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Effects of the main extraction parameters on chemical and microbial characteristics of compost tea
Waste Management xxx (2016) xxx–xxx
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Effects of the main extraction parameters on chemical and microbial characteristics of compost tea M.K. Islam a, T. Yaseen a, A. Traversa b, M. Ben Kheder c, G. Brunetti b, C. Cocozza b,⇑ a
International Centre for Advanced Mediterranean Agronomic Studies, Via Ceglie 9, 70010 Valenzano, Bari, Italy Dipartimento di Scienze del Suolo, della Pianta e degli Alimenti, University of Bari ‘‘Aldo Moro”, Via G. Amendola 165/A, 70126 Bari, Italy c Centre Technique d’Agriculture Biologique, B.P. 54, 4042 Chatt Meriem, Sousse, Tunisia b
a r t i c l e
i n f o
Article history: Received 15 September 2015 Revised 18 March 2016 Accepted 22 March 2016 Available online xxxx Keywords: Compost extract Compost/water ratio Extraction time Storage duration Storage temperature
a b s t r a c t The rising popularity of compost tea as fertilizer or foliar spray against pathogens has encouraged many researchers to evaluate its performance without standardizing its quality, so obtaining inconsistent and controversial results. The fertilizing and pesticide-like effects of compost tea are due to its chemical and microbiological properties. Therefore, this study aimed to identify the best combination of the compost tea extraction parameters for exalting both chemical and microbiological features. A factorial design was adopted to evaluate the effects of compost/water ratio, extraction time, storage duration and storage temperature in different combination on physical, chemical and microbiological characteristics of compost tea, and the results were elaborated through different statistical analyses. Compost tea nutrients and microorganisms were influenced by compost/water ratio and extraction time. In addition, the storage duration affected the microbial populations, whereas the storage temperature influenced only the fungal population of compost tea. Results suggested that the best combination of the studied parameters was: 1:2.5 compost/water ratio, 2 days of extraction time and the compost tea should be utilized immediately after the extraction, since the storage reduced the microbial populations. Ó 2016 Elsevier Ltd. All rights reserved.
1. Introduction Conventional agriculture is generally characterized by the use of a great amount of external inputs such as synthetic fertilizers and pesticides, growth regulators, resulting in heavy reliance on not-renewable resources, reduced biodiversity, soil degradation, chemical residues in food, and health risks to farm workers handling pesticides (Matson et al., 1997; Drinkwater et al., 1998; Tillman, 1999; Zhu et al., 2000; Reganold et al., 2001). As a consequence, sustainable agriculture, and in particular organic farming, emerges with the aim of solving a series of environmental, safety, and health problems arising from the modern conventional agriculture (Biao et al., 2003; Adl et al., 2011). Sustainable farmers attempt to close nutrient cycle on their farms, maximize re-use, employ rotation system, protect environmental quality, and enhance beneficial biological interactions and processes (Vandermeer, 1995; Casado and de Molina, 2009). The fertility management in sustainable agriculture relies on a long-term integrated approach in order to produce higher yields (Badgly and ⇑ Corresponding author at: Dipartimento di Scienze del Suolo, della Pianta, e degli Alimenti, Università di Bari, Via Amendola 165/A, 70126 Bari, Italy. E-mail address: [email protected] (C. Cocozza).
Perfetto, 2007) rather than the more short-term solutions common in conventional agriculture (Watson et al., 2002). Compost tea (CT) is a water extract of compost (Riggle, 1996; Ksheem et al., 2015) and it can be used to correct the nutrient deficiency during the crop production and/or to protect the cultivations. In particular, CT applied to soil affects the rhizosphere of the plant by carrying nutrients and microorganisms (Bess, 2000), whereas CT sprayed on leaf surfaces typically alters the set of organisms on the foliage through the inoculation of beneficial microorganisms that are antagonistic towards various plant pathogens (Noble and Coventry, 2005; Pane et al., 2011) and through the supply of microbial by-products and nutrients that help the survival of phyllosphere microorganisms. One of the problems in exploring the effects of CT on plants is the lack of a standardized extraction process. Thus, it is not surprising that the results from various experiments with CT are inconsistent and often conflicting. In fact, the quality of CT may vary considerably because of differences in procedures used for preparation of the extracts, source, composition, quality, and maturity of the compost (Weltzien, 1992), length of storage, and possibly other factors (Al-Dahmani et al., 2003). St. Martin (2014) showed that the plant diseases suppressiveness of CT is higher for the vermicompost and vermicasting in comparison to compost, regardless
http://dx.doi.org/10.1016/j.wasman.2016.03.042 0956-053X/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Islam, M.K., et al. Effects of the main extraction parameters on chemical and microbial characteristics of compost tea. Waste Management (2016), http://dx.doi.org/10.1016/j.wasman.2016.03.042
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their composition. Scheuerell and Mahaffee (2006) studied the effect of 30 compost teas on gray mold and their results agreed with prior reports of compost classes composed of manures and plant material appearing to be more suitable for producing disease suppressive compost teas than other classes of compost. Weltzien (1990, 1992) has shown that composts prepared with animal manures determined the most suppressive CT, compared to those obtained from composts prepared only with plant residues. In addition, compost should be 2–6 months old to be more suppressive (Tränkner, 1992). The compost/water ratio utilized for the extraction of CT can be considered a crucial parameter, in order to obtain CT of high quality. Previous studies (Zhang et al., 1998; Weltzien, 1990) have shown a suppressiveness potential of CT obtained using a compost/water ratio between 1:1 and 1:50, although most of the studies utilize a compost/water ratio between 1:3 and 1:10 (Scheuerell and Mahaffee, 2002). Few reports about the effect of extraction time on the final quality of the compost tea obtained under aerobic conditions are available in literature. In particular, Cantisano (1998) considered 1 day an optimal extraction time in order to obtain the best result for foliar feeding, and 7– 14 days to obtain the best result for disease control. Ingham (1999) suggested 18–24 h as the optimal extraction time of CT, coinciding with the maximum activity of the microbial biomass. With regards to the storage of CT, if the mode of action is mainly due to competition, the compost tea suppressiveness is reduced with storage, whereas if the mode of action is due to stable metabolites secreted into the water, this reduction not occurred (Scheuerell and Mahaffee, 2002). A previous study (Yohalem et al., 1994) has shown that a compost tea can be stored for four months without losing its effectiveness in terms of suppression of conidia germination. In addition, at 20 °C no loss of efficacy of CT was observed, whereas a reduction of its efficacy was observed at 4 °C. Since the fertilizing and the pesticide-like effects of compost teas are due to their chemical and microbiological properties, the aim of the present paper was to identify the best combination of compost/water ratio, extraction time, and storage duration and temperature to exalt the aforementioned features