Genetically Engineered Erwinia carotovora in Aquatic Microcosms: Survival and Effects on Functional Groups of Indigenous Bacteria

Abstract

The survival of genetically engineered Erwinia carotovora L-864, with a kanamycin resistance gene inserted in its chromosome, was monitored in the water and sediment of aquatic microcosms. The density of genetically engineered and wild-type E. carotovora strains declined at the same rate, falling in 32 days below the level of detection by viable counts. We examined the impact of the addition of genetically engineered and wild-type strains on indigenous bacteria belonging to specific functional groups important in nutrient cycling. For up to 16 days, the densities of total and proteolytic bacteria were significantly higher (P < 0.05) in microcosms inoculated with genetically engineered or wild-type E. carotovora, but by 32 days after inoculation, they had decreased to densities similar to those in control microcosms. Inoculation of genetically engineered or wild-type E. carotovora had no apparent effect on the density of amylolytic and pectolytic bacteria in water and sediment. Genetically engineered and wild-type E. carotovora did not have significantly different effects on the densities of specific functional groups of indigenous bacteria (P > 0.05).

Survival and Ecological Effects of Genetically Engineered Erwinia Carotovora in Soil and Aquatic Microcosms

“…Genetically engineered microcosm survival, intraspecific competition, and effect on indigenous bacteria were studied in aquatic microcosms constructed in 850-ml glass jars using 300 ml of sediment and 550 ml of water [9]. Microcosms were inoculated with 10 7 CFU/ml of genetically engineered and/or wildtype E. carotovora and incubated at 20ЊC on a 12-h light cycle.…”

“…In these investigations, a battery of genetically engineered Erwinia carotovora subspecies carotovora strains were used as models in aquatic and soil microcosms. Erwinia carotovora subspecies carotovora is a phytopathogenic enterobacterium responsible for soft rot in many agricultural plant tissues [9]. Erwinia carotovora produces a battery of enzymes (pectate lyases, cellulases, phospholipases, and proteases) that are responsible for plant tissue breakdown [10].…”

“…Previous studies by Orvos and colleagues [14] and Scanferlato and colleagues [15] addressed the survival, competi- Water and sediment [9,14] L-872 GEM Deficient in a pI 10.4 endo-pectate lyase gene (Kan R ) Survival Soil [17] tion, and effects on indigenous bacterial communities of genetically engineered E. carotovora in a single loam soil type. The present study is an extension of that work and expands upon the findings of GEM survival, competition, and effects on indigenous organisms while taking into account the extent of genetic manipulation of the GEM strain.…”

“…Genetically engineered E. carotovora L-833 was used in the aquatic microcosms and was constructed from the wildtype by partially removing in vitro an extracellular pectate lyase gene on a plasmid (pDR40) by deletion mutation, inserting a 1.4-kb DNA fragment from Tn903 conferring kanamycin resistance, and chromosomally inserting, by gene replacement, part of the plasmid (pSV41) into the chromosome [9,16]. This strain, L-833, has a 30% reduction in its ability to rot potato tuber tissue compared to the wildtype [9]. Strain L-864, used in the aquatic microcosms, is a spontaneous rifampin (150 g/ml)resistant mutant of L-833 that is also kanamycin resistant [9,14].…”

“…This strain, L-833, has a 30% reduction in its ability to rot potato tuber tissue compared to the wildtype [9]. Strain L-864, used in the aquatic microcosms, is a spontaneous rifampin (150 g/ml)resistant mutant of L-833 that is also kanamycin resistant [9,14].…”