Effect of compost, nitrogen salts, and NPK fertilizers on methane oxidation potential at different temperatures

ENVIRONMENTAL BIOTECHNOLOGY

Effect of compost, nitrogen salts, and NPK fertilizerson methane oxidation potential at different temperatures

Louis-B. Jugnia & Yaseen Mottiar & Euphrasie Djuikom &

Alexandre R. Cabral & Charles W. Greer

Received: 17 March 2011 /Revised: 9 August 2011 /Accepted: 18 August 2011 /Published online: 6 September 2011# Springer-Verlag 2011

Abstract The effects of compost, nitrogen salts, andnitrogen–phosphorous–potassium (NPK) fertilizers on themethane oxidation potential (MOP) of landfill cover soil atvarious temperatures were assessed. For this, we used batchassays conducted at 5°C, 15°C, and 25°C with microcosmscontaining landfill cover soil slurries amended with theseelements. Results indicated variable impacts dependent onthe type of amendment and the incubation temperature. Fora given incubation temperature, MOP varied from onecompost to another and with the amount of compost added,except for the shrimp/peat compost. With this lattercompost, independent of the amount, MOP values remainedsimilar and were significantly higher than those obtainedwith other composts. Amendment with most of the testednitrogen salts led to similar improvements in methanotro-phic activity, except for urea. MOP with NPK fertilizeraddition was amongst the highest in this study; theminimum value obtained with NPK (20–0–20) suggestedthe importance of P for methanotrophs. MOP generallyincreased with temperature, and nutrient limitation becameless important at higher temperatures. Overall, at each of

the three temperatures tested, MOP with NPK fertilizeramendments provided the best results and was comparableto those observed with the addition of the shrimp/peatcompost. The results of this study provide the first evidenceof the following: (1) compost addition to improve meth-anotrophic activity in a landfill cover soil should considerthe amount and type of compost used and (2) theimportance of using NPK fertilizers rather than nitrogensalts, in enhancing this activity, primarily at low temper-atures. One can also consider the potential beneficial impactof adding these elements to enhance plant growth, which isan advantage for MOP.

Keywords Methanotrophs . Potential activity . Compost .

Nitrogen salts . NPK fertilizers

Introduction

Methane (CH4) is a potent greenhouse gas that absorbsinfrared radiation more effectively than CO2 and has aglobal warming potential 25 times higher than that of CO2

(IPCC 2007). CH4 is the product of the anaerobicdegradation of organic matter by methanogenic Archaea(Zinder 1993). Landfills represent a significant source ofanthropogenic CH4 emissions contributing as much as 17%(Wuebbles and Hayhoe 2002) of the 70% of CH4 emissionsattributed to human activities (IPCC 2007).

Landfills are an important source of greenhouse gasesthat cause global climate change (IPCC 2007). Afterlandfill closure, vertical migration transports CH4, producedby methanogenic bacteria deep within the anaerobic regionof the landfill, to the aerobic environment near the surface,where it may be subjected to microbiological oxidation bymethane-oxidizing bacteria (methanotrophs) (Whalen et al.

L.-B. Jugnia (*) :Y. Mottiar : C. W. GreerNational Research Council Canada,Biotechnology Research Institute,6100 Royalmount Avenue,Montreal, QC, Canada H4P 2R2e-mail: [email protected]

E. DjuikomFaculty of Sciences, University of Douala,P.O. Box 24157, Douala, Cameroon

A. R. CabralDepartment of Civil Engineering, Université de Sherbrooke,2500 boulevard de l’Université,Sherbrooke, QC, Canada J1K 2R1

Appl Microbiol Biotechnol (2012) 93:2633–2643DOI 10.1007/s00253-011-3560-4

1990; Jones and Nedwell 1993; Humer and Lechner 2001).CH4 emissions from landfills can be mitigated by microbialCH4 oxidation in landfill cover soils (Boeckx et al. 1996;Hilger and Humer 2003; Kallistova et al. 2005; Cabral et al.2010)—known as biocovers.

