Dynamics of bacteriophages in gut of giant pandas reveal a potential regulation of dietary intake on bacteriophage composition
Graphical abstract
Introduction
Similar dietary habits can lead to similar gut microbiome among animals of different phylogenetic origins (Hu et al., 2017; McKenney et al., 2018). However, this situation is not observed in giant pandas. In a study of gut microbiota of 59 different mammals, a profile clustered according to dietary habits was seen for most mammals (Ley et al., 2008), but the gut microbiota of giant pandas and red pandas that are mainly herbivores were clustered with that of carnivores. This finding was confirmed by other studies (McKenney et al., 2018; Xue et al., 2015). A series of cellulolytic activity assays of the gut microbiota of giant pandas (Guo et al., 2018a; McKenney et al., 2018; Xue et al., 2015; Yang et al., 2018; Zhang et al., 2018) revealed that the gut microbiome of giant pandas are similar to that of their carnivorous ancestors, regardless of their shift to a bamboo diet during evolution. Thus, the relationship between the dietary habits and gut microbiome of giant pandas remains uncertain.
Giant pandas have two major diet phases in life: milk diet (nursing stage, generally during 0–1.5 years of age) and bamboo diet (after the nursing stage). Although the gut microbiome of giant pandas displays extensive seasonal variations (Wu et al., 2017; Xue et al., 2015), it is consistently dominated by members of the Enterobacteriaceae (e.g. Escherichia, Shigella, Salmonella) in both cubs and adults (Guo et al., 2018a). During the nursing stage, the gut microbiota of cubs are abundant in milk-digesting bacteria, such as various species of Lactobacillus, Lactococcus, Leuconostoc, and Streptococcus (Guo et al., 2018a; McKenney et al., 2018; Xue et al., 2015; Yang et al., 2018; Zhang et al., 2018). Although the main diet of adult giant pandas is bamboo, their gut microbiota have an extremely low abundance of typical cellulolytic bacteria, such as Ruminococcus and Bacteroides (M. Chen et al., 2015; Li et al., 2010; Xue et al., 2015). Instead, Bacillus and Clostridia (Xue et al., 2015; Z. Zhou et al., 2015) with limited cellulolytic activity are abundant (Xue et al., 2015; Zhang et al., 2018; Zhu et al., 2011), which would account for the tiny loss of cellulose (~ 8%) in fecal bamboos (Dierenfeld et al., 1982).
In the ecosystem, bacteriophages may act as modulators (lysogenic phages) or predators (lytic phages) leading to changes in the composition of bacterial microbiome (Coutinho et al., 2017; Lim et al., 2015). The regulatory roles of bacteriophage are attributed to specific phage genes, such as auxiliary metabolic genes (Hurwitz et al., 2013; Thompson et al., 2011), genes encoding for proteins involved in homologous recombination, and antibiotic resistance genes (ARGs) (Casjens, 2003; Enault et al., 2017). A bacterial genome may carry functional or non-functional lysogenic phages as prophages that may be activated to become lytic, due to alterations in animal's living environment, bacterial DNA damage or mutation, or expression of certain phage homologous genes (Barksdale and Arden, 1974; Casjens, 2003; Coutinho et al., 2018; Fornelos et al., 2018).
A recent study demonstrated the importance of balanced relationship between bacteriophages and diet-related bacteria in the stability of gut microbiota (Rivett and Bell, 2018). We hypothesized that the diets have effects on the composition of bacteriophages in gut microbiota of giant pandas. In this study, we investigated the diversity and dynamics of gut bacteria and bacteriophages in giant pandas under different dietary conditions.
Section snippets
Fecal samples and DNA extraction
A giant panda family with four members living in the Macao Giant Panda Pavilion was investigated. This giant panda family includes the following members: Father, born in July 2007, designated P1; mother, born in August 2008, designated P2; elder brother, born on June 26, 2016, designated P3; Younger brother, also born on June 26, 2016, designated P4. The two cubs were born 3 h apart. They were fed with breast milk until 1.5 months of age, and then switched to formula milk (mixtures of dog and
Sequencing data processing, assembly and annotation
After removing reads of low sequencing quality and those containing adapter or eukaryotic (e.g., bamboo) gene sequences (Table S1), a total of 817.8 G clean data, including 495.6 G from adult and 322.1 G from cub samples (Table S2). The bacterial and viral reads of each sample were separately assembled as scaffolds (Table S3). The final non-redundant gene set contained 5,730,523 microbial genes (Table S4).
Dynamics of gut bacteriophages in the gut microbiota of giant pandas
Based on sequence reads, 641 bacteriophage species from 54 genera, 18 families, and 2
Discussion
Although a number of studies on the gut microbiome of giant pandas have been reported in recent years (Guo et al., 2018b; Wu et al., 2017; Xue et al., 2015; Yang et al., 2018; Zhang et al., 2018; Zhang et al., 2017), few of them focused on virome dynamics (Yang et al., 2018; Zhang et al., 2017), especially that of bacteriophages and bacteria in the gut microbiota of giant pandas. As bacteriophages interact with their host bacteria, they play a role in maintaining the functionality and stability
Conclusion
We investigated the dynamics of bacteriophages and their host bacteria in the gut microbiome of giant pandas with different dietary habits. Results showed that there was a high abundance of Escherichia phage, Enterobacteria phage, and Shigella phage, and a low abundance of milk-digesting bacteriophages such as Lactobacillus phage, Leuconostoc phage, and Streptococcus phage in the gut of adult giant pandas. A majority of gut bacteriophages of giant pandas were phages inside host cells (PIH). The
Ethics statement
Sample collection and all experiments were performed in a manner to minimize risk to the giant pandas and the environment. All experimental protocols of this study were approved by Institute of Chinese Medical Sciences - Animal Ethics Committee (ICMS-AEC) of the University of Macau.
Funding
This work was supported by Fundo dos Pandas (Macao, China), the Macau Science and Technology Development Fund (FDCT) and the Ministry of Science and Technology of China (MOST) joint funding scheme (Ref. No. FDCT 017/2015/AMJ), Research Committee, University of Macau (MYRG2016-00129-ICMS-QRCM), and GDAS Special Project of Science and Technology Development (2018GDASCX-0107).
CRediT authorship contribution statement
Min Guo:Conceptualization, Formal analysis, Methodology, Software, Writing - review & editing.Guilin Liu:Data curation, Formal analysis.Jianwei Chen:Software, Formal analysis.Jinmin Ma:Software.Jinzhong Lin:Formal analysis.Ying Fu:Formal analysis.Guangyi Fan:Software.Simon Ming-Yuen Lee:Supervision, Resources, Funding acquisition, Writing - review & editing.Libiao Zhang:Funding acquisition, Writing - review & editing.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgements
We thank the Macao Giant Panda Pavilion of the Civic and Municipal Affairs Bureau (Macao, China) for providing fecal samples of giant pandas. We also thank Shandong Technology Innovation Center of Synthetic Biology for their advices in the work.
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Contributed equally.