CARBON SOURCE UTILIZATION OF SOIL EXTRACTED MICROORGANISMS AS A TOOL TO DETECT THE EFFECTS OF SOIL SUPPLEMENTED WITH GENETICALLY-ENGINEEREDAND NON-ENGINEERED CORYNEBACTERIUM-GLUTAMICUM AND A RECOMBINANT PEPTIDE AT THE COMMUNITY-LEVEL
W. Vahjen et al., CARBON SOURCE UTILIZATION OF SOIL EXTRACTED MICROORGANISMS AS A TOOL TO DETECT THE EFFECTS OF SOIL SUPPLEMENTED WITH GENETICALLY-ENGINEEREDAND NON-ENGINEERED CORYNEBACTERIUM-GLUTAMICUM AND A RECOMBINANT PEPTIDE AT THE COMMUNITY-LEVEL, FEMS microbiology, ecology, 18(4), 1995, pp. 317-328
Substrate utilization of microbial cells extracted from soil with a 0.
85% aqueous sodium chloride solution, was determined to estimate effec
ts on soil microorganisms at the community level with microtiter plate
s (Biolog GN)) containing 95 different sources of organic carbon. A co
nsistent pattern of utilized substrates was obtained after 24 h of mic
rotiter plate incubation at 28 degrees C. The absorbance values (OD590
) obtained from a microtiter plate reader after background correction
were transformed by using the average absorbance values of oxidized su
bstrates as a threshold to distinguish between well utilized and poorl
y or non-utilized substrates and thereby reduce variances between repl
icates. Doubling times of the extracted soil microorganisms in the mic
rotiter plates were tested with 12 substrates and ranged from 1.96 h t
o 3.23 h, depending on the carbon source. The carbon source utilizatio
n assay was used to assess the effects of soil inoculation with Coryne
bacterium glutamicum with and without a genetically engineered plasmid
(pUN1: 6.3 kb), which encoded for the synthesis of the mammalian prot
ease inhibiting peptide, aprotinin. Additionally, aprotinin itself was
added at two concentrations to soil samples. An identical decrease in
the number of carbon sources utilized, especially carbohydrates,occur
red upon soil inoculation with both C. glutamicum strains after inocul
ation with 10(6) cells g(-1) soil. This effect was only detectable dur
ing the first three weeks of incubation, as long as cell numbers of C.
glutamicum (pUN1) were above 10(5) cfu g(-1). Soil amendment with apr
otinin resulted in utilization of additional substrates, most of them
carbohydrates. With 0.1 mg aprotinin g(-1) soil this stimulation laste
d 2 days and with 10 mg g(-1) it lasted for 7 days.