Several previous investigations have assessed the role ofcompost as a biofilter in landfill cover soils (Humer andLechner 1999; Straska et al. 1999; Wilshusen et al. 2004;Cabral et al. 2010). Organic composts are thought topromote microbial growth by improving the porosity,water-holding capacity, pH, and nutrient supply (Hilgerand Humer 2003). Accordingly, there is an increased use ofcompost in biocovers to mitigate CH4 emissions fromlandfills, and recent investigations in this direction haveshown promising results (Humer and Lechner 2001; Barlazet al. 2004; Maurice and Lagerkvist 2004; Wilshusen et al.2004; Kettunen et al. 2006; Powelson et al. 2006; Jugnia etal. 2008; Aït-Benichou et al. 2009). Organic amendments tosoil generally result in increased enzymatic activity (Bandickand Dick 1999; Garcia-Gil et al. 2000; Perucci et al. 2000).Also, Saison et al. (2006) reported that compost amendmentaffects the prevalence and overall activity of the soilmicrobial community.

Soil-mediated CH4 oxidation provides a biological miti-gation strategy to reduce both atmospheric and in situ-produced CH4 (Amaral et al. 1998). Compost amendment tosoil is a common practice generally used to improve soilfertility and physical structure (Shiralipoura et al. 1992;Arthur et al. 2011). Numerous types of compost exist for thisuse, and the amount applied can vary from site to site.Previous studies have addressed the impact of compost onthe soil microbial community in general (Pérez-Piqueres etal. 2006; Saison et al. 2006) or methanotrophs specifically(Seghers et al. 2003). However, the specific impact ofcompost amendments on the methanotrophic capacity of soilis still not well understood and should be investigated insofaras, among other impacts, excessive use of organic amend-ment is likely to intensify non-methanotrophic microbialactivity and thereby limit the CH4 oxidation capacity of thesoil. Also, nitrogen salts and nitrogen–phosphorous–potassi-um (NPK) fertilizers are commonly used to improve theproductivity of soils, with potential impacts on the methano-trophic activity of these environments.

This study examined the combined effects of temper-ature and amendments of different types and amounts ofcompost on the CH4 oxidation potential (MOP) of alandfill cover soil. For this, samples from a landfill coversoil with a well-established community of methanotrophs(Aït-Benichou et al. 2009) were chosen. To our knowledge,no previous work has compared the rates of MOP in a landfillcover soil amended with different types and amounts ofcompost under various temperature regimes. Similarly, weexamined the effects of nitrogen salts and NPK fertilizers.

Understanding changes in methanotrophic activity induced bythese treatments is of economic and environmental impor-tance to optimize the mitigation of the net flux of CH4 fromlandfills.

Materials and methods

Soil and compost samples

Landfill cover soil was collected from the surface of anexisting landfill located at Saint-Nicephore, Quebec (Canada),and were sieved to 2 mm. This soil had been exposed tobiogas (containing ∼55% CH4) for about 4 years (Jugnia etal. 2008), and its particle size distribution was: 25.2% sand,67.3% silt, and 7.5% clay. The collected soil samples werestored at 4°C and processed within 1 week. Five differenttypes of compost (green waste, biosolid waste, pulp andpaper residue, manure, and shrimp/peat) were obtained froma local producer (Les Composts du Québec, Quebec) andsieved to 2 mm. Subsamples of compost were sterilized bygamma irradiation at MDS Nordion (Laval, Quebec). Allsamples were stored in the dark at 4°C until use. Physico-chemical properties of the landfill cover soil and the differenttypes of compost that were used are presented in Table 1.The moisture content was determined by gravimetric loss at105ºC overnight whereas the organic matter content wasmeasured by gravimetric loss upon ignition at 550ºC for 2hours. The other analyses were performed by an externalcertified laboratory (Bodycote Testing Group, Pointe-Claire,Quebec) following the Standard Method for the Evaluationof Water and Wastewater for pH (ref. S.M. 4500-H) and totalN (ref. S.M. 4500-NH3 B.D.H.) or according to the CEAEQstandard for NH4

+ (http://www.ceaeq.gouv.qc.ca/methodes/pdf/MA303N10.pdf), total P (http://www.ceaeq.gouv.qc.ca/methodes/pdf/MA200Met12.pdf), NO2–NO3, and PO4

3−

(http://www.ceaeq.gouv.qc.ca/methodes/pdf/MA300Ions13.pdf).

