Lab-scale <i>in vitro</i> Mass Production of the Entomopathogenic Nematode <i>Heterorhabditis bacteriophora</i> Using Liquid Culture Fermentation Technology

Devang Upadhyay; Sivanadane Mandjiny; Rebecca Bullard-Dillard; Meredith Storms; Michael Menefee; Leonard D. Holmes

doi:doi:10.11648/j.bio.20150306.19

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Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology

Devang Upadhyay, Sivanadane Mandjiny, Rebecca Bullard-Dillard, Meredith Storms, Michael Menefee, Leonard D. Holmes

Published in American Journal of Bioscience and Bioengineering (Volume 3, Issue 6)

Received: 8 December 2015 Accepted: 17 December 2015 Published: 30 December 2015

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Abstract

The objective of this research is to develop fermentation methodology for the production of the biocontrol agent Heterorhabditis bacteriophora. Deployment of this organism will reduce the use of chemical insecticides which threaten the environment. This study shows how to produce the entomopathogenic nematode (EPN) Heterorhabditis bacteriophora and its bacterial symbiont Photorhabdus luminescens utilizing an in vitro, monoxenic liquid culture. EPNs were cultured in three different bioreactor working volumes of 1.5, 4 and 7 liters with initial nematode inoculation concentrations of approximately 2x10³/mL. Liquid nematode media was conditioned with the bacterial symbiont 24 hours prior to nematode inoculation. Within three days after inoculation, infective juveniles (IJs) developed into self-fertilizing hermaphrodites and eventually produced IJ offspring. Maximum nematode densities were obtained seven days post-nematode inoculation. All three working volumes (1.5, 4 and 7 liters) produced final yields of 4.6x10⁴ ± 2000 IJs/mL, 4.2x10⁴ ± 2200 IJs/mL and 3.9x10⁴ ± 2000 IJs/mL, respectively. In vitro scale-up technology can be further optimized for production of this biocontrol agent by improving media formulation, process parameters, bioreactor design and inoculation times that will maximize nematode yield.

Published in	American Journal of Bioscience and Bioengineering (Volume 3, Issue 6)
DOI	10.11648/j.bio.20150306.19
Page(s)	203-207
Creative Commons	This is an Open Access article, distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution and reproduction in any medium or format, provided the original work is properly cited.
Copyright	Copyright © The Author(s), 2024. Published by Science Publishing Group

Keywords

Heterorhabditis bacteriophora, Photorhabdus luminescens, Entomopathogenic Nematode, Fermentation Technology, Biocontrol Agent

