Shotgun metagenomics reveals significant gut microbiome features in different grades of acute pancreatitis

https://doi.org/10.1016/j.micpath.2021.104849Get rights and content

Highlights

  • Gut microbiota in AP patients was characterized by decreased species richness.

  • The representative gut microbiota in MAP, MSAP, and SAP was StreptococcusEscherichia-coli, and Enterococcus, respectively.

  • Glutathione metabolism, lipopolysaccharide biosynthesis, and amino acid metabolism were enriched in MAP, MSAP, and SAP.

Abstract

Background

Acute pancreatitis (AP) has a broad spectrum of severity and is associated with considerable morbidity and mortality. Dysbiosis of gut microbiota may be associated with AP severity.

Aims

We aimed to evaluate the composition and functional effects of gut microbiota in different grades of AP severity.

Methods

We carried out shotgun metagenomic sequencing on rectal swab samples from three patients with mild acute pancreatitis (MAP), three with moderately severe acute pancreatitis (MSAP), three with severe acute pancreatitis (SAP) and three normal control persons (NOR). Differences analysis in gut microbiota composition and functional enrichment was performed.

Results

Gut microbiota in AP patients was characterized by decreased species richness. The most representative gut microbiota in mild acute pancreatitis (MAP), moderately severe acute pancreatitis (MSAP), and severe acute pancreatitis (SAP) was Streptococcus, Escherichia-coli, and Enterococcus, respectively. Each of the three AP-associated genera could differentiate AP from healthy control population. Representative pathways associated with the glutathione metabolism, lipopolysaccharide biosynthesis, and amino acid metabolism (valine, leucine and isoleucine degradation) were enriched in MAP, MSAP, and SAP, respectively.

Conclusions

The study shows a potential association of gut microbiome composition and function to the progression of AP severity.

Introduction

Acute pancreatitis (AP) is the most common gastrointestinal disease that requires acute admission and causes tremendous pain and socioeconomic burden [1]. The worldwide incidence of AP ranges from 4.9 to 80 cases per 100,000 individuals annually [2]. According to the Revised Atlanta Classification, AP is stratified into mild acute pancreatitis (MAP), moderately severe acute pancreatitis (MSAP), and severe acute pancreatitis (SAP) [3]. The outcome of AP patients with different severity is widely divergent. For instance, patients with MAP only need supportive care such as fluid and analgesia and recover within a few days. But in the non-mild form of AP, an inflammatory cascade leads to local and systemic complications, claims remarkable morbidity and mortality, and makes the management of AP challenging [4]. The pathogenetic mechanism underlying the progression of AP severity is yet to be elucidated [5].

Gut microbiota plays an essential role in maintaining immune homeostasis and the biological barrier of the intestine [6]. Under pathological conditions such as AP, perturbations to the gut microbiota could disrupt gut barrier, increase the intestinal permeability, and lead to bacterial translocation, which in turn triggers secondary infectious complications [7]. Several bacteria, including Escherichia, Shigella, Enterococcus, and Enterobacteriaceae family, have been found in the pancreas, indicating that their translocation from the gut could lead to infected pancreatic necrosis in non-mild AP [8]. Indeed Li et al. reported that the abundance of potentially pathogenic bacteria such as Enterococcus and Enterobacteriaceae is significantly increased, and that of beneficial bacteria such as Bifidobacterium is significantly decreased in gut microbiota of patients with MAP and SAP [9]. Our previous work revealed different gut bacteria composition in AP patients with different grades of severity. To be specific, Bacteroides, Escherichis-Shigella, and Enterococcus was the dominant gut microbiota species in MAP, MSAP, and SAP, respectively. Besides, Anaerococcus and Enterococcus were significantly increased and Eubacterium hallii decreased in non-mild AP patients [10]. Similar findings were also reported by Zhu Y et al. [7]. These studies together indicated a potential association between gut microbiota dysbiosis and AP progression. However, what remains obscure is what functional effects gut microbiota may exert upon the worsening of AP.

Shotgun metagenomic sequencing of the whole DNA provides valuable information about the functions of the microbial community [11]. The sequencing technology is used to obtain the entire genomic content of the microbiome and achieve accurate taxonomic classification and functional assignments. Also, it is able identify novel functional genes, antibiotic resistance genes, microbial pathways, and functional dysbiosis of the gut microbiome [12]. In our previous study, we performed 16S rRNA sequencing to find potential pathogenic microorganisms in patients with different severity of AP [10]. In order to further evaluate the composition and functional effects of gut microbiota in different grades of AP severity, we conducted a metagenomic shotgun survey on intestinal microbiota of MAP, MSAP, SAP patients and healthy controls, in order to characterize the AP-related composition and functional changes.