Soil slurries and CH4 oxidation potential

Microcosm experiments for the determination of MOP wereperformed with soil slurries in aerobic 120-mL glass serumbottles. Ten grams of sample was used, and three levels(5%, 15%, and 25% w/w) of compost amendment weretested. Microcosm slurries were prepared by mixing 10 g ofsoil (control) or compost/soil mixture (5%, 15%, and 25%w/w) with 10 mL of deionized water. For experiments withnitrogen salts and NPK fertilizers, stock solutions ofNH4Cl, (NH4)2SO4, NH4NO3, NaNO3, urea, and NPKfertilizers at rates of 20–20–20, 20–6–20, and 20–0–20were prepared so that the addition of 10 mL of either one ofthese solutions to 10 g of soil yielded 600 μg of total

2634 Appl Microbiol Biotechnol (2012) 93:2633–2643

nitrogen (60 mg/kg) added in the microcosm. In all cases,10 mL of liquid was enough to form a slurry and therebyeliminate variations in MOP caused by differences in soilmoisture.

The glass serum bottles were capped with Teflon™-lined rubber stoppers and sealed with aluminum crimps.Pure CH4 (5 mL) was added to each bottle (equivalent toapproximately 5% v/v) after withdrawal of an equalvolume of the gas phase. All treatments were prepared intriplicate for each incubation temperature (5°C, 10°C, or25°C, representing low, medium, and high temperatureswithin the wide range of temperature from 0°C to 30°C,generally reported for methanotrophic activity), andslurries were not shaken. The MOP was evaluated bymeasuring the headspace concentration of CH4 over thecourse of the incubation period. Measurements wereperformed by gas chromatography (SRI 8610 C gaschromatograph, SRI, Torrance, CA, USA) as described inRoy and Greer (2000). For gas determinations, 0.2 mL ofthe headspace gas was injected into the gas chromatographsystem equipped with a Porapack Q column, and peakswere integrated using the Peak Simple II software (SRI,Torrance, CA, USA).

Statistics

A linear regression through the linear portion of theplot of CH4 concentration vs. time describing the MOPwas obtained with measurements taken using repeatedsampling in the same triplicate microcosms throughoutthe incubation period. MOP values were determinedfrom the slope of the regression. For statistical analysis, ageneral linear model with unbalanced repeated measures(type III) analysis was applied using the PASW statistics18 package (formerly SPSS Statistics, SPSS Inc.). Thepredicted means from the general linear model werecompared using the least significant difference method

where a P value of less than 0.05 was considered to besignificant.

Results

Characteristics of the landfill cover soil and the differentcomposts

The results of the physicochemical analysis of the landfillcover soil and the different composts are presented inTable 1. The pH of the landfill cover soil was alkaline, andthe different composts were neutral or slightly alkaline.Nitrogen and phosphorus concentrations were higher in thecomposts than in the landfill cover soil. However, thebacteria-assimilable forms (NH4

+, NO2–NO3, and PO43−)

represented only a small fraction of the total concentrationavailable. Indeed, NH4

+ concentrations were generally lessthan 5 mg of N kg−1dry weight (DW) soil (i.e., 0–0.2% of totalnitrogen) and lower than the NO2–NO3 concentrations,representing 0.2–1.86% of total nitrogen, except for theshrimp/peat compost which contained NO2–NO3 as 4.5%of the total nitrogen. Also, PO4

3− concentrations were lessthan 5 mg of P kgDW soil

−1 except for the manure compostwhich contained 848 mg of P kgDW soil

−1 (i.e., 8.23% oftotal phosphorous). All composts contained similar levelsof organic matter, ranging from 20% to 26%, which was fargreater than that present in the landfill cover soil.