References

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[3]	Shapiro-Ilan D, Gaugler R (2002) Production technology for entomopathogenic nematodes and their bacterial symbionts. Journal of Industrial Microbiology & Biotechnology. 28: 137 –146.
[4]	Patterson W, Upadhyay D, Mandjiny S, Bullard-Dillard R, Storms M, Menefee M, Holmes L (2015) Attractant Role of Bacterial Bioluminescence of Photorhabdus luminescens on a Galleria mellonella Model. American journal of life sciences 3(4): 290-294.
[5]	Strauch O, Ehlers R (1998) Food signal production of Photorhabdus luminescens inducing the recovery of entomopathogenic nematodes Heterorhabditis spp. in liquid culture. Applied Microbiology and Biotechnology 50: 369-374.
[6]	Gerdes E, Upadhyay D, Mandjiny S, Bullard-Dillard R, Storms M, Menefee M, Holmes L (2015) Photorhabdus Luminescens: Virulent Properties and Agricultural Applications. American Journal of Agriculture and Forestry 3(5): 171-177.
[7]	Yoo S, Brown I, Gaugler R (2000) Liquid media development for Heterorhabditis bacteriophora: lipid source and concentration. Applied Microbiology and Biotechnology 54: 759-763.
[8]	Ehlers RU (2001) Mass production of entomopathogenic nematodes for plant protection. Applied Microbiology and Biotechnology 56: 623-633.
[9]	Gaugler R (1997) Alternative paradigms for commercializing biopesticides. Phytoparasitica 25: 179-182.
[10]	Ehlers RU, ShapiroIlan DI (2005) Mass production 65-78pp. In: Nematode as biological control agent. Grewal, Ehler and Shapiro-Ilan, CAB International, Wallingford, UK.
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[12]	Wouts W (1981) Mass production of the entomopathogenous nematode Heterorhabditis bacteriophora (Nematode: Heterorhabditidae) on artificial media. Journal of Nematology 13: 467-469.
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[15]	Strauch O, Stoessel S, Ehlers R (1994) Culture conditions define automictic or amphimictic reproduction in entomopathogenic rhabditid nematodes of the genus Heterorhabditis. Fundamental & Applied Nematology 17(6): 575-582.
[16]	Kooliyottil R, Upadhyay D, Inman III FL, Mandjiny S, Holmes LD (2013) A Comparative Analysis of Entomoparasitic Nematodes Heterorhabditis bacteriophora and Steinernema carpocapsae. Open Journal of Animal Science 3(4): 326-333.
[17]	Jessen P, Strauch O, Wyss U, Luttmann R, Ehlers RU (2000) Carbon dioxide triggers dauer juvenile recovery of entomopathogenic nematodes (Heterorhabditis species.) Nematology 2: 310–324.
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[19]	Inman FL, Holmes LD (2012b) The effects of trehalose on the bioluminescence and pigmentation of the phase I variant of Photorhabdus luminescens. Journal of Life Science 6: 119-129.
[20]	Upadhyay D, Kooliyottil R, Mandjiny S, Inman III F, Holmes L (2013) Mass production of the beneficial nematode Steinernema carpocapsae utilizing a fed-batch culturing process. ESci Journal of Plant Pathology 02(01): 52-58.
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Devang Upadhyay, Sivanadane Mandjiny, Rebecca Bullard-Dillard, Meredith Storms, Michael Menefee, et al. (2015). Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology. American Journal of Bioscience and Bioengineering, 3(6), 203-207. https://doi.org/10.11648/j.bio.20150306.19

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ACS Style

Devang Upadhyay; Sivanadane Mandjiny; Rebecca Bullard-Dillard; Meredith Storms; Michael Menefee, et al. Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology. Am. J. BioSci. Bioeng. 2015, 3(6), 203-207. doi: 10.11648/j.bio.20150306.19

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AMA Style

Devang Upadhyay, Sivanadane Mandjiny, Rebecca Bullard-Dillard, Meredith Storms, Michael Menefee, et al. Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology. Am J BioSci Bioeng. 2015;3(6):203-207. doi: 10.11648/j.bio.20150306.19

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@article{10.11648/j.bio.20150306.19,
  author = {Devang Upadhyay and Sivanadane Mandjiny and Rebecca Bullard-Dillard and Meredith Storms and Michael Menefee and Leonard D. Holmes},
  title = {Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology},
  journal = {American Journal of Bioscience and Bioengineering},
  volume = {3},
  number = {6},
  pages = {203-207},
  doi = {10.11648/j.bio.20150306.19},
  url = {https://doi.org/10.11648/j.bio.20150306.19},
  eprint = {https://article.sciencepublishinggroup.com/pdf/10.11648.j.bio.20150306.19},
  abstract = {The objective of this research is to develop fermentation methodology for the production of the biocontrol agent Heterorhabditis bacteriophora. Deployment of this organism will reduce the use of chemical insecticides which threaten the environment. This study shows how to produce the entomopathogenic nematode (EPN) Heterorhabditis bacteriophora and its bacterial symbiont Photorhabdus luminescens utilizing an in vitro, monoxenic liquid culture. EPNs were cultured in three different bioreactor working volumes of 1.5, 4 and 7 liters with initial nematode inoculation concentrations of approximately 2x103/mL. Liquid nematode media was conditioned with the bacterial symbiont 24 hours prior to nematode inoculation. Within three days after inoculation, infective juveniles (IJs) developed into self-fertilizing hermaphrodites and eventually produced IJ offspring. Maximum nematode densities were obtained seven days post-nematode inoculation. All three working volumes (1.5, 4 and 7 liters) produced final yields of 4.6x104 ± 2000 IJs/mL, 4.2x104 ± 2200 IJs/mL and 3.9x104 ± 2000 IJs/mL, respectively. In vitro scale-up technology can be further optimized for production of this biocontrol agent by improving media formulation, process parameters, bioreactor design and inoculation times that will maximize nematode yield.},
 year = {2015}
}

