences showed high percentage identities at nucleotide and amino acid levels. In contrast, G sequences showed high diversity between A and B groups. Most changes observed in the deduced G protein sequence were amino acid substitutions in the extracellular domain, and changes in stop codon usage leading to different lengths in the G proteins. High content of serine and threonine residues were also shown, suggesting that this protein would be highly glycosylated. The potential sites for N- and O-glycosylation seem to have a different conservation pattern between the two main groups. This is the first report on the genetic variability of the G and F protein genes of hMPV strains in South America. Two main genetic groups and at least three subgroups were revealed among Argentinean hMPV strains. The F protein seems to be highly conserved, whereas the G protein showed extensive diversity between groups A and B.     

Epidemiology of respiratory viruses in bronchoalveolar lavage samples in a tertiary hospital

BackgroundThe prevalence of respiratory viruses in adults is largely underexplored, as most studies focus on children. Additionally, in severely ill or immunocompromised adults, where respiratory infections are mostly attributed to bacteria and fungi; respiratory viruses can lead to severe complications.ObjectivesTo evaluate the epidemiology of respiratory viruses in bronchoalveolar lavage fluid (BAL) specimens from patients with lower respiratory tract disease. The study population consisted of different groups including immunocompetent patients (control patients), solid organ transplant recipients, patients with haematological malignancies and other immunocompromised adults.Study designA total of 134 BAL fluid specimens collected during 2009–2011 were retrospectively assessed with the new commercial multiplex real-time PCR FTD Respiratory 21 Plus^®, targeting 18 different viruses and 2 atypical bacterial pathogens.ResultsViral or atypical bacterial pathogens were detected in 29.1% of BAL fluid specimens. Coronaviruses were most prevalent (13.4%), followed by rhinoviruses (5.2%), RSV (4.5%) and bocaviruses (3.7%). Comparing the total number of viruses detected, a statistically significant difference was observed between the control group and patients with haematological malignancies (27.5% vs. 57.1%, p < 0.05).ConclusionIn conclusion, our study highlights the high prevalence of respiratory viruses in BAL fluid specimens from adult patients with lower respiratory tract disease. The methods to be used should be sensitive and cover a wide range of potential pathogens. The specific patient population can also influence the detection rates of respiratory viruses.     

Diversity in Glycosaminoglycan Binding Amongst hMPV G Protein Lineages

We have previously shown that hMPV G protein (B2 lineage) interacts with cellular glycosaminoglycans (GAGs). In this study we examined subtypes A1, A2 and B1 for this interaction. GAG-dependent infectivity of available hMPV strains was demonstrated using GAG-deficient cells and heparin competition. We expressed the G protein ectodomains from all strains and analysed these by heparin affinity chromatography. In contrast to the B2 lineage, neither the A2 or B1 G proteins bound to heparin. Sequence analysis of these strains indicated that although there was some homology with the B2 heparin-binding domains, there were less positively charged residues, providing a likely explanation for the lack of binding. Although sequence analysis did not demonstrate well defined positively charged domains in G protein of the A1 strain, this protein was able to bind heparin, albeit with a lower affinity than G protein of the B2 strain. These results indicate diversity in GAG interactions between G proteins of different lineages and suggest that the GAG-dependency of all strains may be mediated by interaction with an alternative surface protein, most probably the conserved fusion (F) protein. Analysis of both native and recombinant F protein confirmed that F protein binds heparin, supporting this conclusion.