Effect of compost types and amount on MOP performanceat different temperatures

As shown in Fig. 1, of the different types of compostconsidered, CH4 consumption was fastest in soil amendedwith shrimp/peat compost irrespective of the amount added.The complete oxidation of CH4 in these microcosmsoccurred within 4 days at 25°C, between 7 and 12 days at

Table 1 Physicochemical properties of the landfill cover soil and the different composts

Parameters Landfill Composts

cover soil Green waste Biosolid waste Pulp and paper Manure Shrimp/peat

pH 8.6 7.7 7.6 7.6 7.8 7.0

Organic matter (%) 1.14 22.68 20.01 21.87 25.27 20.26

NH4+a <5 <5 <6 <5 <5 <5

NO2–NO3a 1.1 <0.8 24.1 184 <0.8 752

PO43−a <0.5 <4.3 <5.0 <2.5 848 <3.2

Total Na 241 11,400 10,700 9,890 11,800 15,700

Total Pa 595 1,780 6,100 5,080 10,300 9,810

a Units are in milligrams per kilogram of dry weight soil

Appl Microbiol Biotechnol (2012) 93:2633–2643 2635

15°C, and in less than 30 days at 5°C (Fig. 1). This was notthe case with the other composts for which the completeoxidation of CH4 required 6–8 days at 25°C, 21–30 days at15°C, and more than 30 days at 5°C (Fig. 1). Samplesamended with biosolid waste and manure compostexhibited activities that were somewhat faster and moreconsiderable at 5°C and 15°C with an increasing amount ofthese composts in soil samples (Fig. 1).

Overall, the concentration–time profiles of all theexperiments in Fig. 1 indicated an impact of the amountof compost on CH4 oxidation: at low temperatures, therewas an increasing variability among the different com-posts dependent on the amount of compost added.Samples amended with shrimp/peat compost exhibitedthe highest MOP, with average values of 0.7, 2.0, and5.7 μg CH4gDW soil

−1h−1 at 5°C, 10°C, and 25°C,respectively. When excluding these values, the lowestMOP, achieved with 5% compost, was between0.24 μg CH4gDW soil

−1h−1 at 5°C and 2.45 μg CH4gDWsoil

−1h−1 at 25°C (Fig. 2). These values increasedconsiderably with 25% compost and fluctuated between

0.32 μg CH4gDW soil−1h−1 at 5°C and 4.03 μg CH4gDW

soil−1h−1 at 25°C (Fig. 2).

Effect of nitrogen salts and NPK fertilizers on MOPperformance at different temperatures

CH4 consumption was also improved when soils wereamended with nitrogen salts or NPK fertilizers, althoughthe effect was less pronounced with urea and NPK (20–0–20). In soil samples amended with different nitrogensalts, the profiles of CH4 consumption over time wereclose to each other, and complete CH4 consumption wasachieved in less than 5 days at 25°C, between 63 and80 days at 15°C, and more than 80 days at 5°C (Fig. 3).NPK (20–20–20) and NPK (20–6–20) exhibited similarprofiles of CH4 consumption over time, with oxidationcompleted within 21–42 days at 15°C and 5°C, whileNPK (20–0–20) required more than 79 days for completeoxidation at 15°C and 5°C (Fig. 3). For nitrogen salts andfertilizers, the time elapsed for complete CH4 consump-tion increased with decreasing incubation temperature

Fig. 1 Profiles of CH4 consumption over time from microcosms amended with variable amounts of different composts and incubated at differenttemperatures. Triplicate samples were averaged, and error bars correspond to one standard deviation

2636 Appl Microbiol Biotechnol (2012) 93:2633–2643

Fig. 2 The MOP (± standard error) from microcosms amended with variable amounts of different composts. At a given temperature, differentletters indicate significant differences (P<0.05) between the different treatments

Appl Microbiol Biotechnol (2012) 93:2633–2643 2637

Fig. 3 Profiles of CH4 consumption over time from microcosms amended with different nitrogen salts and NPK fertilizers and incubated atdifferent temperatures. Triplicate samples were averaged, and error bars correspond to one standard deviation

2638 Appl Microbiol Biotechnol (2012) 93:2633–2643

(Fig. 3), a trend that was observed with compostaddition.