of compost tea. 2. Materials and methods 2.1. Compost and irrigation water Compost was obtained at the CIHEAM-IAM (Centre International de Hautes Etudes Agronomiques Méditerranéennes – Istituto Agronomico Mediterraneo) composting facility, located in Valenzano (Apulia region, Southern Italy), using olive pruning residues, bovine manure and wheat straw as raw materials. Since the production and utilization of the CT mainly occurs on farm, quite common agricultural wastes (pruning residues, straw and manures) have been utilized in the experimentation. Therefore, the results of this study could have broad applicability, in the framework of a sustainable agriculture. A pile of 25 m3 was prepared respecting the microbial requirements and its temperature was kept P55 °C for at least three days by turning and irrigating periodically the windrow in order to obtain the hygienization of the biomasses. The curing phase was characterized by less frequent turning and continued till the ninetieth day in order to achieve further stabilization. At the end of the composting, five samples were collected randomly spanning the whole compost pile. Table 1 reports the main features of the final compost. 2.2. Compost tea production and characterization The compost was thoroughly mixed to insure physical/chemical uniformity of the sample before the extraction of CT. The CT was
Table 1 Chemical properties of compost used for compost tea extraction. Compost parameters
Unit
Value
pH (H2O) (3:50, w/v) Electrical conductivity (1:10, w/v) Organic carbon Organic matter Total nitrogen C:N ratio Total P (as P) Total Ca Total K (as K) Total Mg Total Na Total Fe Total Cd Total Cr Total Cu Total Mn Total Ni Total Pb Total Zn Total Hg
dS m 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1
9.5 ± 0.13 2.1 ± 0.36 158.9 ± 12.5 274.0 ± 21.6 16.7 ± 0.12 9.5 5.6 ± 0.35 103.4 ± 12.3 78.4 ± 9.6 75.6 ± 9.0 56.6 ± 6.7 14.7 ± 2.1 2.0 ± 0.2 22.4 ± 3.3 65.0 ± 7.6 625.9 ± 32.4 17.8 ± 2.6 24.7 ± 1.7 160.4 ± 24.7 <0.4
obtained suspending the final compost in the irrigation water obtained from the artesian well at CIHEAM-IAM. The main characteristics of the irrigation water are reported in Table 2. The bucket-bubbler method was used for compost tea extraction. This is a simple and feasible method in which the mixture is deliberately aerated, allowing large numbers of beneficial organisms to populate this mixture (Scheuerell, 2003; Kelley, 2004; Ingham, 2005). Each extraction unit consisted of a plastic container open at top and with a tap at bottom, a 6 W pump with two outlets, two plastic pipes with two control valves and two bubblers. For CT extraction, ten liters of water were added in each extraction unit. Two bubblers were immerged and placed at the bottom of the bucket and the pumps were put on to start bubbling. After a while, compost was added slowly at different rates per treatment. Treatments consisted of three levels of compost/water ratio (CWR: 1/2.5, 1/5 and 1/10) replicated three times: therefore, the extraction units were nine. Further treatments were: three levels of extraction time (ET: 2 days, 4 days and 6 days); five levels of storage duration (SD: 0 week, 1 week, 2 weeks, 3 weeks and 4 weeks); and two levels of storage temperature [ST: room temperature and cool temperature (4 °C)]. Eighteen samples of CT were collected from each extraction unit at the end of each extraction time (54 samples from a set of compost/water ratio replicates) using a plastic beaker and then filtered by a strainer. Six samples of 54 were immediately analyzed (time 0), whereas 48 were stored in 100 mL sterile plastic bottles for different duration (from one to four weeks). Twenty four samples were preserved at room temperature (24–28 °C) and twenty four at cool temperature (4 °C). CT samples were characterized for their main physical and chemical properties according to the Italian Official Methods of Fertilizers Analyses (Trinchera et al., 2006).
Table 2 Chemical properties of water used for compost tea extraction. Water parameters
Unit
pH Electrical conductivity Cl HCO3 Ca++ Mg++ K+ Na+
dS m mg L mg L mg L mg L mg L mg L
Value 1 1 1 1 1 1 1
7.4 1.05 113.4 457.5 162.2 18.2 7.8 50.6
Please cite this article in press as: Islam, M.K., et al. Effects of the main extraction parameters on chemical and microbial characteristics of compost tea. Waste Management (2016), http://dx.doi.org/10.1016/j.wasman.2016.03.042
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2.3. Microbiological analysis The CT samples were characterized by the means of total bacteria, fungi and actinomycetes colonies. The CT samples were serially diluted from 10 1 to 10 5; aliquots of 100 lL were spread in Petri dishes amended with semi selective nutrient yeast dextrose agar (NYDA) media for the determination of bacteria and fungi, and with the selective starch casein agar (SCA) for the actinomycetes quantification. The NYDA (for 1 L: 10 g D-Glucose, 5 g yeast extract, 8 g nutrient broth, 18 g agar) was amended with ampicillin 250 mg L 1 and streptomycin sulfate 250 mg L 1 for fungi detection, and with cycloheximide 150 mg L 1 for bacteria detection. The antibiotic solutions were added to the respective semi selected NYDA medium after filtration using 0.45 lm then 0.22 lm filters. The SCA medium was obtained as described by Hirsch and Christensen (1983). At the end of each extraction time, CT samples were plated on Petri dishes, incubated at 24 °C and bacteria, fungi and actinomycetes colonies (CFU) were counted after 2–3 days, 4–5 days and 12–14 days, respectively. 2.4. Statistical analysis Each CT sample was analyzed in 6 replicates. The effects of compost/water ratio, extraction time, and storage duration and temperature on physical, chemical and microbiological characteristics of CT were examined using a four-way factorial ANOVA without replication. All interaction terms were included except for the fourth-order term (Sokal and Rohlf, 1981; Hettler and Hare, 1998). Microbiological data were log-transformed and then statistically evaluated for each extraction time (Mills and Wassel, 1980; Hughes et al., 2001; Papineau et al., 2005; Enticknap et al., 2006). The effects of extraction time, storage duration and storage temperature on physical, chemical and microbiological characteristics of CT were examined using a three-way factorial ANOVA. Analyses were done by GLM procedure of SAS/STATÒ software (SAS Institute Inc., 2000). Main effects of means were compared by Duncan’s multiple range test (Duncan, 1955) at 0.05 level of probability. Means of two-way interactions were compared by Bonferroni all pairwise multiple comparison test (Savin, 1980). Means of three way interactions were compared by posthoc LSD test (Sokal and Rohlf, 1981). Regression analyses were done by JMP software (SAS Institute Inc., 2007). 3. Results 3.1. Chemical properties of compost tea According to the results presented in Table 3, the pH of CT appeared significantly influenced only by the storage duration and, a lesser extent, by the storage temperature. In particular, the average pH of CT (8.8) decreased with increasing of storage duration and reached a value of 8.1 after 4 weeks. With the increase of storage duration, organic acid anions contained in the original compost, i.e. oxalate, citrate, and malate, were released due to the microbial decomposition, which balances the excess of cations (Noble et al., 1996; Yan et al., 1996). Besides, the atmospheric CO2 or CO2 produced by microbial activities dissolved in CT forming carbonic acid, so determining a decrease of pH over time. An increase of CT storage temperature could cause a decrease in its viscosity, an increase in the mobility of its ions in solution and an increase in the number of ions and protons in the same solution due to the dissociation of molecules (Zumdahl, 1993).