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TY  - JOUR
T1  - Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology
AU  - Devang Upadhyay
AU  - Sivanadane Mandjiny
AU  - Rebecca Bullard-Dillard
AU  - Meredith Storms
AU  - Michael Menefee
AU  - Leonard D. Holmes
Y1  - 2015/12/30
PY  - 2015
N1  - https://doi.org/10.11648/j.bio.20150306.19
DO  - 10.11648/j.bio.20150306.19
T2  - American Journal of Bioscience and Bioengineering
JF  - American Journal of Bioscience and Bioengineering
JO  - American Journal of Bioscience and Bioengineering
SP  - 203
EP  - 207
PB  - Science Publishing Group
SN  - 2328-5893
UR  - https://doi.org/10.11648/j.bio.20150306.19
AB  - The objective of this research is to develop fermentation methodology for the production of the biocontrol agent Heterorhabditis bacteriophora. Deployment of this organism will reduce the use of chemical insecticides which threaten the environment. This study shows how to produce the entomopathogenic nematode (EPN) Heterorhabditis bacteriophora and its bacterial symbiont Photorhabdus luminescens utilizing an in vitro, monoxenic liquid culture. EPNs were cultured in three different bioreactor working volumes of 1.5, 4 and 7 liters with initial nematode inoculation concentrations of approximately 2x103/mL. Liquid nematode media was conditioned with the bacterial symbiont 24 hours prior to nematode inoculation. Within three days after inoculation, infective juveniles (IJs) developed into self-fertilizing hermaphrodites and eventually produced IJ offspring. Maximum nematode densities were obtained seven days post-nematode inoculation. All three working volumes (1.5, 4 and 7 liters) produced final yields of 4.6x104 ± 2000 IJs/mL, 4.2x104 ± 2200 IJs/mL and 3.9x104 ± 2000 IJs/mL, respectively. In vitro scale-up technology can be further optimized for production of this biocontrol agent by improving media formulation, process parameters, bioreactor design and inoculation times that will maximize nematode yield.
VL  - 3
IS  - 6
ER  -

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Author Information

Devang Upadhyay

Sartorius Stedim Biotechnology Laboratory, Biotechnology Research and Training Center, the University of North Carolina at Pembroke, Pembroke, USA
Sivanadane Mandjiny

Department of Chemistry and Physics, the University of North Carolina at Pembroke, Pembroke, USA
Rebecca Bullard-Dillard

School of Graduate Studies and Research, the University of North Carolina at Pembroke, Pembroke, USA
Meredith Storms

College of Arts & Sciences, the University of North Carolina at Pembroke, Pembroke, USA
Michael Menefee

Thomas Family Center for Entrepreneurship, the University of North Carolina at Pembroke, Pembroke, USA
Leonard D. Holmes

Sartorius Stedim Biotechnology Laboratory, Biotechnology Research and Training Center, the University of North Carolina at Pembroke, Pembroke, USA

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APA Style

Devang Upadhyay, Sivanadane Mandjiny, Rebecca Bullard-Dillard, Meredith Storms, Michael Menefee, et al. (2015). Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology. American Journal of Bioscience and Bioengineering, 3(6), 203-207. https://doi.org/10.11648/j.bio.20150306.19

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ACS Style

Devang Upadhyay; Sivanadane Mandjiny; Rebecca Bullard-Dillard; Meredith Storms; Michael Menefee, et al. Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology. Am. J. BioSci. Bioeng. 2015, 3(6), 203-207. doi: 10.11648/j.bio.20150306.19

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AMA Style

Devang Upadhyay, Sivanadane Mandjiny, Rebecca Bullard-Dillard, Meredith Storms, Michael Menefee, et al. Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology. Am J BioSci Bioeng. 2015;3(6):203-207. doi: 10.11648/j.bio.20150306.19

