The Effect of Heavy Metals on the Population Structure of R. leguminosarum bv. viciae in Sewage Sludge Contaminated Soils

Abstract

The disposal of sewage sludge on agricultural lands for use as a source of nutrients appears to be convenient to both city authorities and farmers. Unfortunately, sludges contain appreciable amount of heavy metals which remain in the cultivated layer for a very long time. This constitutes a major source of heavy metal contamination in these soils. The purpose of this work was to determine changes in the population size and structure of R. leguminosarum bv. viciae in two soils, Old arable soils (pH 6–7) and Old woodland soils (pH 5.1–6), from a long term sewage sludge experiment at Braunschweig, Germany, across a gradient of heavy metal concentrations to find whether the population size and structure of R. leguminosarum bv. viciae has changed due to long-term sewage sludge application. The most probable number (MPN) plant infection method was employed for estimating soil rhizobial population size. Plasmid profiles and PCR/RFLP analysis of the highly variable intergenic spacer region (IGS) which separates 16S and 23S ribosomal rRNA genes were used to characterize the indigenous population structure R. leguminosarum bv. viciae. The results showed that the number of R. leguminosarum bv. viciae decreased with increasing Zn concentration in both the Old arable and the Old-woodland soils. Reduction in the population size in the Old woodland soil was greater than that of the Old arable soil. Changes in the population structure of rhizobia based on plasmid profiles showed that: 1) Rhizobial population structure in the sewage sludge treatment plots were quite different to those in the control plots. 2) The least number of plasmid types and the greatest mean number of plasmid bands per isolate (complexity of plasmid profiles) was found in the most contaminated plots. 3) Diversity of R. leguminosarum bv. viciae in both soils increased at low levels of sewage sludge application but diversity was reduced with total Zn concentration above 100 mg kg−1 in the Old arable and above 170 mg kg−1 in the Old woodland soils and least diversity in both soils was detected in the most contaminated plots.