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Amorphous poly-β-hydroxybutyrate (PHB)-accumulating Bacillus spp. as biocontrol agents in crustacean culture

Joseph Laranja (UGent)
(2017)
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Promoter
(UGent) , (UGent) and Edgar Amar
Organization
Abstract
The latest FAO report showed that almost half of aquatic food products worldwide are produced from the aquaculture sector, thus making the sector a significant contributor of food fish products for the current 7.3 billion people in the planet. However, the development and expansion of aquaculture has always been hampered by diseases, thereby affecting its worldwide production and trade. Hence, sustainable measures to combat diseases for aquatic farmed animals have been the primary focus for research for the further expansion of the aquaculture sector. In this research, Poly-beta-hydroxybutyrate (PHB)-accumulating Bacillus spp. was explored and tested as new biological control agents for aquaculture. The (novel) strategy of using amorphous PHB (an antimicrobial compound) where the biopolymer is still contained in (live) bacteria that are often associated with strong probiotic effects (e.g. Bacillus spp.) may further advanced the capabilities of both PHB and the probiotic strategy as important sustainable disease-controlling measures for aquaculture. In this research, the study has demonstrated for the first time the strategy for isolating and rapid screening of PHB-accumulating Bacillus species from pasteurized shrimp pond sediments using the aid of MALDI-TOF MS as a tool for dereplication (i.e. bacterial grouping). The superior PHB-accumulating Bacillus isolate identified in this study was Bacillus sp. JL47 and has the capability to accumulate amorphous PHB at 28.6% PHB (on cell dry weight) after 24 h or 55% PHB after 48 h of culture in LB medium with 2% glucose. Under the fully controlled axenic culture system, the study has demonstrated the protective effects of the superior PHB accumulator Bacillus sp. JL47 strain in gnotobiotic Artemia against pathogenic Vibrio campbellii infection. The study also showed that the protective effect of Bacillus sp. JL47 was superior when the bacterium contains high amount of amorphous PHB, irrespective of the cell density used suggesting that the amorphous PHB present in the Bacillus cell is a main determinant in the protective effects of the PHB-accumulating Bacillus sp. JL47 on Artemia. Furthermore, the study has demonstrated the beneficial effects of using PHB-accumulating Bacillus spp. in Penaeus monodon postlarvae cultured in tanks wherein the shrimps fed the PHB-accumulating Bacillus spp. showed an improved survival, enhanced growth performance and higher resistance against pathogenic Vibrio campbellii LMG 21363, with a maximum attained for the shrimp supplied of the highest PHB accumulator (i.e. Bacillus sp. JL47). Likewise, feeding the P. monodon postlarvae with Artemia enriched with the PHB-accumulating Bacillus sp. JL47 at 0.5 g L-1 (wet weight) resulted in an up-regulation in the expression of prophenoloxidase and transglutaminase genes in the shrimp before and after the Vibrio campbellii challenge. The data suggest that the protective effect of the PHB-accumulating Bacillus sp. JL47 demonstrated in the previous in vivo work can be attributed to the immune-enhancing effects of the PHB-accumulating Bacillus sp. JL47 in the shrimp. Lastly, the study also investigated the (possible) contribution of phasin protein (i.e. a PHB-granule associated protein covering the surface of the amorphous PHB) on the survival of gnotobiotic Artemia challenged with pathogenic V. campbellii by cloning the protein and overproduced it in E. coli cells. The results however showed no protective effect of the recombinant E. coli containing the overproduced phasin protein in Artemia during Vibrio campbellii infection. In conclusion, the results presented in this research demonstrated the efficiency of PHB-accumulating Bacillus spp. as new biological control agents for aquaculture. The possible synergistic effect of amorphous PHB and the other probiotic properties of the Bacillus spp. may have contributed this effect. However, further research is needed to elucidate (clearly) the relative contributions of the stored PHB and the probiotic activity of the bacilli in the host.

