Elsevier

Fish & Shellfish Immunology

Volume 47, Issue 2, December 2015, Pages 664-673
Fish & Shellfish Immunology

Full length article
Live Edwardsiella tarda vaccine enhances innate immunity by metabolic modulation in zebrafish

https://doi.org/10.1016/j.fsi.2015.09.034Get rights and content

Highlights

  • GC/MS based metabolomics is used to reveal metabolic mechanisms in Zebrafish in response to live E.tarda vaccine.

  • The E.tarda vaccine activates biosynthesis of unsaturated fatty and the TCA cycle.

  • Hydroxyl radical is limited during stimulation.

  • Oleate induces effective protection against E.tarda infection.

Abstract

Control of bacterial infection resides in the core of human health and sustainable animal breeding. Vaccines as an economic and efficient immunoprophylaxis have been widely accepted, but mechanisms for vaccines do not fully understand. Information regarding to metabolome in response to vaccines is not available. Here we explore the metabolic features by using GC/MS based metabolic profile and trace metabolic mechanisms in zebrafish (Dario rerio) in response to live Edwardsiella tarda vaccine. Pathway enrichment analysis shows that live vaccine activates biosynthesis of unsaturated fatty acids and the TCA cycle and reduces aminoacyl-tRNA biosynthesis, suggesting a metabolic characteristic feature in response to the live vaccine. We further demonstrate that hydroxyl radical is limited during stimulation. Finally, we reveal oleate induces effective protection against E. tarda infection. These results have implications for immunity study that metabolic regulation contributes to immune protection. Our findings enable us to propose novel therapeutic strategies on metabolism against bacterial infections.

Introduction

Recently developed metabolomics is a high-throughput approach to detect and profile changes in metabolites which are the end products of cellular processes. It can discover the metabolic pathways which are linked to biological mechanisms, shape the understanding of medicines how to play an action and reprogram the metabolism by exogenous complementation [1]. Gas chromatography/mass spectrometry (GC/MS), liquid chromatography-mass spectrometry (LC/MS) and nuclear magnetic resonance (NMR) are the three most used analytical technologies to investigate metabolic profiling [2]. Out of them, GC/MS has been widely used since it possesses three golden standards, high chromatographic resolution, very reproducible chromatography and searchable mass spectra libraries [3].

Immunometabolism is an emerging field at the interface between immune response and metabolism, showing how immune cells regulate metabolism and metabolic pathways regulates immune response outcome [4]. The multilevel interactions between the metabolic and immune systems promote our understanding of pathogenic mechanisms and offer substantial therapeutic promise. Investigation on immunometabolism has been reported in Drosophila melanogaster infected with Listeria monocytogenes, mice and human infected with Staphylococcus aureus, human infected with Mycobacterium tuberculosis using metabolomics approach [5], [6], [7]. Very recently, we have used the GC/MS based metabolomics identifies biomarkers that differentiate death from survival carps infected with Edwardsiella tarda [8]. We have demonstrated that crucial biomarkers identified by the metabolomics approach may elevate survival of tilapia and zebrafish infected with bacterial pathogens [9], [10], [11]. Furthermore, the developed functional metabolomics discloses an unknown mechanism of fatty acids by which IFN-α2b protects host from microbial infections [10]. These results indicate that investigation on metabolomics contributes to an understanding of immunological mechanisms and immune metabolic regulation.

Immunoprophylaxis is an effective measure to prevent and control infections [12]. Live vaccine, activated vaccine and subunit vaccine are three types of vaccines which have been widely used in the immunoprophylaxis. Gram-negative enteric pathogen E. tarda causes an infectious disease, called edwardsiellosis, in a wide range of hosts including fish, amphibians, reptiles and mammals [13], [14], [15], [16], [17]. This infection leads to serious economic losses and a threat to humans [18], [19]. Commercial vaccines against E. tarda are not available, although empirical vaccine candidate development strategy works well [20], [21], [22], [23]. Limited understanding of protective mechanisms of the experimental vaccines leads to an increased interest in exploring metabolic reconstruction by vaccines. The goal of the present study is to investigate metabolite profiling of zebrafish in response to live E. tarda vaccines for understanding vaccine mechanisms and identifying crucial metabolites that modulate innate immune response. The results obtained are reported as follows.

Section snippets

Bacterial strain, growth condition and vaccine preparation

E. tarda EIB202 was provided by Professor Yuanxing Zhang, East China University of Science and Technology. All chemicals were purchased from Sigma–Aldrich. Single colony of E. tarda EIB202 was picked from plate and propagated overnight in 5 mL tryptic soybroth (TSB) medium at 200 rpm (Sky-111B, SuKun, China) at 30 °C. Aliquot 1 mL culture was diluted into 1:100 using fresh TSB medium and grown at 30 °C. Bacterial cells were harvested at an OD600 of 1.0 by centrifugation at 6000 g (Universal

Protective efficiency of bacterial immunostimulation and metabolomics profiling of zebrafish humoral fluid

To identify the functional pathways and metabolites that were required for immunity after vaccination with live E. tarda, we investigated metabolomes of zebrafish exposed or not exposed to the bacterium as outlined in Fig. 1A. Zebrafish was infected by E. tarda and the resulting accumulating death showed that significant immune protection was found at p < 0.05 (Log-rank test) between the exposure and control (Fig. 1B). On the other hand, we collected humoral fluid at 48 h after these injections

Discussion

A line of evidences has demonstrated that vaccine induces not only adaptive immunity but also innate immunity [35]. Recently, regulation of metabolites to immunity has been reported [36]. Elevated crucial metabolites identified from a stress-related metabolome enhance the metabolome's ability against the stress such as bacterial infection [9]. The present study reveals that vaccine can mounts metabolic strategy to cope with microbial infection. This finding is also consistent with our recent

Acknowledgments

This work was sponsored by grants from Science and Technology Program of Guangzhou (201504010025), NSFC Projects (41276145, 31272702), “973” Project (2012CB114406), Doctoral Fund of Ministry of Education of China (20120171110008).

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