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The electrical conductivity (EC) for different levels of compost/ water ratio and extraction time differed significantly (Table 3). In particular, the EC values increased from 1:10 to 1:2.5 compost/ water ratio and with the increasing of extraction time. The influence on EC exerted by these parameters could be explained by the fact that with the increased of compost quantity and extraction time, total solute concentration in CT increased. Kim et al. (2015) tested teas obtained from an oriental medicinal herbs compost, a vermicompost, a rice straw compost, and a mixtures of three composts on the growth of different plants and reported an increased of EC after one day of incubation with a maximum value showed after 5 days of incubation. The total nitrogen (TN) was significantly influenced by compost/water ratio, extraction time and storage duration (Table 3). Fig. 1 illustrates the interaction between extraction time and storage duration, at compost/water ratio of 1:2.5, on TN of CT. This combination was chosen since it showed the highest TN content among the ones studied. At 0 week of storage, means of TN after 2, 4 and 6 days of extraction were 1.1, 0.7 and 0.8 g L 1, respectively. Generally speaking, the extraction time did not influence or influenced negatively the TN content of the CT regardless the storage duration. The organic matter (OM) subjected to composting contains various organic materials such as cellulose, lignin, non-cellulosic carbohydrates (hemicelluloses, starch, and mono- and oligosaccharides), proteins and lipids which are metabolized along different biochemical pathways, i.e. mineralization, transformation or stabilization through the formation of humic-like substances (Plaza et al., 2005; Senesi et al., 2007), and finally release N (Said-Pullicino et al., 2007). The biochemical transformations of OM that take place during composting are determined by microorganisms whose metabolism occurs in the water-soluble phase. As a consequence, dissolved OM fraction of compost mainly contributes to the TN of CT. Nitrogen supply from organic amendments depends on both the initial availability of N in the amendments and also on the longer-term rate of mineralization. Thus, interactions among compost/water ratio, extraction time and storage duration entirely depends on the chemical composition of compost organic nitrogen. The estimated equation for compost/water ratio ⁄ extraction time ⁄ storage duration interaction on TN is: TN (g L 1) = 1.369 CWR ⁄ 0.4 ET ⁄ 0.022 + SD ⁄ 0.003 (R2 = 0.90, p < 0.0001, n = 90). The organic carbon (OC) was significantly influenced by compost/water ratio, extraction time and storage duration (Table 3). Fig. 2 illustrates the interaction between extraction time and storage duration, at compost/water ratio of 1:2.5, on OC of CT. At 0 week of storage, means of total OC after 2, 4 and 6 days of extraction were 9.6, 6.5 and 7.5 g L 1, respectively. In general, the extraction time did not influence or influenced negatively the OC content regardless the storage duration. Results show that the highest OC was obtained from compost/water ratio of 1:2.5. Also in this case, dissolved OC is the organic fraction containing organic materials utilized as an energy source, bio-originating macromolecules such as enzymes, polysaccharides and proteins, breakdown products and the repolymerized compounds (Said-Pullicino et al., 2007). Water-extractable OC (WEOC) fraction of compost mainly contributed to the total OC of CT and its response to extraction factors depends on its stability and microbial activities. Generally, it is observed a decrease of OC from 2 to 6 days of extraction. This result can be explained by that the WEOC dissolved fully within 2 days of extraction and, after that, part of it is used by CT microorganisms or lost as CO2 due to their metabolism. The estimated equation for compost/water ratio ⁄ extraction time ⁄ storage duration interaction on OC is: OC (g L 1) = 11.813 CWR ⁄ 3.282 ET ⁄ 0.238 SD ⁄ 0.011 (R2 = 0.89, p < 0.0001, n = 90).
Please cite this article in press as: Islam, M.K., et al. Effects of the main extraction parameters on chemical and microbial characteristics of compost tea. Waste Management (2016), http://dx.doi.org/10.1016/j.wasman.2016.03.042
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Table 3 ANOVA (P-values) of effect of CWR, ET, SD and ST on compost tea quality. Bold figures indicate significant P-values. Source of variation i
CWR ETj SDk STl CWR ⁄ ET CWR ⁄ SD CWR ⁄ ST ET ⁄ SD ET ⁄ ST SD ⁄ ST CWR ⁄ ET ⁄ SD CWR ⁄ ET ⁄ ST CWR ⁄ SD ⁄ ST ET ⁄ SD ⁄ ST Error (CWR ⁄ ET ⁄ SD ⁄ ST) a b c d e f g h i j k l
df
pH
ECa
TNb
OCc
OMd
CNRe
BPDf
FPDg
APDh
2 2 4 1 4 8 2 8 2 4 16 4 8 8 16
0.3567 0.6032 <0.0001 0.039 0.2306 0.7283 0.4234 0.844 0.5775 0.3005 0.6382 0.2559 0.5312 0.933
<0.0001 <0.0001 0.2042 1 0.126 0.342 0.1824 0.6572 0.2691 0.8234 0.3667 0.6512 0.59 0.2651
<0.0001 <0.0001 <0.0001 0.5093 0.036 0.0031 0.3685 0.0054 0.8603 0.1282 0.0008 0.8712 0.1218 0.0882
<0.0001 <0.0001 0.2696 0.6498 0.6562 0.5324 0.4088 0.4867 0.7257 0.4717 0.0333 0.9822 0.361 0.5698
<0.0001 <0.0001 0.2602 0.7032 0.6476 0.5074 0.4217 0.5121 0.7294 0.474 0.041 0.9774 0.3729 0.5977
0.0851 0.0019 <0.0001 0.3517 0.0379 0.0122 0.9957 0.0005 0.1935 0.7586 0.0152 0.5574 0.3193 0.3516
<0.0001 0.0263 <0.0001 0.0049 0.3949 0.4125 0.5975 0.0057 0.629 0.3341 0.6518 0.6713 0.8772 0.0844
<0.0001 0.0205 <0.0001 0.002 0.6899 0.5993 0.1714 0.0088 0.6451 0.4484 0.7778 0.7849 0.8106 0.5669
<0.0001 0.1987 0.0188 0.6046 0.6251 0.3557 0.1124 0.0773 0.9375 0.5301 0.7532 0.3686 0.834 0.9714
Electrical conductivity. Total nitrogen. Organic carbon. Organic matter. Carbon/nitrogen ratio. Bacteria population density. Fungal population density. Actinomycetes population density. Compost/water ratio. Extraction time. Storage duration. Storage temperature.
CWR 1:2.5
Fig. 1. Effect of the interaction among compost/water ratio (CWR) 1:2.5, extraction time and storage duration on compost tea total N.
CWR 1:2.5
ratio (CNR) of CT. The highest CNR (13.0) was obtained after 6 days of extraction time and 0 week of storage, while the lowest one was recorded after 4 days of extraction time and 2 weeks of storage. The changes in the CNR reflect OM decomposition and depends on the cumulative effect of OC and TN that were previously described. The estimated equation for compost/water ratio ⁄ extraction time ⁄ storage duration interaction on carbon to nitrogen ratio is: CNR = 10.367 CWR ⁄ 0.198 ET ⁄ 0.096 SD ⁄ 0.207 (R2 = 0.33, p < 0.0001, n = 90). CNR was mainly affected by storage duration possibly because of comparatively higher loss of carbon than nitrogen during the storage period as CO2 released from the microbial activity. 3.2. Microbial properties of compost tea The interaction between extraction time and storage duration on bacteria population density (BPD) resulted the sole significant combination (Table 3), and is illustrated in Fig. 4. The highest
CWR 1:10
Fig. 2. Effect of the interaction among the compost/water ratio (CWR) 1:2.5, extraction time and storage duration on the organic carbon content of the compost tea.
The same trend was obviously obtained for the organic matter (OM) of CT (Table 3). Fig. 3 depicts the interaction between extraction time and storage duration, at compost/water ratio of 1:10, on carbon/nitrogen
Fig. 3. Effect of the interaction among compost/water ratio (CWR) 1:10 and extraction time (ET) and storage duration (SD) on the C/N ratio (CNR) of compost tea.