Section snippets

Subjects recruitment

The 2012 revised Atlanta classification stratified the clinical severity of AP patients into three categories: mild AP (MAP), moderately severe AP (MSAP), and severe AP (SAP) [3]. All the patients with a diagnosis of AP based on the 2012 Revised Atlanta Classification and admitted to Peking Union Medical College Hospital, Beijing, China were eligible for inclusion if they were enrolled within 48 h of the onset of symptoms from June to December 2019. A total of three MAP, three MSAP, and three

Clinical characteristics of patients and healthy subjects

A total of 9 AP patients (3 MAP, 3 MSAP, and 3 SAP) and 3 NOR were enrolled in this study. The NOR group were all women and they were 42.0 ± 14.2 years old with a BMI of 20.9 ± 2.3. The multivariate test showed that there was no individual selection bias in age and gender (supplementary Figure 3). The clinical characteristics of AP groups were shown in Table 1. Age (p = 0.473) and gender (p = 0.99) were comparable between the AP groups and the NOR group. Among the AP patients statistically

Discussion

In this study, we performed the Shotgun metagenomic sequencing approach to a cohort of 12 individuals, including 9 AP patients (3 MAP, 3 MSAP, and 3 SAP) and 3 NOR. We found remarkably different outcomes (Table 1) and significant dysbiosis of microbiome composition and function in the AP patients. Moreover, the composition and function of microbiota in SAP were different from MAP and MSAP, implying the possible association of the dysbiosis of microbial composition and function to AP severity.

Conclusions

We conducted a shotgun metagenomics survey on the gut microbiome of AP patients with three different severity grades for the first time. We identified several new AP-related bacteria and functional gene pathways, which extend the current knowledge about the role of gut microbiota in AP. Our findings may be useful for future studies investigating the mechanism of AP worsening and developing strategies for the treatment of AP.

Author statement

YSS, XYY and FYY contributed to collection of clinical data and fecal samples, and interpretation of data, and drafting of the article. WD, MX, and XJ contributed to the concept and design of the study, interpretation of data, and the critical revision of the study methods. CGR contributed to the critical revision of the article for relevant intellectual content. ZHD made critical review of the article for valuable intellectual content. WD and XJ contributed to the drafting of the article, and

Funding

This work was supported by the Beijing Natural Science Foundation (No. 7192162) and Grants from Peking Union Medical College (2019XK320036, 2019zlgc0503).

Declaration of competing interest

None.

Acknowledgments

The authors want to thank Dr. Zhang Yan for the editing of this paper.

References (62)

  • M.S. Desai et al.

    A dietary fiber-deprived gut microbiota degrades the colonic mucus barrier and enhances pathogen susceptibility

    Cell

    (2016)
  • Y. Zhu et al.

    Gut microbiota dysbiosis worsens the severity of acute pancreatitis in patients and mice

    J. Gastroenterol.

    (2019)
  • Q. Li et al.

    Bacteremia in patients with acute pancreatitis as revealed by 16S ribosomal RNA gene-based techniques

    Crit. Care Med.

    (2013)
  • X.Y. Li et al.

    Role of gut microbiota on intestinal barrier function in acute pancreatitis

    World J. Gastroenterol.

    (2020)
  • S. Yu et al.

    Identification of dysfunctional gut microbiota through rectal swab in patients with different severity of acute pancreatitis

    Dig. Dis. Sci.

    (2020)
  • S. Abubucker et al.

    Metabolic reconstruction for metagenomic data and its application to the human microbiome

    PLoS Comput. Biol.

    (2012)
  • W.L. Wang et al.

    Application of metagenomics in the human gut microbiome

    World J. Gastroenterol.

    (2015)
  • M. Singer et al.

    The third international consensus definitions for sepsis and septic shock (Sepsis-3)

    Jama

    (2016)
  • C.M. Bassis et al.

    Comparison of stool versus rectal swab samples and storage conditions on bacterial community profiles

    BMC Microbiol.

    (2017)
  • L.M. Biehl et al.

    Usability of rectal swabs for microbiome sampling in a cohort study of hematological and oncological patients

    PloS One

    (2019)
  • P.B. Eckburg et al.

    Diversity of the human intestinal microbial flora

    Science (New York, NY)

    (2005)
  • Nadia Gaci et al.

    Archaea and the human gut:New beginning of an old story

    World J. Gastroenterol.

    (2014)
  • A.I. Bordin et al.

    Effects of administration of live or inactivated virulent rhodococccus equi and age on the fecal microbiome of neonatal foals

    PloS One

    (2013)
  • J.J. Farrell et al.

    Variations of oral microbiota are associated with pancreatic diseases including pancreatic cancer

    Gut

    (2012)
  • V. Eeckhaut et al.

    Anaerostipes butyraticus sp. nov., an anaerobic, butyrate-producing bacterium from Clostridium cluster XIVa isolated from broiler chicken caecal content, and emended description of the genus Anaerostipes

    Int. J. Syst. Evol. Microbiol.

    (2010)
  • D. Ciocan et al.

    Characterization of intestinal microbiota in alcoholic patients with and without alcoholic hepatitis or chronic alcoholic pancreatitis

    Sci. Rep.

    (2018)
  • J.C. Marshall et al.

    The microbiology of multiple organ failure: the proximal gastrointestinal tract as an occult reservoir of pathogens

    (1988)
  • K. Bianca et al.

    Changes in colon gene expression associated with increased colon inflammation in interleukin-10 gene-deficient mice inoculated with Enterococcus species

    BMC Immunol.

    (2010)
  • C. Tan et al.

    Dysbiosis of intestinal microbiota associated with inflammation involved in the progression of acute pancreatitis

    Pancreas

    (2015)
  • G. Falony et al.

    Population-level analysis of gut microbiome variation

    Science (New York, NY)

    (2016)
  • H.Y. Gaisano et al.

    Supramaximal cholecystokinin displaces Munc18c from the pancreatic acinar basal surface, redirecting apical exocytosis to the basal membrane

    J. Clin. Invest.

    (2001)
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