At a given temperature, changes in MOP were lesssignificant between the different nitrogen salts than theNPK fertilizers tested, but the differences became moresignificant with shifts in temperature, notably between 15°Cand 25°C (Fig. 4). MOP ranged from 0.20 to 0.70 μg CH4

gDW soil−1h−1 at 5°C, fluctuated between 0.31 and

0.97 μg CH4gDW soil−1h−1 at 15°C, and increased signifi-

cantly from 2.35 to 7.18 μg CH4gDW soil−1h−1 at 25°C

(Fig. 4). With the exception of urea, the MOP at 25°C wassimilar for NPK fertilizers and the nitrogen salts tested(Fig. 4). This was not the case at 5 and 15°C, since NPKamendment provided greater MOP than amendment withnitrogen salts. Indeed, the maximum value (0.40 μg CH4

gDW soil−1h−1) obtained with nitrogen salts was similar to the

minimum value (0.30 μg CH4gDW soil−1h−1) observed with

NPK (20–0–20) (Fig. 4).When comparing all the samples incubated at 25°C

with those containing compost amended at the 25%level, MOP values in soils amended with NPKfertilizers were the greatest and comparable to thoseobtained with the shrimp/peat compost. They werefollowed in decreasing order by the nitrogen salts andthe other types of compost. At 5°C and 15°C, the additionof NPK fertilizers provided the best results, followed bycomposts, which were better than the nitrogen salts.Also, for nearly all amendment types, MOP generallyincreased with increasing temperature, especially between15°C and 25°C (Figs. 3 and 4).

Discussion

The results indicated that CH4 oxidation in a landfill coversoil was improved by amendment with compost, nitrogensalts, and NPK fertilizers. Several studies have reportedsimilar findings (Hilger et al. 2000; Humer and Lechner2001; De Visscher and Van Cleemput 2003; Barlaz et al.2004; Maurice and Lagerkvist 2004; Aït-Benichou et al.2009; Albanna and Fernandes 2009; Lee et al. 2009).

The CH4 oxidation capacity of landfill cover soils isquite variable. Albanna and Fernandes (2009) reportedvalues of CH4 oxidation capacities of landfill cover soilsincubated at three different temperatures (5°C, 22°C, and35°C), ranging from 3.3 to 12.3 μg CH4gDW soil

−1h−1 witha clayey–silt soil and from 3.5 to 10.5 μg CH4gDW soil

−1h−1

with a clay soil. Using engineered soil with a mixture ofsewage sludge compost and de-inking waste amended withsand or bark chips incubated at 4–6°C, Kettunen et al.(2006) reported CH4 oxidation rates of 3.75 and2.45 μg CH4gDW soil

−1h−1, respectively. Einola et al.(2007) found CH4 consumption rates ranging between

0.48 and 3.68 μg CH4gDW soil−1h−1 for landfill cover soils

incubated between 1°C and 6°C. Compared to others, ourresults fall within the lower range of those previouslypublished for landfill cover soils, probably because weadded water in our microcosms to form a slurry andtherefore minimized variations caused by soil moisture.Whalen et al. (1990) and Czepiel et al. (1996) confirmedthat gas diffusion at soil saturation is limited by thediffusion coefficient of CH4 in water, which is four ordersof magnitude lower than in the air.

Effect of compost type and content on MOP performance

The MOP values from our experimental microcosmsamended with compost were always higher than thoseobserved in no amendment control microcosms thatcontained only the landfill cover soil. Since thecompost was sterile, this is an indication that compostaddition to this landfill cover soil provided somerequired missing abiotic factors, such as organic matterand nutrients that fuelled the activity of indigenous soilmethanotrophs. Previous studies have established aclear relationship between methanotrophic activity and/or diversity, and organic matter concentration (Christophersenet al. 2000; Bin et al. 2009). Our results agree with bothfield observations at landfills and results from laboratorystudies that have demonstrated that the MOP in mineralsoils can be enhanced by adding organic material such assewage sludge (Kightley et al. 1995) and compost (Humerand Lechner 2001; Barlaz et al. 2004; Maurice andLagerkvist 2004; Aït-Benichou et al. 2009). Also,Wilshusen et al. (2004) and Kettunen et al. (2006) reportedthat compost should be capable of enhancing the oxidationof CH4 at rates two to three times higher than those ofmineral soils.

