Elsevier

Journal of Clinical Virology

Volume 76, March 2016, Pages 45-50
Journal of Clinical Virology

A case report demonstrating the utility of next generation sequencing in analyzing serial samples from the lung following an infection with influenza A (H7N9) virus

https://doi.org/10.1016/j.jcv.2015.12.013Get rights and content

Highlights

  • Bacterial pneumonia is a well-recognized sequela of patient suffering from influenza.

  • Lung microbiome of serial BAL samples were characterized by using NGS longitudinally.

  • Lung microbiome of the patient was dominated by Acinetobacter baumannii after the viral invasion.

  • Kinetics of serum cytokines related to viral and secondary bacterial infection were analyzed.

Abstract

Background

Bacterial pneumonia is a well-recognized sequela of patient suffering from influenza, and a key factor, with cytokine dysregulation, that contribute to severe disease and mortality.

Objectives

To obtain a comprehensive assessment of lung microbial community dynamics in a fatal influenza H7N9 case during the whole clinical course, we undertook a longitudinal study.

Study design

Serial bronchoalveolar lavage fluid samples were collected from a H7N9 patient after illness onset, and the microbiome was characterized by using next-generation sequencing and microbiological approaches. Furthermore, the kinetics of circulating cytokine storms related to viral and secondary bacterial infection were analyzed.

Results

Within complex and dynamic communities, the lung microbiome with H7N9 infection were dominated by gram-negative bacteria, Acinetobacter baumannii after the viral invasion and during the whole clinical course. Sputum and blood culture confirmed the secondary bacterial infection with multidrug-resistant A. baumannii 9 days later. The dynamics of the bacterial infection with carbapenem-resistant A. baumannii correlated with antibiotic therapy. Our observations also indicated that sustained high levels of host inflammatory factors, consisting of a set of distinct cytokines associated with disease stage, may contribute to disease progression and death.

Conclusions

This study demonstrates an initial attempt to explore the dynamic microbiome involved inH7N9 infection and its response to antimicrobial therapy, as well as host cytokine response to infection by using next-generation sequencing. These type of investigations with longitudinal follow-up to understand dynamics of microbial community and cytokines involved in lung infection may provide opportunities for development and optimization of targeted antimicrobial therapy and even new therapeutic strategies.

Section snippets

Background

Influenza and pneumonia remain the leading infectious causes of death worldwide [1], resulting in almost 3.5 million deaths yearly and are a persistent and pervasive public health issue [2]. Bacterial pneumonia and cytokine dysregulation in patients suffering from influenza are key factors that contribute to severe disease and mortality [3], [4], [5]. The microbes involved in lung infection are complex, with organisms ranging in virulence from commensal to highly pathogenic, and viral-bacterial

Objectives

We describe an initial attempt to explore the structure and dynamics of the microbiome correlated with lung infection by using NGS. Serial bronchoalveolar lavage fluid (BALF) samples were collected from a fatal H7N9-related case with severe pneumonia and acute respiratory distress syndrome (ARDS) and analyzed by NGS. The comprehensive pulmonary microbial community kinetics, and host inflammatory response during therapy were studied.

Patient

A 61-year-old female patient with severe pneumonia was admitted on 18 July 2013 [10], [11]. She had been healthy except for cholecystectomy 30 years prior. She started to experience fever and general malaise on 13 July (day 0 after illness onset) and sought medical attention on 14 July. A chest radiograph on 15 July revealed interstitial pneumonia in the left upper lung. Progressive dyspnea developed on 16 July and follow-up chest radiographs showed ground grass opacity and lower lung

Deep sequencing summary

In total eight BALF samples were collected at intervals of 2 days during the whole clinical course and processed and run on Ion Torrent PGM, retrospectively. The turnaround time of the whole process from each sample preparation to data back from the NGS was about 48 h.

To test for the presence of potential contaminants in the reagents used for nucleic acids extraction and sequencing library preparation, we prepared a control sample consisting of a defined template (Lambda gDNA, Life Technologies,

Discussion

We demonstrate an initial attempt to use unbiased NGS to explore the dynamics of lung microbiome with novel influenza A (H7N9) virus infection. The NGS method allowed us to perform metagenomic analyses of pulmonary infection within a reasonable timeframe. These findings strongly suggest that NGS methods can complement conventional diagnostics and also highlight their potential to aid clinical personnel and health agencies in making appropriate decisions. Our observations also indicate that

Funding

This work was supported by the National S&T Major Project, “China Mega-Project for Infectious Disease” (grant No. 2013ZX10004-001), the Nonprofit Industry Research Project of Chinese National Health and Family Planning Commission (grant No. 201402001), and National Science Fund for Distinguished Young Scholars (grant number 81425001/H0104) for Dr. Bin Cao, and the PUMC Youth FUND (grant No. 33320140031).

Competing interests

The authors have no competing interests to report.

Ethical approval

No ethical approval is required for this study.

References (22)

  • M.R. Wilson et al.

    Actionable diagnosis of neuroleptospirosis by next-generation sequencing

    N. Engl. J. Med.

    (2014)
  • Cited by (8)

    View all citing articles on Scopus
    View full text