Please cite this article in press as: Islam, M.K., et al. Effects of the main extraction parameters on chemical and microbial characteristics of compost tea. Waste Management (2016), http://dx.doi.org/10.1016/j.wasman.2016.03.042
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BPD (6.43 log CFU L 1) was obtained after 4 days of extraction with zero week of storage duration. With the increase of storage duration, BPD decreased at variable rates depending on interaction and the lowest BPD (5.77 log CFU L 1) was obtained after 6 days of extraction with 4 weeks of storage. The compost/water ratio significantly affected the BPD (Table 3) since the average BPD recorded from compost/water ratio of 1:2.5, 1:5 and 1:10 were 6.15, 6.04 and 5.93 log CFU L 1, respectively. In addition, the storage temperature influenced significantly the BPD. In fact, the BPD was 6.09 log CFU L 1 at cool temperature and reduced to 5.99 log CFU L 1 at room temperature. These findings agree with Ingham (2005) who reported that there is a balance between extraction of nutrients and growth of organisms, giving an optimal time for tea production, depending on the extract conditions of brewing. The estimated equations for extraction time ⁄ storage duration interaction on bacteria population density is BPD (log CFU L 1) = 6.295 ET ⁄ 0.004 SD ⁄ 0.017 (R2 = 0.39, p < 0.0001, n = 90). The storage duration affect the bacterial activity possibly due to the consumption of nutrients, microbial competition and production of toxic molecules. Fig. 5 shows the significant interaction (Table 3) between extraction time and storage duration on fungal population density (FPD). It was observed that FPD decreased with increasing of extraction time and storage duration. The highest FPD (5.03 log CFU L 1) was obtained after 2 days of extraction with zero week of storage, whereas the lowest FPD (4.02 log CFU L 1) was recorded after 6 days of extraction with 4 weeks of storage. The compost/water ratio significantly influenced the FPD: the average FPD recorded were 4.62, 4.43 and 4.33 log CFU L 1 from compost/water ratio of 1:2.5, 1:5 and 1:10, respectively. The FDP was significantly affected by the storage temperature too: the FPD was 4.53 log CFU L 1 at cool temperature and reduced to 4.39 log CFU L 1 at room temperature. Both BPD and FPD increased from 1:10 to 1:2.5 compost/water ratio and at cool temperature. The estimated equation for extraction time ⁄ storage duration interaction on fungal population density is FPD (log CFU L 1) = 4.852 ET ⁄ 0.033 SD ⁄ 0.018 (R2 = 0.39, p < 0.0001, n = 90). As like as bacterial population, the storage duration affects negatively the FPD possibly because of the same reasons described above. According to results in Table 3, the compost/water ratio had significant effect on actinomycetes population density (APD). Means of APD recorded from compost/water ratio of 1:2.5, 1:5 and 1:10 were 5.85, 5.71 and 5.49 log CFU L 1, respectively. The storage duration significantly influenced the APD only after 4 weeks of storage, when the APD was found slightly lower. According to a previous study (Scheuerell and Mahaffee, 2006), nitrogen rich feedstock, such as compost containing manures,
Fig. 4. Effect of the interaction of storage duration and extraction time on the bacteria population density (BPD) of compost tea.
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Fig. 5. Effect of the interaction of storage duration and extraction time on the fungal population density (FPD) of compost tea.
produces a compost tea with higher bacterial content, respect to the fungal ones. Also Kim et al. (2015) reported that microbial communities of different aerated compost teas were predominantly bacteria.
3.3. Correlations Table 4 shows the correlation of compost/water ratio, extraction time, storage duration and storage temperature with CT quality parameters together with their significance probability. It is evident that the compost/water ratio showed significant (P < 0.001) correlation with EC, TN, OC and microbial populations. In particular, the chemical parameters were almost equally influenced by the dilution, while the actinomycetes population was more negatively affected by the dilution in comparison to bacteria and fungi. The storage duration had significant (P < 0.001) negative correlation with pH, CNR bacterial and fungal populations, while the actinomycetes population showed a lower significance (P < 0.05). Apparently, the microbial populations have contributed to the reduction of the pH with their metabolism, while their survival has been reduced significantly with time. The storage temperature showed a significant correlation only with fungal population since it has suffered for the room temperature storage in comparison to the bacteria and actinomycetes.
4. Discussion Results of hundreds of analyses indicated that compost/water ratio significantly affected compost tea electrical conductivity, total nitrogen, organic carbon and matter, and microorganisms densities with an increasing of these parameters from 1:10 to 1:2.5 compost/water ratio. Among several compost/water ratios from 1:2 to 1:50 (v/v), St. Martin (2014) showed that the denser CTs were more effective towards different pathogens. In addition, Scheuerell and Mahaffee (2004) and Cayuela et al. (2008) found that dilution decreased disease suppression of Pythium dampingoff of cucumber and Pythium capsici, respectively. Further, Evans et al. (2013) found positive effects of a manure compost tea obtained at compost/water ratio of 1:3 after 48 h of fermentation on the pathogen Botrytis cinerea. About the fertilizing effect of CT, Wang et al. (2014) reported no effect on the yield of zucchini using a chicken manure based vermicompost tea (1:10, v/v, continuously aerated for 12 h) as fertilizer. Also Hewidy et al. (2015) showed no effect on broccoli yield of a CT obtained from the organic fraction of the municipal solid wastes and pruning residues (1:5, v/v, continuously aerated for 24 h).
Please cite this article in press as: Islam, M.K., et al. Effects of the main extraction parameters on chemical and microbial characteristics of compost tea. Waste Management (2016), http://dx.doi.org/10.1016/j.wasman.2016.03.042
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Table 4 Pearson correlation coefficient (r) of CWR, ET, SD and ST with CT quality parameters. Bold figures indicate significant P-values. Parameters CWR ETj SDk STl *** * NS a b c d e f g h i j k l
i
ECa
pH NS
0.098 0.056NS 0.741*** 0.149NS
TNb ***
0.976 0.110 NS 0.027NS 0.000NS
OCc ***
0.936 0.101NS 0.093NS 0.010NS
OMd ***
0.933 0.135NS 0.035NS 0.010NS
CNRe ***
0.929 0.134NS 0.040NS 0.015NS
BPDf NS
0.098 0.188NS 0.534*** 0.050NS
FPDg ***
0.363 0.024NS 0.656*** 0.190NS
APDh ***
0.396 0.184NS 0.595*** 0.231*
0.748*** 0.127NS 0.215* 0.035NS
P < 0.001. P < 0.05. P P 0.05. Electrical conductivity. Total nitrogen. Organic carbon. Organic matter. Carbon/nitrogen ratio. Bacteria population density. Fungal population density. Actinomycetes population density. Compost/water ratio. Extraction time. Storage duration. Storage temperature.
