phoD Alkaline Phosphatase Gene Diversity in Soil

Abstract

bPhosphatase enzymes are responsible for much of the recycling of organic phosphorus in soils. The PhoD alkaline phosphatase takes part in this process by hydrolyzing a range of organic phosphoesters. We analyzed the taxonomic and environmental distribution of phoD genes using whole-genome and metagenome databases. phoD alkaline phosphatase was found to be spread across 20 bacterial phyla and was ubiquitous in the environment, with the greatest abundance in soil. To study the great diversity of phoD, we developed a new set of primers which targets phoD genes in soil. The primer set was validated by 454 sequencing of six soils collected from two continents with different climates and soil properties and was compared to previously published primers. Up to 685 different phoD operational taxonomic units were found in each soil, which was 7 times higher than with previously published primers. The new primers amplified sequences belonging to 13 phyla, including 71 families. The most prevalent phoD genes identified in these soils were affiliated with the orders Actinomycetales (13 to 35%), Bacillales (1 to 29%), Gloeobacterales (1 to 18%), Rhizobiales (18 to 27%), and Pseudomonadales (0 to 22%). The primers also amplified phoD genes from additional orders, including Burkholderiales, Caulobacterales, Deinococcales, Planctomycetales, and Xanthomonadales, which represented the major differences in phoD composition between samples, highlighting the singularity of each community. Additionally, the phoD bacterial community structure was strongly related to soil pH, which varied between 4.2 and 6.8. These primers reveal the diversity of phoD in soil and represent a valuable tool for the study of phoD alkaline phosphatase in environmental samples. P hosphorus (P) is an essential macronutrient for all living cells (1). Despite its relative abundance in soils, P is one of the main limiting nutrients for terrestrial organisms (2). P is present in organic and inorganic forms in soil, but only the inorganic orthophosphate ions in soil solutions are readily available for plants (3). To sustain crop productivity, large amounts of P fertilizers are therefore used in agriculture, both as inorganic fertilizers (e.g., triple super phosphate) and organic fertilizers (e.g., manure). After application, some of the inorganic P is rapidly taken up by plants and microorganisms, while the remaining P is immobilized as insoluble and bound P forms in the soil. Microorganisms can access and recycle P from these recalcitrant P forms by solubilization of inorganic P and by mineralization of organic P via enzymatic processes mediated primarily by phosphatases, which hydrolyze the orthophosphate group from organic compounds (3). When facing P scarcity, microorganisms upregulate expression of functional genes coding for phosphatases (phosphomonoesterases, phosphodiesterases, phytases), high-affinity phosphate transporters, and enzymes for phosphonate utilization, which together constitute the Pho regulon (4). The phosphomonoesters which are hydrolyzed ...

Quantification of bacterial non-specific acid ( phoC) and alkaline ( phoD ) phosphatase genes in bulk and rhizosphere soil from organically managed soybean fields

“…This is likely a result of the potential phosphatase assays being conducted at pH 6.5, rather than pH 4.0 for acid phosphatase and pH 11.0 for alkaline phosphatase. In contrast to our results, Ragot et al (2015) reported pH to be the main variable influencing the phoD bacterial community, although they had a wider range of pH from 4.2 to 6.8 and a greater variation in soil texture across the six sites included in their study.…”

“…For example, Ragot et al (2015) found between one and nine phoD gene copies per species, with 71% of the genomes containing only one copy. It is unclear how many phoC genes may be present in different bacterial genomes.…”

“…Acid (ACP, EC 3.1.3.2) and alkaline phosphatases (ALK, EC 3.1.3.1) are non-specific phosphohydrolases capable of hydrolysing simple phosphate monoesters to acquire orthophosphate (Nannipieri et al, 2011;Ragot et al, 2015) and are often described by their optimal pH activity (Eivazi and Tabatabai, 1977). Phosphatase activity is typically highest in the rhizosphere (defined as the soil surrounding a plant root), with higher levels of microbial activity compared to bulk soil (Hinsinger, 2001;Marschner et al, 2004).…”

Land‐use influences phosphatase gene microdiversity in soils

“…) indicates that phosphomonoesterases are more abundant in soil than phytases: phoD in particular is much more abundant than the other phosphatase genes. This is consistent with analysis of phoA , phoD and phoX abundance in publically available bacterial genomes and metagenome datasets (Tan et al ., ; Ragot et al ., ; Ragot et al ., ). In the case of the phosphomonoesterases (where grassland soil contained the greatest abundance), phoD represented 52%GE compared with 3%GE and 8%GE for phoA and phoX respectively.…”

A Keystone Methylobacterium Strain in Biofilm Formation in Drinking Water

“…To test the primers, the Standard MyTaq Mix Protocol (Bioline) [59] was used with 0.5 µL of 10 µM of each primer, 0.2 µL of 1 U Bioline MyTaq TM DNA polymerase (BIO-21106), 5 µL of 5X MyTaq TM Buffer (containing 5 mM deoxynucleoside triphosphates (dNTPs) and 15 mM magnesium chloride (MgCl 2 )), and 1 µL of 4.6 ng/µL template DNA in a final volume of 20 µL. This template DNA was derived from a sample from the pure culture with OD quantified at 0.32 and DNA quantified at 46 ng/µL.…”