Analysis of bacterial communities on alkaline phosphatase genes in soil supplied with organic matter

Abstract

We studied the effects of the application of organic matter (OM) and chemical fertilizer (CF) on soil alkaline phosphatase (ALP) activity and ALP‐harboring bacterial communities in the rhizosphere and bulk soil in an experimental lettuce field in Hokkaido, Japan. The ALP activity was higher in soils with OM than in soils with CF, and activity was higher in the rhizosphere for OM than in the bulk soil. Biomass P and available P in the soil were positively related to the ALP activity of the soil. As a result, the P concentration of lettuce was higher in OM soil than in CF soil. We analyzed the ALP‐harboring bacterial communities using polymerase chain reaction based denaturing gradient gel electrophoresis (DGGE) on the ALP genes. Numerous ALP genes were detected in the DGGE profile, regardless of sampling time, fertilizer treatment or sampled soil area, which indicated a large diversity in ALP‐harboring bacteria in the soil. Several ALP gene fragments were closely related to the ALP genes of Mesorhizobium loti and Pseudomonas fluorescens. The community structures of the ALP‐harboring bacteria were assessed using principal component analysis of the DGGE profiles. Fertilizer treatment and sampled soil area significantly affected the community structures of ALP‐harboring bacteria. As the DGGE bands contributing to the principal component were different from sampling time, it is suggested that the major bacteria harboring the ALP gene shifted. Furthermore, there was, in part, a significant correlation between ALP activity and the community structure of the ALP‐harboring bacteria. These results raise the possibility that different ALP‐harboring bacteria release different amounts and/or activity of ALP, and that the structure of ALP‐harboring bacterial communities may play a major role in determining overall soil ALP activity.

phoD Alkaline Phosphatase Gene Diversity in Soil

“…For comparative analysis of PHOD and ALPS primers ALPS-F730/ALPS-R110 (5=-CA GTGGGACGACCACGAGGT-3=/5=-GAGGCCGATCGGCATGTCG-3=) (11), phoD genes were amplified in pooled duplicate DNA extracts at a concentration of 20 ng l Ϫ1 using the PCR conditions described above, with an annealing temperature at 58°C for PHOD primers and at 57°C for ALPS primers. Samples were then sequenced using 454 GS-FLXϩ pyrosequencing (Roche) by Research and Testing Laboratory, with a resulting yield between 1,642 and 13,998 reads per library.…”

“…They were based on phosphatase gene sequences from seven isolates and first used to examine the different soil alkaline phosphatase community structures resulting from mineral and organic fertilization. Alkaline phosphatase genes belonging to the Actinobacteria, Alpha-, Beta-, and Gammaproteobacteria, and Cyanobacteria classes were identified by clon-ing, giving the first insight into alkaline phosphatase diversity in soil (11).…”

“…Advances in culture-independent techniques have provided new tools for the study of microbial communities in the environment. The first functional gene probes to target alkaline phosphatase genes were the primers developed by Sakurai et al named ALPS primers (11). They were based on phosphatase gene sequences from seven isolates and first used to examine the different soil alkaline phosphatase community structures resulting from mineral and organic fertilization.…”

Bacterial alkaline phosphomonoesterase in the rhizospheres of plants grown in Chilean extreme environments

“…The abundance and the compositions of APase-harboring bacteria have been related to P availability in soils fertilized with manure and P inorganic fertilizers (Chhabra et al 2013;Fraser et al 2015a;Sakurai et al 2008;Tan et al 2013). However, little is known of APase-harboring bacterial assemblages in pristine soils.…”

“…P-limiting conditions, particularly in alkaline soils (Nannipieri et al 2011;Sharma et al 2005;Turner 2010). Three homologous genes encoding APase in prokaryotes have been identified: phoD (Sakurai et al 2008), phoA (Zappa et al 2001), and phoX (Sebastian and Ammerman 2009). The phoD gene has been used as a molecular marker to provide information regarding the distribution and diversity of APase bacterial assemblages in soils (Tan et al 2013).…”