Significant decrease of total N, organic carbon and organic matter contents of compost tea were observed when the extraction time increased, together with a slight reduction of bacterial and fungal populations. Ingham and Alms (1999) reported that the optimum extraction time is usually between 18 and 36 h, when the microbial biomass activity is the highest. St. Martin and Brathwaite (2012) showed that brewing cycles of 18 h for aerated compost tea can be used to produce CT with optimal Pythium ultimum in vitro inhibitive properties using lawn clipping and banana leaf composts. It was observed that the storage duration did not significantly influence the chemical properties of compost tea, with the only exception of pH and total N. In contrast, fungi and bacteria populations appeared more influenced from the storage duration since they decreased with increasing the storage duration possibly because of the competition for nutrients and oxygen among microorganisms and the release of metabolic toxic molecules. The same trend, but at lesser extent, was observer for actinomycetes populations. Long storage times negatively impacts upon the diversity of microorganisms, as well as nutrients carried by the tea for plant use (Bess, 2000). Number and activity of organisms reduces significantly with storage and, although this reduction is acceptable for a soil application of compost tea, it is not acceptable for a foliar application of compost tea (Ingham, 2005). McQuilken et al. (1994) found that the age of CT had some effects on its subsequent activity against both germination and mycelial growth of B. cinerea. In particular, a 3–12 day old CT still reduced the conidial germination, whereas a >18 day old CT significantly declined its inhibition properties. In addition, a 3–5-day old CT showed the highest inhibition against the mycelia growth. The storage temperature influenced the fungal and bacterial populations and, at a lesser extent, the pH of compost tea. To our knowledge, no study has been done in relation to the compost tea storage temperature, even if the tea temperature is considered very important for enzyme activity. In particular, at temperatures below the optimum, enzymes do not catalyze any reaction and membranes solidify, whereas at temperatures above the optimum, denaturation of enzymes, transport carriers and proteins occurs (Madigan et al., 2015). Many interactions and individual effect of the studied four factors (compost/water ratio, extraction time, storage duration and storage temperature) on compost tea quality parameters were found significant and those were elaborated by regression equations. Significance tests of predicted model were performed in
comparison with actual data. Test of significance of individual factor or interaction factors at individual level revealed that many quality parameters of compost tea can be predicted by compost/ water ratio and extraction time. According to the results, the best combination between chemical and microbial properties of CT can be obtained using a compost/water ratio of 1:2.5 and an extraction time of two days. If the CT production is devoted to exploit solely one or more microbial features, no storage should be applied since the microbial populations of CT decline with time. Therefore, the CT should be prepared and immediately used in agriculture, otherwise a cool storage is necessary to preserve the microbial populations of CT. However, the sprinkler and drip irrigation systems clog when a very dense CT is used for fertigation. It would be better to utilize surface irrigation systems or, if possible, to adapt slurry spreaders for soil application, and wet somehow the aerial parts of crops for the foliar application. Finally, further studies are needed to expand the validity of the proposed equations for different compost source and different maturity level, and to evaluate the performance of extracted compost tea on field crops. Acknowledgements Authors are very grateful to Prof. Nicola Senesi for his help to improve the quality of the manuscript. Authors wish to thank Maria Giuseppina Borracci for her moral support. References Adl, S., Iron, D., Kolokolnikov, T., 2011. A threshold area ratio of organic to conventional agriculture causes recurrent pathogen outbreaks in organic agriculture. Sci. Total Environ. 409, 2192–2197. Al-Dahmani, J.H., Abbasi, P.A., Miller, S.A., Hoitink, H.A.J., 2003. Suppression of bacterial spot of tomato with foliar sprays of compost extracts under greenhouse and field conditions. Plant Dis. 87, 913–919. Badgly, C., Perfetto, I., 2007. Can organic agriculture feed the world? Renew. Agric. Food Syst. 22, 80–85. Bess, V.H., 2000. Understanding compost tea. Biocycle 41 (10), 71–72. Biao, X., Xiaorong, W., Zhuhong, D., Yaping, Y., 2003. Critical impact assessment of organic agriculture. J. Agric. Environ. Ethics 16, 297–311. Cantisano, A., 1998. Compost teas. Organic Ag Advisors Letter. Colfax, California. Casado, G.I.G., de Molina, M.G., 2009. Preindustrial agriculture versus organic agriculture: the land cost of sustainability. Land Use Policy 26, 502–510. Cayuela, M., Millner, P., Meyer, S., Roig, A., 2008. Potential of olive mill waste and compost as biobased pesticides against weeds, fungi, and nematodes. Sci. Total Environ. 399, 11–18.
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Effects of the main extraction parameters on chemical and microbial characteristics of compost tea M.K. Islam a, T. Yaseen a, A. Traversa b, M. Ben Kheder c, G. Brunetti b, C. Cocozza b,⇑ a
International Centre for Advanced Mediterranean Agronomic Studies, Via Ceglie 9, 70010 Valenzano, Bari, Italy Dipartimento di Scienze del Suolo, della Pianta e degli Alimenti, University of Bari ‘‘Aldo Moro”, Via G. Amendola 165/A, 70126 Bari, Italy c Centre Technique d’Agriculture Biologique, B.P. 54, 4042 Chatt Meriem, Sousse, Tunisia b
a r t i c l e
i n f o
Article history: Received 15 September 2015 Revised 18 March 2016 Accepted 22 March 2016 Available online xxxx Keywords: Compost extract Compost/water ratio Extraction time Storage duration Storage temperature
a b s t r a c t The rising popularity of compost tea as fertilizer or foliar spray against pathogens has encouraged many researchers to evaluate its performance without standardizing its quality, so obtaining inconsistent and controversial results. The fertilizing and pesticide-like effects of compost tea are due to its chemical and microbiological properties. Therefore, this study aimed to identify the best combination of the compost tea extraction parameters for exalting both chemical and microbiological features. A factorial design was adopted to evaluate the effects of compost/water ratio, extraction time, storage duration and storage temperature in different combination on physical, chemical and microbiological characteristics of compost tea, and the results were elaborated through different statistical analyses. Compost tea nutrients and microorganisms were influenced by compost/water ratio and extraction time. In addition, the storage duration affected the microbial populations, whereas the storage temperature influenced only the fungal population of compost tea. Results suggested that the best combination of the studied parameters was: 1:2.5 compost/water ratio, 2 days of extraction time and the compost tea should be utilized immediately after the extraction, since the storage reduced the microbial populations. Ó 2016 Elsevier Ltd. All rights reserved.
1. Introduction Conventional agriculture is generally characterized by the use of a great amount of external inputs such as synthetic fertilizers and pesticides, growth regulators, resulting in heavy reliance on not-renewable resources, reduced biodiversity, soil degradation, chemical residues in food, and health risks to farm workers handling pesticides (Matson et al., 1997; Drinkwater et al., 1998; Tillman, 1999; Zhu et al., 2000; Reganold et al., 2001). As a consequence, sustainable agriculture, and in particular organic farming, emerges with the aim of solving a series of environmental, safety, and health problems arising from the modern conventional agriculture (Biao et al., 2003; Adl et al., 2011). Sustainable farmers attempt to close nutrient cycle on their farms, maximize re-use, employ rotation system, protect environmental quality, and enhance beneficial biological interactions and processes (Vandermeer, 1995; Casado and de Molina, 2009). The fertility management in sustainable agriculture relies on a long-term integrated approach in order to produce higher yields (Badgly and ⇑ Corresponding author at: Dipartimento di Scienze del Suolo, della Pianta, e degli Alimenti, Università di Bari, Via Amendola 165/A, 70126 Bari, Italy. E-mail address: [email protected] (C. Cocozza).