The MOP observed within each group of samples treatedseparately with the same amounts of the different compostsunder study at a given incubation temperature varied fromone type of compost to another (Fig. 2). This suggestsdifferences between the type of compost, with the quantityand quality of the elements in the compost poweringmethanotrophic activity. The observed increase in MOPwith increasing amounts of compost, except with shrimp/peat compost, is evidence of this quantitative effect. Moonaet al. (2010) reached the same conclusion using increasingratios of earthworm cast to improve methanotrophic activityin a filter bed material consisting of a mixture of earthwormcast and rice paddy soil in a biocover. In contrast, resultswith shrimp/peat compost tend to highlight the differenceamong composts in terms of quality. With the shrimp/peatcompost and at each of the three temperatures examined,MOP values remained similar despite differences in theamount of this compost added.

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Fig. 4 The MOP (± standard error) from microcosms amended with different nitrogen salts and NPK fertilizers. At a given temperature, differentletters indicate significant differences (P<0.05) between the different treatments

2640 Appl Microbiol Biotechnol (2012) 93:2633–2643

The MOP values obtained with shrimp/peat compostwere generally higher than those observed with the othercomposts, possibly because it is a better source of essentialelements for methanotrophic activity. Small differences,principally at 5°C and 15°C, were detected betweencomposts with, in most cases, the highest MOP valuesattributed to manure and/or biosolid composts. As such,shrimp/peat compost followed by manure and biosolidcompost could be considered the most effective additives.However, since excessive use of organic amendments islikely to intensify non-methanotrophic microbial activityand thereby limit CH4 oxidation capacity, shrimp/peat couldbe considered the most effective compost. In this study, thereported differences between compost types in terms ofquality and quantity to improving methanotrophic activityare new information that have never been reported withlandfill cover soil, although previous studies have focusedon compost type.

At all three temperatures considered in this study, MOPvalues with the lowest amount of shrimp/peat compostprovided similar results to the highest amounts of othercomposts. The main physicochemical property of shrimp/peat compost that differed from the other composts was therelatively high concentration of bioavailable nitrogen, in theform of NO2–NO3. This prompted us to examine hownitrogen may impact MOP.

Effect of nitrogen salts and NPK fertilizers on MOPperformance

As the methanotrophic community developed, the mineralnitrogen content of the soil decreased (De Visscher et al.1999), and results of other studies reported an increase ofmethane oxidation rates possibly due to a relief of nitrogenlimitation (Hilger et al. 2000; De Visscher and Van Cleemput2003; Lee et al. 2009). We hypothesized that the form ofnitrogen may be relevant in this biological phenomenon, sowe tested a range of nitrogen salts and NPK fertilizers.

In general, our results indicated that there was a lack ofinhibition and even CH4 oxidation stimulation followingthe addition of various nitrogen salts. This is consistentwith results of previous studies (Bodelier et al. 2000; Hilgeret al. 2000; Sitaula et al. 2000; De Visscher et al. 2001; DeVisscher and Van Cleemput 2003), but contrasts with resultsfrom Kightley et al. (1995) reporting that NO3

− had aninhibitory effect on CH4 oxidation in landfill cover soils.De Visscher et al. (2001) observed that amending thelandfill cover soil with nitrifying sludge or compost initiallyinhibited CH4 oxidation, followed by a stimulation after afew days. In general, the reported information about CH4

oxidation in the presence of nitrogen salts is oftencontradictory (Hütsch et al. 1994; Dunfield et al. 1995;Willison et al. 1995; Delgado and Mosier 1996; Bodelier

and Laanbroek 2004). In the present study, most forms ofnitrogen tested were found to be equally suitable as similarchanges in methanotrophic activity were observed. Theprimary exception was urea, an NH4

+-producing com-pound, whose inhibitory effect on methanotrophs has beendemonstrated (Dubey and Singh 2000; Zheng et al. 2008).The mechanism responsible for this effect on CH4 uptakeis complex because it involves both methanotrophs andnitrifiers. Some have hypothesized that NH4

+ is acompetitive inhibitor of CH4 monoxygenase (Bédard andKnowles 1989).