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@article{10.11648/j.bio.20150306.19,
  author = {Devang Upadhyay and Sivanadane Mandjiny and Rebecca Bullard-Dillard and Meredith Storms and Michael Menefee and Leonard D. Holmes},
  title = {Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology},
  journal = {American Journal of Bioscience and Bioengineering},
  volume = {3},
  number = {6},
  pages = {203-207},
  doi = {10.11648/j.bio.20150306.19},
  url = {https://doi.org/10.11648/j.bio.20150306.19},
  eprint = {https://article.sciencepublishinggroup.com/pdf/10.11648.j.bio.20150306.19},
  abstract = {The objective of this research is to develop fermentation methodology for the production of the biocontrol agent Heterorhabditis bacteriophora. Deployment of this organism will reduce the use of chemical insecticides which threaten the environment. This study shows how to produce the entomopathogenic nematode (EPN) Heterorhabditis bacteriophora and its bacterial symbiont Photorhabdus luminescens utilizing an in vitro, monoxenic liquid culture. EPNs were cultured in three different bioreactor working volumes of 1.5, 4 and 7 liters with initial nematode inoculation concentrations of approximately 2x103/mL. Liquid nematode media was conditioned with the bacterial symbiont 24 hours prior to nematode inoculation. Within three days after inoculation, infective juveniles (IJs) developed into self-fertilizing hermaphrodites and eventually produced IJ offspring. Maximum nematode densities were obtained seven days post-nematode inoculation. All three working volumes (1.5, 4 and 7 liters) produced final yields of 4.6x104 ± 2000 IJs/mL, 4.2x104 ± 2200 IJs/mL and 3.9x104 ± 2000 IJs/mL, respectively. In vitro scale-up technology can be further optimized for production of this biocontrol agent by improving media formulation, process parameters, bioreactor design and inoculation times that will maximize nematode yield.},
 year = {2015}
}

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TY  - JOUR
T1  - Lab-scale in vitro Mass Production of the Entomopathogenic Nematode Heterorhabditis bacteriophora Using Liquid Culture Fermentation Technology
AU  - Devang Upadhyay
AU  - Sivanadane Mandjiny
AU  - Rebecca Bullard-Dillard
AU  - Meredith Storms
AU  - Michael Menefee
AU  - Leonard D. Holmes
Y1  - 2015/12/30
PY  - 2015
N1  - https://doi.org/10.11648/j.bio.20150306.19
DO  - 10.11648/j.bio.20150306.19
T2  - American Journal of Bioscience and Bioengineering
JF  - American Journal of Bioscience and Bioengineering
JO  - American Journal of Bioscience and Bioengineering
SP  - 203
EP  - 207
PB  - Science Publishing Group
SN  - 2328-5893
UR  - https://doi.org/10.11648/j.bio.20150306.19
AB  - The objective of this research is to develop fermentation methodology for the production of the biocontrol agent Heterorhabditis bacteriophora. Deployment of this organism will reduce the use of chemical insecticides which threaten the environment. This study shows how to produce the entomopathogenic nematode (EPN) Heterorhabditis bacteriophora and its bacterial symbiont Photorhabdus luminescens utilizing an in vitro, monoxenic liquid culture. EPNs were cultured in three different bioreactor working volumes of 1.5, 4 and 7 liters with initial nematode inoculation concentrations of approximately 2x103/mL. Liquid nematode media was conditioned with the bacterial symbiont 24 hours prior to nematode inoculation. Within three days after inoculation, infective juveniles (IJs) developed into self-fertilizing hermaphrodites and eventually produced IJ offspring. Maximum nematode densities were obtained seven days post-nematode inoculation. All three working volumes (1.5, 4 and 7 liters) produced final yields of 4.6x104 ± 2000 IJs/mL, 4.2x104 ± 2200 IJs/mL and 3.9x104 ± 2000 IJs/mL, respectively. In vitro scale-up technology can be further optimized for production of this biocontrol agent by improving media formulation, process parameters, bioreactor design and inoculation times that will maximize nematode yield.
VL  - 3
IS  - 6
ER  -

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