Perfetto, 2007) rather than the more short-term solutions common in conventional agriculture (Watson et al., 2002). Compost tea (CT) is a water extract of compost (Riggle, 1996; Ksheem et al., 2015) and it can be used to correct the nutrient deficiency during the crop production and/or to protect the cultivations. In particular, CT applied to soil affects the rhizosphere of the plant by carrying nutrients and microorganisms (Bess, 2000), whereas CT sprayed on leaf surfaces typically alters the set of organisms on the foliage through the inoculation of beneficial microorganisms that are antagonistic towards various plant pathogens (Noble and Coventry, 2005; Pane et al., 2011) and through the supply of microbial by-products and nutrients that help the survival of phyllosphere microorganisms. One of the problems in exploring the effects of CT on plants is the lack of a standardized extraction process. Thus, it is not surprising that the results from various experiments with CT are inconsistent and often conflicting. In fact, the quality of CT may vary considerably because of differences in procedures used for preparation of the extracts, source, composition, quality, and maturity of the compost (Weltzien, 1992), length of storage, and possibly other factors (Al-Dahmani et al., 2003). St. Martin (2014) showed that the plant diseases suppressiveness of CT is higher for the vermicompost and vermicasting in comparison to compost, regardless
http://dx.doi.org/10.1016/j.wasman.2016.03.042 0956-053X/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Islam, M.K., et al. Effects of the main extraction parameters on chemical and microbial characteristics of compost tea. Waste Management (2016), http://dx.doi.org/10.1016/j.wasman.2016.03.042
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their composition. Scheuerell and Mahaffee (2006) studied the effect of 30 compost teas on gray mold and their results agreed with prior reports of compost classes composed of manures and plant material appearing to be more suitable for producing disease suppressive compost teas than other classes of compost. Weltzien (1990, 1992) has shown that composts prepared with animal manures determined the most suppressive CT, compared to those obtained from composts prepared only with plant residues. In addition, compost should be 2–6 months old to be more suppressive (Tränkner, 1992). The compost/water ratio utilized for the extraction of CT can be considered a crucial parameter, in order to obtain CT of high quality. Previous studies (Zhang et al., 1998; Weltzien, 1990) have shown a suppressiveness potential of CT obtained using a compost/water ratio between 1:1 and 1:50, although most of the studies utilize a compost/water ratio between 1:3 and 1:10 (Scheuerell and Mahaffee, 2002). Few reports about the effect of extraction time on the final quality of the compost tea obtained under aerobic conditions are available in literature. In particular, Cantisano (1998) considered 1 day an optimal extraction time in order to obtain the best result for foliar feeding, and 7– 14 days to obtain the best result for disease control. Ingham (1999) suggested 18–24 h as the optimal extraction time of CT, coinciding with the maximum activity of the microbial biomass. With regards to the storage of CT, if the mode of action is mainly due to competition, the compost tea suppressiveness is reduced with storage, whereas if the mode of action is due to stable metabolites secreted into the water, this reduction not occurred (Scheuerell and Mahaffee, 2002). A previous study (Yohalem et al., 1994) has shown that a compost tea can be stored for four months without losing its effectiveness in terms of suppression of conidia germination. In addition, at 20 °C no loss of efficacy of CT was observed, whereas a reduction of its efficacy was observed at 4 °C. Since the fertilizing and the pesticide-like effects of compost teas are due to their chemical and microbiological properties, the aim of the present paper was to identify the best combination of compost/water ratio, extraction time, and storage duration and temperature to exalt the aforementioned features of compost tea. 2. Materials and methods 2.1. Compost and irrigation water Compost was obtained at the CIHEAM-IAM (Centre International de Hautes Etudes Agronomiques Méditerranéennes – Istituto Agronomico Mediterraneo) composting facility, located in Valenzano (Apulia region, Southern Italy), using olive pruning residues, bovine manure and wheat straw as raw materials. Since the production and utilization of the CT mainly occurs on farm, quite common agricultural wastes (pruning residues, straw and manures) have been utilized in the experimentation. Therefore, the results of this study could have broad applicability, in the framework of a sustainable agriculture. A pile of 25 m3 was prepared respecting the microbial requirements and its temperature was kept P55 °C for at least three days by turning and irrigating periodically the windrow in order to obtain the hygienization of the biomasses. The curing phase was characterized by less frequent turning and continued till the ninetieth day in order to achieve further stabilization. At the end of the composting, five samples were collected randomly spanning the whole compost pile. Table 1 reports the main features of the final compost. 2.2. Compost tea production and characterization The compost was thoroughly mixed to insure physical/chemical uniformity of the sample before the extraction of CT. The CT was
Table 1 Chemical properties of compost used for compost tea extraction. Compost parameters
Unit
Value
pH (H2O) (3:50, w/v) Electrical conductivity (1:10, w/v) Organic carbon Organic matter Total nitrogen C:N ratio Total P (as P) Total Ca Total K (as K) Total Mg Total Na Total Fe Total Cd Total Cr Total Cu Total Mn Total Ni Total Pb Total Zn Total Hg
dS m 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 g kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1 mg kg 1
9.5 ± 0.13 2.1 ± 0.36 158.9 ± 12.5 274.0 ± 21.6 16.7 ± 0.12 9.5 5.6 ± 0.35 103.4 ± 12.3 78.4 ± 9.6 75.6 ± 9.0 56.6 ± 6.7 14.7 ± 2.1 2.0 ± 0.2 22.4 ± 3.3 65.0 ± 7.6 625.9 ± 32.4 17.8 ± 2.6 24.7 ± 1.7 160.4 ± 24.7 <0.4
obtained suspending the final compost in the irrigation water obtained from the artesian well at CIHEAM-IAM. The main characteristics of the irrigation water are reported in Table 2. The bucket-bubbler method was used for compost tea extraction. This is a simple and feasible method in which the mixture is deliberately aerated, allowing large numbers of beneficial organisms to populate this mixture (Scheuerell, 2003; Kelley, 2004; Ingham, 2005). Each extraction unit consisted of a plastic container open at top and with a tap at bottom, a 6 W pump with two outlets, two plastic pipes with two control valves and two bubblers. For CT extraction, ten liters of water were added in each extraction unit. Two bubblers were immerged and placed at the bottom of the bucket and the pumps were put on to start bubbling. After a while, compost was added slowly at different rates per treatment. Treatments consisted of three levels of compost/water ratio (CWR: 1/2.5, 1/5 and 1/10) replicated three times: therefore, the extraction units were nine. Further treatments were: three levels of extraction time (ET: 2 days, 4 days and 6 days); five levels of storage duration (SD: 0 week, 1 week, 2 weeks, 3 weeks and 4 weeks); and two levels of storage temperature [ST: room temperature and cool temperature (4 °C)]. Eighteen samples of CT were collected from each extraction unit at the end of each extraction time (54 samples from a set of compost/water ratio replicates) using a plastic beaker and then filtered by a strainer. Six samples of 54 were immediately analyzed (time 0), whereas 48 were stored in 100 mL sterile plastic bottles for different duration (from one to four weeks). Twenty four samples were preserved at room temperature (24–28 °C) and twenty four at cool temperature (4 °C). CT samples were characterized for their main physical and chemical properties according to the Italian Official Methods of Fertilizers Analyses (Trinchera et al., 2006).