At 5°C and 15°C, the maximum MOP values observedwith nitrogen salt amendment matched the lowest valuesobserved with NPK fertilizers, suggesting that NPKfertilizers were more efficient than nitrogen salts inimproving methanotrophic activity. Although increasedCH4 oxidation rates may be due to a relief from nitrogenlimitation, Zheng et al. (2008) reported that potassium (K)fertilizer may play an important role in maintainingmethanotrophic metabolic processes. Similarly, a fieldexperiment by Babu et al. (2006) suggested amendmentwith K to stimulate the activity of methanotrophic bacterialpopulations. On the other hand, the fact that the lowest CH4

uptake rates at 5°C and 15°C were obtained with NPK (20–0–20) amendment suggests the importance of phosphorousin the metabolism of the indigenous methanotrophiccommunity. Methanotrophs can rely on intracellular poly-phosphate (Trotsenko and Shishkina 1990), which, in ourcase, might have been accumulated during growth of themethanotroph community within the landfill cover soil.However, the importance of P appears to be less relevant at25°C, given that all forms of the NPK fertilizer additionstimulated CH4 uptake to similar extents. At high temper-atures, rather than nutrients limiting methanotrophic activ-ity, other complex processes that occur in the environmentat these temperatures (organic matter degradation) contributeto releasing of, amongst other components, appropriatenutrients that can sustain methanotrophic activity. This pointsto the role of temperature in enhancing methanotrophicactivity, as substantiated by the fact that differences observedat 5°C and 15°C following additions of NPK fertilizer and/ornitrogen salts were not noticeable at 25°C, except for urea.

Effect of temperature on MOP

Temperature has a direct impact on biological reactions.The positive response of the CH4 consumption rate toincreasing temperature was clear and consistent acrossamendment types and levels, in agreement with publishedobservations regarding temperature and methanotrophy(Czepiel et al. 1996; Christophersen et al. 2000; Gebert etal. 2003; Börjesson et al. 2004; Park et al. 2005; Jugnia et al.2006, 2008; Kettunen et al. 2006). This generally occurs

Appl Microbiol Biotechnol (2012) 93:2633–2643 2641

when the CH4 concentration, among other factors, is notlimiting (Pawlowska and Stepniewski 2006; Einola et al.2007). The rate of CH4 consumption by methanotrophs insoil is affected by the rate of substrate (CH4 and O2) supplyas well as the amount and activity of CH4 monooxygenasethat catalyzes the process. Accordingly, Boeckx and VanCleemput (1996) observed a low response of CH4 consump-tion to temperature (average Q10, 1.9) in landfill cover soilincubated at 10 μl L−1 CH4 in contrast to the higher Q10

values (1.9–7.26) in previous studies at >1% CH4 concen-trations (Czepiel et al. 1996; Christophersen et al. 2000; DeVisscher et al. 2001; Börjesson et al. 2004), as well as in thepresent study (Q10, 2.54–3.43). The Q10 values generallyincreased with temperature in this study with the exceptionof the shrimp compost. Our observation that at hightemperatures nutrients were no longer the major factorregulating methanotrophic activity is consistent with a recentreport by Albanna and Fernandes (2009) that the temperatureeffect on CH4 oxidation was so significant that it masked theeffects of moisture content and nutrient availability.

Another important finding of this study is our observationthat amendment with NPK fertilizers (containing [P] >0)improved CH4 oxidation to a greater extent than nitrogensalts, notably at 5°C and 15°C. To the best of our knowledge,such a comparison is something that has not been reported sofar for landfill cover soils. This new insight will beparticularly informative for landfill operators when consid-ering biocover design in cold climates. In addition, this hasimplications for landfill cover soil where it would be possibleto provide nutrients to promote growth and maintain thevegetation system. A good vegetation system minimizeserosion by stabilizing the surface of the cover, and itcontributes to aeration and moisture control of the soil.

Acknowledgments We gratefully acknowledge the financial supportby the National Research Council Canada through the BiotechnologyResearch Institute including a summer research studentship for Y.M.Also, E.D. was supported by the fellowship program “ProfesseurInvité du Sud” from “l'Agence Universitaire de la Francophonie”,sponsored by A.R.C. We also acknowledge the contribution of WasteManagement and NSERC (Canada) under the Cooperative Researchand Development grant CRD 379885-08.

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