ELSEVIER FEMS Microbiology Ecology I8 ( 1995) 3 17-328 Carbon source utilization of soil extracted microorganisms as a tool to detect the effects of soil supplemented with genetically engineered and non-engineered zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA Corynebacterium glutamicwm and a recombinant peptide at the community level Wilfried Vahjen, Jean-Charles Munch, Christoph C. Tebbe * Received 21 April 1995; revised 10 August 1995; accepted 8 September 1995 zyxwvutsrqponmlkjihgfedcbaZYX Abstract Substrate utilization of microbial cells extracted from soil with a 0.85% aqueous sodium chloride solution, was determined to estimate effects on soil microorganisms at the community level with microtiter plates (Biolog GN”) containing 95 different sources of organic carbon. A consistent pattern of utilized substrates was obtained after 24 h of microtiter plate incubation at 28°C. The absorbance values COD,,,) obtained from a microtiter plate reader after background correction were transformed by using the average absorbance values of oxidized substrates as a threshold to distinguish between well utilized and poorly or non-utilized substrates and thereby reduce variances between replicates. Doubling times of the extracted soil microorganisms in the microtiter plates were tested with 12 substrates and ranged from 1.96 h to 3.23 h, depending on the carbon source. The carbon source utilization assay was used to assess the effects of soil inoculation with zyxwvuts Cnqnehucteriwn glutumicum with and without a genetically engineered plasmid (pUN1; 6.3 kb), which encoded for the synthesis of the mammalian protease 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, occurred upon soil inoculation with both C. glutumicum strains after inoculation with IO6 cells g-’ soil. This effect was only detectable during the first three weeks of incubation, as long as cell numbers of C. glutamicum (pUNI) were above 10s cfu g-1. Soil amendment with aprotinin resulted in utilization of additional substrates. most of them carbohydrates. With 0.1 mg aprotinin g- ’ soil this stimulation lasted 2 days and with 10 mg g-’ it lasted for 7 days. Kewwds: Soil microbial community: Carbon source utilization; Aprotinin; Genetically engineered microorganism: Risk assessment 1. Introduction isms is their establishment as members of natural One major concern associated with the environ- mental release of genetically engineered microorgan- microbial communities. By this process important functional groups may be displaced and as a conse- quence alterations in biodegradation and nutrient cy- I Corresponding author. Tel: f49 (53 1) 596 736; Fax: +49 (53 I) 596 375: E-mail: tebbe@bb.fal.de cling could occur [l-3]. To obtain information on such potential alterations, the analysis of the micro- bial community structure and the detection of changes 0168-6496/95/$09.50 0 1995 Federation of European Microbiological Societies. All rights reserved SSDI 0168-6496(95)00072-O Downloaded from https://academic.oup.com/femsec/article-abstract/18/4/317/503686 by guest on 31 May 2020