Table 2 Chemical properties of water used for compost tea extraction. Water parameters
Unit
pH Electrical conductivity Cl HCO3 Ca++ Mg++ K+ Na+
dS m mg L mg L mg L mg L mg L mg L
Value 1 1 1 1 1 1 1
7.4 1.05 113.4 457.5 162.2 18.2 7.8 50.6
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2.3. Microbiological analysis The CT samples were characterized by the means of total bacteria, fungi and actinomycetes colonies. The CT samples were serially diluted from 10 1 to 10 5; aliquots of 100 lL were spread in Petri dishes amended with semi selective nutrient yeast dextrose agar (NYDA) media for the determination of bacteria and fungi, and with the selective starch casein agar (SCA) for the actinomycetes quantification. The NYDA (for 1 L: 10 g D-Glucose, 5 g yeast extract, 8 g nutrient broth, 18 g agar) was amended with ampicillin 250 mg L 1 and streptomycin sulfate 250 mg L 1 for fungi detection, and with cycloheximide 150 mg L 1 for bacteria detection. The antibiotic solutions were added to the respective semi selected NYDA medium after filtration using 0.45 lm then 0.22 lm filters. The SCA medium was obtained as described by Hirsch and Christensen (1983). At the end of each extraction time, CT samples were plated on Petri dishes, incubated at 24 °C and bacteria, fungi and actinomycetes colonies (CFU) were counted after 2–3 days, 4–5 days and 12–14 days, respectively. 2.4. Statistical analysis Each CT sample was analyzed in 6 replicates. The effects of compost/water ratio, extraction time, and storage duration and temperature on physical, chemical and microbiological characteristics of CT were examined using a four-way factorial ANOVA without replication. All interaction terms were included except for the fourth-order term (Sokal and Rohlf, 1981; Hettler and Hare, 1998). Microbiological data were log-transformed and then statistically evaluated for each extraction time (Mills and Wassel, 1980; Hughes et al., 2001; Papineau et al., 2005; Enticknap et al., 2006). The effects of extraction time, storage duration and storage temperature on physical, chemical and microbiological characteristics of CT were examined using a three-way factorial ANOVA. Analyses were done by GLM procedure of SAS/STATÒ software (SAS Institute Inc., 2000). Main effects of means were compared by Duncan’s multiple range test (Duncan, 1955) at 0.05 level of probability. Means of two-way interactions were compared by Bonferroni all pairwise multiple comparison test (Savin, 1980). Means of three way interactions were compared by posthoc LSD test (Sokal and Rohlf, 1981). Regression analyses were done by JMP software (SAS Institute Inc., 2007). 3. Results 3.1. Chemical properties of compost tea According to the results presented in Table 3, the pH of CT appeared significantly influenced only by the storage duration and, a lesser extent, by the storage temperature. In particular, the average pH of CT (8.8) decreased with increasing of storage duration and reached a value of 8.1 after 4 weeks. With the increase of storage duration, organic acid anions contained in the original compost, i.e. oxalate, citrate, and malate, were released due to the microbial decomposition, which balances the excess of cations (Noble et al., 1996; Yan et al., 1996). Besides, the atmospheric CO2 or CO2 produced by microbial activities dissolved in CT forming carbonic acid, so determining a decrease of pH over time. An increase of CT storage temperature could cause a decrease in its viscosity, an increase in the mobility of its ions in solution and an increase in the number of ions and protons in the same solution due to the dissociation of molecules (Zumdahl, 1993).
3
The electrical conductivity (EC) for different levels of compost/ water ratio and extraction time differed significantly (Table 3). In particular, the EC values increased from 1:10 to 1:2.5 compost/ water ratio and with the increasing of extraction time. The influence on EC exerted by these parameters could be explained by the fact that with the increased of compost quantity and extraction time, total solute concentration in CT increased. Kim et al. (2015) tested teas obtained from an oriental medicinal herbs compost, a vermicompost, a rice straw compost, and a mixtures of three composts on the growth of different plants and reported an increased of EC after one day of incubation with a maximum value showed after 5 days of incubation. The total nitrogen (TN) was significantly influenced by compost/water ratio, extraction time and storage duration (Table 3). Fig. 1 illustrates the interaction between extraction time and storage duration, at compost/water ratio of 1:2.5, on TN of CT. This combination was chosen since it showed the highest TN content among the ones studied. At 0 week of storage, means of TN after 2, 4 and 6 days of extraction were 1.1, 0.7 and 0.8 g L 1, respectively. Generally speaking, the extraction time did not influence or influenced negatively the TN content of the CT regardless the storage duration. The organic matter (OM) subjected to composting contains various organic materials such as cellulose, lignin, non-cellulosic carbohydrates (hemicelluloses, starch, and mono- and oligosaccharides), proteins and lipids which are metabolized along different biochemical pathways, i.e. mineralization, transformation or stabilization through the formation of humic-like substances (Plaza et al., 2005; Senesi et al., 2007), and finally release N (Said-Pullicino et al., 2007). The biochemical transformations of OM that take place during composting are determined by microorganisms whose metabolism occurs in the water-soluble phase. As a consequence, dissolved OM fraction of compost mainly contributes to the TN of CT. Nitrogen supply from organic amendments depends on both the initial availability of N in the amendments and also on the longer-term rate of mineralization. Thus, interactions among compost/water ratio, extraction time and storage duration entirely depends on the chemical composition of compost organic nitrogen. The estimated equation for compost/water ratio ⁄ extraction time ⁄ storage duration interaction on TN is: TN (g L 1) = 1.369 CWR ⁄ 0.4 ET ⁄ 0.022 + SD ⁄ 0.003 (R2 = 0.90, p < 0.0001, n = 90). The organic carbon (OC) was significantly influenced by compost/water ratio, extraction time and storage duration (Table 3). Fig. 2 illustrates the interaction between extraction time and storage duration, at compost/water ratio of 1:2.5, on OC of CT. At 0 week of storage, means of total OC after 2, 4 and 6 days of extraction were 9.6, 6.5 and 7.5 g L 1, respectively. In general, the extraction time did not influence or influenced negatively the OC content regardless the storage duration. Results show that the highest OC was obtained from compost/water ratio of 1:2.5. Also in this case, dissolved OC is the organic fraction containing organic materials utilized as an energy source, bio-originating macromolecules such as enzymes, polysaccharides and proteins, breakdown products and the repolymerized compounds (Said-Pullicino et al., 2007). Water-extractable OC (WEOC) fraction of compost mainly contributed to the total OC of CT and its response to extraction factors depends on its stability and microbial activities. Generally, it is observed a decrease of OC from 2 to 6 days of extraction. This result can be explained by that the WEOC dissolved fully within 2 days of extraction and, after that, part of it is used by CT microorganisms or lost as CO2 due to their metabolism. The estimated equation for compost/water ratio ⁄ extraction time ⁄ storage duration interaction on OC is: OC (g L 1) = 11.813 CWR ⁄ 3.282 ET ⁄ 0.238 SD ⁄ 0.011 (R2 = 0.89, p < 0.0001, n = 90).
