Nitric oxide reductase-targeted real-time PCR quantification of denitrifier populations in soil.

The quantification of denitrifying bacteria is a component in the further understanding of denitrification processes in the environment. Real-time PCR primers were designed to target two segments of the denitrifier population (cnorB(P) [Pseudomonas mandelii and closely related strains] and cnorB(B) [Bosea, Bradyrhizobium, and Ensifer spp.]) in agricultural soils based on functional cnorB (nitric oxide reductase) gene sequences. Total population numbers were measured using 16S rRNA gene real-time PCR. Two soil microcosm experiments were conducted. Experiment 1 examined the response of the indigenous soil microbial population to the addition of 500 mg/kg glucose-C daily over 7 days in soil microcosms. Changes in the total population were correlated (r = 0.83) between 16S rRNA gene copy numbers and microbial biomass carbon estimates. Members of the cnorB(P) population of denitrifiers showed typical r-strategy by being able to increase their proportion in the total population from starting levels of <0.1% to around 2.4% after a daily addition of 500 mg/kg glucose-C. The cnorB(B) guild was not able to increase its relative percentage of the total population in response to the addition of glucose-C, instead increasing copy numbers only in proportion with the total population measured by 16S rRNA genes. Experiment 2 measured population dynamics in soil after the addition of various amounts of glucose-C (0 to 500 mg/kg) and incubation under denitrifying conditions. cnorB(P) populations increased proportionally with the amount of glucose-C added (from 0 to 500 mg/kg). In soil microcosms, denitrification rates, respiration, and cnorB(P) population densities increased significantly with increasing rates of glucose addition. cnorB(B) guild densities did not increase significantly under denitrifying conditions in response to increasing C additions.

Analysis of denitrification genes and comparison of nosZ, cnorB and 16S rDNA from culturable denitrifying bacteria in potato cropping systems.

Bacterial denitrification in agricultural soils is a major source of nitrous oxide, a potent greenhouse gas. This study examined the culturable bacterial population of denitrifiers in arable field soils in potato (Solanum tuberosum L.) production and denitrification genes (nir, nor and nos) and 16S rDNA in those isolates. Enrichments for culturable denitrifiers yielded 31 diverse isolates that were then analysed for denitrification genes. The nitrous oxide reductase (nosZ) gene was found in all isolates. The majority of isolates ( approximately 90%) contained the cnorB nitric oxide reductase gene, with the remainder containing the qnorB gene. Nitrite reductase genes (nirS and nirK) were amplifiable from most of the isolates, and were segregated between species similar to previously isolated denitrifiers. Isolated strains were preliminarily identified using fatty acid methyl ester analysis and further identified using 16S rDNA sequencing. The majority of isolates (21) were classified as Pseudomonas sp., with smaller groups of isolates being most similar to Bosea spp. (4), Achromobacter spp. (4) and two isolates closely related to Sinorhizobium/Ensifer spp. Phylogenetic trees were compared among nosZ, cnorB and 16S rDNA genes for a subset of Pseudomonas strains. The trees were mostly congruent, but some Pseudomonas sp. isolates grouped differently depending on the gene analysed, indicating potential horizontal gene transfer of denitrification genes. Although Bosea spp. are known denitrifiers, to the best of our knowledge this is the first report of isolation and sequencing of denitrification genes from this bacterial genus.

Remobilization and uptake of N by newly planted apple (Malus domestica) trees in response to irrigation method and timing of N application.

Environmentally sound management of N in apple orchards requires that N supply meets demand. In 1997, newly planted apple trees (Malus domestica Borkh. var. Golden Delicious on M.9 rootstock) received daily applications of N for six weeks as Ca(15NO3)(2) through a drip irrigation system at a concentration of 112 mg l(-1) at 2-8, 5-11 or 8-14 weeks after planting. Irrigation water was applied either to meet estimated evaporative demand or at a fixed rate. In 1997, trees were harvested at 5, 8, 11 and 14 weeks after planting; and in 1998 at 3 weeks after full bloom. The amount of fertilizer N recovered was similar in trees in both irrigation treatments, but efficiency of fertilizer use was greater for trees receiving demand-controlled irrigation than fixed-rate irrigation. This was attributed to lower N inputs, greater retention time in the root zone and less N leaching in the demand-controlled irrigation treatments compared with fixed-rate irrigation treatments. Less fertilizer N was recovered by trees receiving an early application of N than a later application of N and this was related to the timing of N supply with respect to tree demand. Demand for root-supplied N was low until 11 weeks after planting, because early shoot and root growth was supported by N remobilized from woody tissue, which involved 55% of the total tree N content at planting. Rapid development of roots > 1 mm in diameter occurred between 11 and 14 weeks after planting, after remobilization ended, and was greater for trees receiving an early application of N than for trees receiving a later application of N. Late-season tree N demand was supplied by native soil N, and uptake and background soil solution N concentrations were higher for trees receiving demand-supplied irrigation compared with fixed-rate irrigation. Total annual N uptake by roots was unaffected by treatments and averaged 6-8 g tree(-1). Nitrogen applications in 1997 affected growth and N partitioning in 1998. Trees receiving early applications of N had more flowers, spur leaves and bourse shoots than trees receiving later applications of N. Consequently, more N was remobilized into fruits in trees receiving early applications of N compared with fruits in trees receiving later applications of N. Demand for N in the young apple trees was low. Early season demand was met by remobilization from woody tissues and the timing of demand for root-supplied N probably depends on whether flowering occurs. Method of N delivery affected the efficiency of N use. We conclude that N demand can be met at soil solution N concentrations of around 20 mg l(-1).

Impact and shear resistance of turf grass racing surfaces for Thoroughbreds.

The 2 dominant processes involved in the horse hoof-to-racing surface interaction are the shock loading of the horse's leg upon impact with the racing surface and rotation of the horse's hoof into the racing surface. These processes were measured as impact resistance (ie, the peak deceleration of a moving body upon impact with the test surface) and as resistance to shear. The objective of the present study was to measure physically (under a variety of conditions) those soil and turf factors related to the 2 processes. It was concluded that thatch accumulation and mowing height of the turf did not have a significant (P less than 0.05) effect on racing surface hardness (therefore, turf management should optimize turf growth and recovery), that turf roots were responsible for an increase in impact resistance and in resistance to shear, that control over soil moisture through irrigation and drainage allowed modification of racing surface hardness, and that soil materials tend to have lower impact resistance (ie, lower shock loading of the horse's leg) and higher resistance to shear (ie, greater resistance to hoof rotation) than do sand materials.