The significant contribution of fungi to soil N2O production across diverse ecosystems

Chen, Huaihai; Mothapo, Nape V.; Shi, Wei

doi:10.1016/j.apsoil.2013.08.011

Cited by 95 publications

(69 citation statements)

References 50 publications

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“…Microscopic observations corroborated bacterial growth on antibiotic-treated solid medium. Incomplete inhibition of all or specific microbial processes is a known weakness of SIRIN analyses and may explain the observed variation in N 2 O production between sites (13,17,64,65). The observed variability in NO 3 Ϫ reduction is consistent with other studies describing a diversity of fungi that are incapable of reducing NO 3 Ϫ to N 2 O (7, 10).…”

Section: Figsupporting

confidence: 79%

Detection and Diversity of Fungal Nitric Oxide Reductase Genes ( p450nor ) in Agricultural Soils

Higgins

Welsh

Orellana

et al. 2016

Appl Environ Microbiol

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Members of the Fungi convert nitrate (NO 3؊ ) and nitrite (NO 2 ؊ ) to gaseous nitrous oxide (N 2 O) (denitrification), but the fungal contributions to N loss from soil remain uncertain. Cultivation-based methodologies that include antibiotics to selectively assess fungal activities have limitations, and complementary molecular approaches to assign denitrification potential to fungi are desirable. Microcosms established with soils from two representative U.S. Midwest agricultural regions produced N 2 O from added NO 3 ؊ or NO 2 ؊ in the presence of antibiotics to inhibit bacteria. Cultivation efforts yielded 214 fungal isolates belonging to at least 15 distinct morphological groups, 151 of which produced N 2 O from NO 2 ؊ . Novel PCR primers targeting the p450nor gene, which encodes the nitric oxide (NO) reductase responsible for N 2 O production in fungi, yielded 26 novel p450nor amplicons from DNA of 37 isolates and 23 amplicons from environmental DNA obtained from two agricultural soils. The sequences shared 54 to 98% amino acid identity with reference P450nor sequences within the phylum Ascomycota and expand the known fungal P450nor sequence diversity. p450nor was detected in all fungal isolates that produced N 2 O from NO 2 ؊ , whereas nirK (encoding the NO-forming NO 2 ؊ reductase) was amplified in only 13 to 74% of the N 2 O-forming isolates using two separate nirK primer sets. Collectively, our findings demonstrate the value of p450nor-targeted PCR to complement existing approaches to assess the fungal contributions to denitrification and N 2 O formation. IMPORTANCEA comprehensive understanding of the microbiota controlling soil N loss and greenhouse gas (N 2 O) emissions is crucial for sustainable agricultural practices and addressing climate change concerns. We report the design and application of a novel PCR primer set targeting fungal p450nor, a biomarker for fungal N 2 O production, and demonstrate the utility of the new approach to assess fungal denitrification potential in fungal isolates and agricultural soils. These new PCR primers may find application in a variety of biomes to assess the fungal contributions to N loss and N 2 O emissions. Denitrification is a key process responsible for loss of fixed nitrogen (N) in soils and sediments and is mediated by both abiotic and microbial processes (1, 2). Of the microorganisms involved in denitrification, members of the Bacteria are well studied and considered key contributors; however, some saprotrophic fungi also conserve energy from the reduction of nitrate (NO 3 Ϫ ) or nitrite (NO 2 Ϫ ) to nitrous oxide (N 2 O), the main end product of fungal denitrification (3-5). Fungi have been implicated in N turnover for over 30 years (1,3,6), but the ecological importance of fungal contributions to denitrification remain uncertain. Potential roles of fungi in soil N loss and greenhouse gas (i.e., N 2 O) emission are underexplored, and monitoring tools (e.g., quantitative or endpoint PCR) to specifically address the presence and abundance of denitrif...

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Section: Figsupporting

confidence: 79%

Detection and Diversity of Fungal Nitric Oxide Reductase Genes ( p450nor ) in Agricultural Soils

Higgins

Welsh

Orellana

et al. 2016

Appl Environ Microbiol

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“…captan, cycloheximide, ketoconazole, benomyl and nystatin) have been employed in past studies 13,17,22,23,24 , including studies investigating denitrification in soils and sediments. 13,25,26,27,28 In these studies, different types of biocides were used and at different application rates. Inhibition efficiency has been variable, and after analysis of published results, it has not been possible to identify the type and the optimum concentration of biocide that is most inhibitory to denitrification in agricultural soils.…”

Section: Introductionmentioning

confidence: 99%

Evaluation of antibacterial and antifungal compounds for selective inhibition of denitrification in soils

