Characterization of Photorhabdus luminescens Growth for the Rearing of the Beneficial Nematode Heterorhabditis bacteriophora

Abstract

Culturing the bioluminescent bacterium Photorhabdus luminescens in nutrient broth (NB) is used to recover phase I cells. These phase I cells were highly luminescent for up to 7 h in this media and the luminosity could also be seen with the naked eye after a 15 min eye adjustment period in a dark room. Red pigmentation is a known trait of phase I cells and was visually distinct within the culture media. The color shade of the red pigment varied on nutrient agar and in NB suggesting that the concentration of the pigment produced is dependent upon density of phase I cells within a specified area. The specific growth rate (l) and doubling time (g) was determined during the logarithmic growth phase to be 0.36 h -1 and 2.1 h, respectively in NB medium. The nematode-bacterium suspension was injected into larvae of Galleria mellonella to test for entomopathogencity. Within 24 h post-injection insect mortality was seen along with dark red pigmentation and extremely high luminosity indicating infection with P. luminescens.

Susceptibility of olive fruit fly, Bactrocera oleae (Diptera: Tephritidae) pupae to entomopathogenic nematodes

“…The ones infected by H. bacteriophora developed a typical red colour due to the pigment produced by the enteric bacteria Photorhabdus luminescens, while the pupae with S. carpocapsae maintained a light-brown coloration (Forst and Nealson 1996;Singh et al 2012) (Fig. 1A and B).…”

Determination of specific oxygen uptake rate ofPhotorhabdus luminescensduring submerged culture in lab scale bioreactor

“…Microorganism and inoculum P. luminescens, isolated earlier and maintained as explained by Singh, Moreau, Inman, and Holmes (2012), was cultured in a liquid medium containing per litre of tap water (tH 2 O): 8 g nutrient broth (Carolina Biological Supply Company, USA), 20 g trehalose (Carolina Biological Supply Company, USA) adjusted to pH 7.2 with 2 N NaOH. The bacterial culture was cultivated in a baffled shake flask shaken at 200 rpm for 36 h at 28°C in an incubated shaker and used as the bioreactor inoculum.…”

“…The two distinct forms identified as phase I and phase II differ from each other with respect to bioluminescence, pigmentation/antibiotic production, secretion of extracellular enzymes and the formation of crystalline inclusion bodies (Wang, Bilgrami, Shapiro-Ilan, & Gaugler, 2007). In vivo and in vitro luminescence of the phase I form of P. luminescens is much greater (∼300 times higher) than that of the phase II variant Owuama, 2001;Singh et al, 2012). The occurrence of phase II variants is problematic for improving the nematode culturing process, since nematode development and reproduction requires the phase I variant of P. luminescens.…”

Mass Production of the Beneficial Nematode Heterorhabditis bacteriophora and Its Bacterial Symbiont Photorhabdus luminescens

“…After inoculation, P. luminescens is grown for 24 h at 28°C, with an air flow rate of 1 vvm and agitation of 100 rpm [64]. The medium for the co-culture is much more complex than bacterial media alone [65]. Published formulations contain varying concentrations of the following: (i) peptone; (ii) yeast extract; (iii) beef extract; (iv) lipid; (v) carbohydrate; (vi) cholesterol; and (vii) salts.…”