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Citation

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Chicago
Laranja, Joseph. 2017. “Amorphous Poly-β-hydroxybutyrate (PHB)-accumulating Bacillus Spp. as Biocontrol Agents in Crustacean Culture”. Ghent, Belgium: Ghent University. Faculty of Bioscience Engineering.
APA
Laranja, J. (2017). Amorphous poly-β-hydroxybutyrate (PHB)-accumulating Bacillus spp. as biocontrol agents in crustacean culture. Ghent University. Faculty of Bioscience Engineering, Ghent, Belgium.
Vancouver
1.
Laranja J. Amorphous poly-β-hydroxybutyrate (PHB)-accumulating Bacillus spp. as biocontrol agents in crustacean culture. [Ghent, Belgium]: Ghent University. Faculty of Bioscience Engineering; 2017.
MLA
Laranja, Joseph. “Amorphous Poly-β-hydroxybutyrate (PHB)-accumulating Bacillus Spp. as Biocontrol Agents in Crustacean Culture.” 2017 : n. pag. Print.
@phdthesis{8533593,
  abstract     = {The latest FAO report showed that almost half of aquatic food products worldwide are produced from the aquaculture sector, thus making the sector a significant contributor of food fish products for the current 7.3 billion people in the planet. However, the development and expansion of aquaculture has always been hampered by diseases, thereby affecting its worldwide production and trade. Hence, sustainable measures to combat diseases for aquatic farmed animals have been the primary focus for research for the further expansion of the aquaculture sector. In this research, Poly-beta-hydroxybutyrate (PHB)-accumulating Bacillus spp. was explored and tested as new biological control agents for aquaculture. The (novel) strategy of using amorphous PHB (an antimicrobial compound) where the biopolymer is still contained in (live) bacteria that are often associated with strong probiotic effects (e.g. Bacillus spp.) may further advanced the capabilities of both PHB and the probiotic strategy as important sustainable disease-controlling measures for aquaculture.
In this research, the study has demonstrated for the first time the strategy for isolating and rapid screening of PHB-accumulating Bacillus species from pasteurized shrimp pond sediments using the aid of MALDI-TOF MS as a tool for dereplication (i.e. bacterial grouping). The superior PHB-accumulating Bacillus isolate identified in this study was Bacillus sp. JL47 and has the capability to accumulate amorphous PHB at 28.6\% PHB  (on cell dry weight) after 24 h or 55\% PHB after 48 h of culture in LB medium with 2\% glucose. Under the fully controlled axenic culture system, the study has demonstrated the protective effects of the superior PHB accumulator Bacillus sp. JL47 strain in gnotobiotic Artemia against pathogenic Vibrio campbellii infection. The study also showed that the protective effect of Bacillus sp. JL47 was superior when the bacterium contains high amount of amorphous PHB, irrespective of the cell density used suggesting that the amorphous PHB present in the Bacillus cell is a main determinant in the protective effects of the PHB-accumulating Bacillus sp. JL47 on Artemia. Furthermore,  the study has demonstrated the beneficial effects of using PHB-accumulating Bacillus spp. in Penaeus monodon postlarvae cultured in tanks wherein the shrimps fed the PHB-accumulating Bacillus spp. showed an improved survival, enhanced growth performance and higher resistance against pathogenic Vibrio campbellii LMG 21363, with a maximum attained for the shrimp supplied of the highest PHB accumulator (i.e. Bacillus sp. JL47). Likewise,  feeding the P. monodon postlarvae with Artemia enriched with the PHB-accumulating Bacillus sp. JL47 at 0.5 g L-1 (wet weight) resulted in an up-regulation in the expression of prophenoloxidase and transglutaminase genes in the shrimp before and after the Vibrio campbellii challenge. The data suggest that the protective effect of the PHB-accumulating Bacillus sp. JL47 demonstrated in the previous in vivo work can be attributed to the immune-enhancing effects of the PHB-accumulating Bacillus sp. JL47 in the shrimp. Lastly, the study also investigated the (possible) contribution of phasin protein (i.e. a PHB-granule associated protein covering the surface of the amorphous PHB) on the survival of gnotobiotic Artemia challenged with pathogenic V. campbellii by cloning the protein and overproduced it in E. coli cells. The results however showed no protective effect of the recombinant E. coli containing the overproduced phasin protein in Artemia during Vibrio campbellii infection.
In conclusion, the results presented in this research demonstrated the efficiency of PHB-accumulating Bacillus spp. as new biological control agents for aquaculture. The possible synergistic effect of amorphous PHB and the other probiotic properties of the Bacillus spp. may have contributed this effect. However, further research is needed to elucidate (clearly) the relative contributions of the stored PHB and the probiotic activity of the bacilli in the host.},
  author       = {Laranja, Joseph},
  isbn         = {9789463570404},
  language     = {eng},
  pages        = {XX, 262},
  publisher    = {Ghent University. Faculty of Bioscience Engineering},
  school       = {Ghent University},
  title        = {Amorphous poly-\ensuremath{\beta}-hydroxybutyrate (PHB)-accumulating Bacillus spp. as biocontrol agents in crustacean culture},
  year         = {2017},
}