Ladan

Jacinthe

2016

Environ. Sci.: Processes Impacts

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Nitrous oxide (N2O) is an atmospheric constituent implicated in climate warming and stratospheric ozone depletion. Both bacteria and fungi participate in N2O production, but information is lacking with regard to the relative contribution of bacterial and fungal denitrifiers to the denitrification process in agricultural soils. The selective inhibition technique (SI) is widely used to assess the contribution of different groups of microbes to soil processes, but success of the technique depends on the effectiveness of the inhibitors.In this study, laboratory experiments were conducted to assess the contribution of bacteria and fungi to denitrification using soils from a woodlot, agricultural fields under conventional plowing (PT), and no-till for either 50 years (long-term) or 11 years (medium-term). A selective inhibition (SI) technique was developed using two bactericides (streptomycin, bronopol) and two fungicides (cycloheximide, captan) applied at different rates (0-32 mg g -1 soil). Regardless of application rate, streptomycin and cycloheximide were not effective inhibitors of denitrification, with degree of inhibition only between 2 and 20% relative to controls. These results are significant given the wide use of these products in SI studies. However, the bactericide bronopol and the fungicide captan effectively inhibited denitrification, with the strongest inhibition observed at an application rate of 16 mg g -1 soil. The ratio of fungal to bacterial denitrification activity (F:B) was generally less than 1, indicating a dominance of bacteria in denitrification activity in the soils investigated. However, an increase in F:B ratio from 0.24 in mediumterm NT to 0.87 in long-term NT soils was noted, suggesting perhaps a progressive increase in the role of fungal denitrifiers with longer duration of NT farming.3

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“…The fungal contribution to N 2 O production in soils is often measured using substrate-induced respiration-inhibition (SIRIN) in combination with cycloheximide, a fungal growth inhibitor, to measure the decrease in the flux of N 2 O in soil samples. These types of studies have shown that the addition of cycloheximide decreased N 2 O production by up to 89% in a perennial ryegrass field (6), 63% to 85% in semiarid soils (7,8), 81% in an arable peat soil (9), 40 to 51% in soil farming systems (10), and 18% in a sandy loam ley grass field (11). In some studies, the decrease in N 2 O flux when inhibiting fungi was greater than that when inhibiting bacteria, clearly demonstrating that fungal denitrification was more important than bacterial denitrification in certain ecosystems (6,8,9).…”

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confidence: 99%

Novel P450nor Gene Detection Assay Used To Characterize the Prevalence and Diversity of Soil Fungal Denitrifiers

Novinscak

Zebarth

Burton

et al. 2016

Appl Environ Microbiol

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Denitrifying fungi produce nitrous oxide (N 2 O), a potent greenhouse gas, as they generally lack the ability to convert N 2 O to dinitrogen. Contrary to the case for bacterial denitrifiers, the prevalence and diversity of denitrifying fungi found in the environment are not well characterized. In this study, denitrifying fungi were isolated from various soil ecosystems, and novel PCR primers targeting the P450nor gene, encoding the enzyme responsible for the conversion of nitric oxide to N 2 O, were developed, validated, and used to study the diversity of cultivable fungal denitrifiers. This PCR assay was also used to detect P450nor genes directly from environmental soil samples. Fungal denitrification capabilities were further validated using an N 2 O gas detection assay and a PCR assay targeting the nirK gene. A collection of 492 facultative anaerobic fungi was isolated from 15 soil ecosystems and taxonomically identified by sequencing the internal transcribed spacer sequence. Twenty-seven fungal denitrifiers belonging to 10 genera had the P450nor and the nirK genes and produced N 2 O from nitrite. N 2 O production is reported in strains not commonly known as denitrifiers, such as Byssochlamys nivea, Volutella ciliata, Chloridium spp., and Trichocladium spp. The prevalence of fungal denitrifiers did not follow a soil ecosystem distribution; however, a higher diversity was observed in compost and agricultural soils. The phylogenetic trees constructed using partial P450nor and nirK gene sequences revealed that both genes clustered taxonomically closely related strains together. IMPORTANCE A PCR assay targeting the P450nor gene involved in fungal denitrification was developed and validated. The newly developedP450nor primers were used on fungal DNA extracted from a collection of fungi isolated from various soil environments and on DNA directly extracted from soil. The results indicated that approximatively 25% of all isolated fungi possessed this gene and were able to convert nitrite to N 2 O. All soil samples from which denitrifying fungi were isolated also tested positive for the presence of P450nor. The P450nor gene detection assay was reliable in detecting a large diversity of fungal denitrifiers. Due to the lack of homology existing between P450nor and bacterial denitrification genes, it is expected that this assay will become a tool of choice for studying fungal denitrifiers.

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The significant contribution of fungi to soil N2O production across diverse ecosystems

Cited by 95 publications

References 50 publications

Detection and Diversity of Fungal Nitric Oxide Reductase Genes ( p450nor ) in Agricultural Soils

Detection and Diversity of Fungal Nitric Oxide Reductase Genes ( p450nor ) in Agricultural Soils

Evaluation of antibacterial and antifungal compounds for selective inhibition of denitrification in soils

Novel P450nor Gene Detection Assay Used To Characterize the Prevalence and Diversity of Soil Fungal Denitrifiers

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