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M.K. Islam et al. / Waste Management xxx (2016) xxx–xxx
Table 3 ANOVA (P-values) of effect of CWR, ET, SD and ST on compost tea quality. Bold figures indicate significant P-values. Source of variation i
CWR ETj SDk STl CWR ⁄ ET CWR ⁄ SD CWR ⁄ ST ET ⁄ SD ET ⁄ ST SD ⁄ ST CWR ⁄ ET ⁄ SD CWR ⁄ ET ⁄ ST CWR ⁄ SD ⁄ ST ET ⁄ SD ⁄ ST Error (CWR ⁄ ET ⁄ SD ⁄ ST) a b c d e f g h i j k l
df
pH
ECa
TNb
OCc
OMd
CNRe
BPDf
FPDg
APDh
2 2 4 1 4 8 2 8 2 4 16 4 8 8 16
0.3567 0.6032 <0.0001 0.039 0.2306 0.7283 0.4234 0.844 0.5775 0.3005 0.6382 0.2559 0.5312 0.933
<0.0001 <0.0001 0.2042 1 0.126 0.342 0.1824 0.6572 0.2691 0.8234 0.3667 0.6512 0.59 0.2651
<0.0001 <0.0001 <0.0001 0.5093 0.036 0.0031 0.3685 0.0054 0.8603 0.1282 0.0008 0.8712 0.1218 0.0882
<0.0001 <0.0001 0.2696 0.6498 0.6562 0.5324 0.4088 0.4867 0.7257 0.4717 0.0333 0.9822 0.361 0.5698
<0.0001 <0.0001 0.2602 0.7032 0.6476 0.5074 0.4217 0.5121 0.7294 0.474 0.041 0.9774 0.3729 0.5977
0.0851 0.0019 <0.0001 0.3517 0.0379 0.0122 0.9957 0.0005 0.1935 0.7586 0.0152 0.5574 0.3193 0.3516
<0.0001 0.0263 <0.0001 0.0049 0.3949 0.4125 0.5975 0.0057 0.629 0.3341 0.6518 0.6713 0.8772 0.0844
<0.0001 0.0205 <0.0001 0.002 0.6899 0.5993 0.1714 0.0088 0.6451 0.4484 0.7778 0.7849 0.8106 0.5669
<0.0001 0.1987 0.0188 0.6046 0.6251 0.3557 0.1124 0.0773 0.9375 0.5301 0.7532 0.3686 0.834 0.9714
Electrical conductivity. Total nitrogen. Organic carbon. Organic matter. Carbon/nitrogen ratio. Bacteria population density. Fungal population density. Actinomycetes population density. Compost/water ratio. Extraction time. Storage duration. Storage temperature.
CWR 1:2.5
Fig. 1. Effect of the interaction among compost/water ratio (CWR) 1:2.5, extraction time and storage duration on compost tea total N.
CWR 1:2.5
ratio (CNR) of CT. The highest CNR (13.0) was obtained after 6 days of extraction time and 0 week of storage, while the lowest one was recorded after 4 days of extraction time and 2 weeks of storage. The changes in the CNR reflect OM decomposition and depends on the cumulative effect of OC and TN that were previously described. The estimated equation for compost/water ratio ⁄ extraction time ⁄ storage duration interaction on carbon to nitrogen ratio is: CNR = 10.367 CWR ⁄ 0.198 ET ⁄ 0.096 SD ⁄ 0.207 (R2 = 0.33, p < 0.0001, n = 90). CNR was mainly affected by storage duration possibly because of comparatively higher loss of carbon than nitrogen during the storage period as CO2 released from the microbial activity. 3.2. Microbial properties of compost tea The interaction between extraction time and storage duration on bacteria population density (BPD) resulted the sole significant combination (Table 3), and is illustrated in Fig. 4. The highest
CWR 1:10
Fig. 2. Effect of the interaction among the compost/water ratio (CWR) 1:2.5, extraction time and storage duration on the organic carbon content of the compost tea.
The same trend was obviously obtained for the organic matter (OM) of CT (Table 3). Fig. 3 depicts the interaction between extraction time and storage duration, at compost/water ratio of 1:10, on carbon/nitrogen
Fig. 3. Effect of the interaction among compost/water ratio (CWR) 1:10 and extraction time (ET) and storage duration (SD) on the C/N ratio (CNR) of compost tea.
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M.K. Islam et al. / Waste Management xxx (2016) xxx–xxx
BPD (6.43 log CFU L 1) was obtained after 4 days of extraction with zero week of storage duration. With the increase of storage duration, BPD decreased at variable rates depending on interaction and the lowest BPD (5.77 log CFU L 1) was obtained after 6 days of extraction with 4 weeks of storage. The compost/water ratio significantly affected the BPD (Table 3) since the average BPD recorded from compost/water ratio of 1:2.5, 1:5 and 1:10 were 6.15, 6.04 and 5.93 log CFU L 1, respectively. In addition, the storage temperature influenced significantly the BPD. In fact, the BPD was 6.09 log CFU L 1 at cool temperature and reduced to 5.99 log CFU L 1 at room temperature. These findings agree with Ingham (2005) who reported that there is a balance between extraction of nutrients and growth of organisms, giving an optimal time for tea production, depending on the extract conditions of brewing. The estimated equations for extraction time ⁄ storage duration interaction on bacteria population density is BPD (log CFU L 1) = 6.295 ET ⁄ 0.004 SD ⁄ 0.017 (R2 = 0.39, p < 0.0001, n = 90). The storage duration affect the bacterial activity possibly due to the consumption of nutrients, microbial competition and production of toxic molecules. Fig. 5 shows the significant interaction (Table 3) between extraction time and storage duration on fungal population density (FPD). It was observed that FPD decreased with increasing of extraction time and storage duration. The highest FPD (5.03 log CFU L 1) was obtained after 2 days of extraction with zero week of storage, whereas the lowest FPD (4.02 log CFU L 1) was recorded after 6 days of extraction with 4 weeks of storage. The compost/water ratio significantly influenced the FPD: the average FPD recorded were 4.62, 4.43 and 4.33 log CFU L 1 from compost/water ratio of 1:2.5, 1:5 and 1:10, respectively. The FDP was significantly affected by the storage temperature too: the FPD was 4.53 log CFU L 1 at cool temperature and reduced to 4.39 log CFU L 1 at room temperature. Both BPD and FPD increased from 1:10 to 1:2.5 compost/water ratio and at cool temperature. The estimated equation for extraction time ⁄ storage duration interaction on fungal population density is FPD (log CFU L 1) = 4.852 ET ⁄ 0.033 SD ⁄ 0.018 (R2 = 0.39, p < 0.0001, n = 90). As like as bacterial population, the storage duration affects negatively the FPD possibly because of the same reasons described above. According to results in Table 3, the compost/water ratio had significant effect on actinomycetes population density (APD). Means of APD recorded from compost/water ratio of 1:2.5, 1:5 and 1:10 were 5.85, 5.71 and 5.49 log CFU L 1, respectively. The storage duration significantly influenced the APD only after 4 weeks of storage, when the APD was found slightly lower. According to a previous study (Scheuerell and Mahaffee, 2006), nitrogen rich feedstock, such as compost containing manures,
Fig. 4. Effect of the interaction of storage duration and extraction time on the bacteria population density (BPD) of compost tea.
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Fig. 5. Effect of the interaction of storage duration and extraction time on the fungal population density (FPD) of compost tea.
produces a compost tea with higher bacterial content, respect to the fungal ones. Also Kim et al. (2015) reported that microbial communities of different aerated compost teas were predominantly bacteria.
3.3. Correlations Table 4 shows the correlation of compost/water ratio, extraction time, storage duration and storage temperature with CT quality parameters together with their significance probability. It is evident that the compost/water ratio showed significant (P < 0.001) correlation with EC, TN, OC and microbial populations. In particular, the chemical parameters were almost equally influenced by the dilution, while the actinomycetes population was more negatively affected by the dilution in comparison to bacteria and fungi. The storage duration had significant (P < 0.001) negative correlation with pH, CNR bacterial and fungal populations, while the actinomycetes population showed a lower significance (P < 0.05). Apparently, the microbial populations have contributed to the reduction of the pH with their metabolism, while their survival has been reduced significantly with time. The storage temperature showed a significant correlation only with fungal population since it has suffered for the room temperature storage in comparison to the bacteria and actinomycetes.
4. Discussion Results of hundreds of analyses indicated that compost/water ratio significantly affected compost tea electrical conductivity, total nitrogen, organic carbon and matter, and microorganisms densities with an increasing of these parameters from 1:10 to 1:2.5 compost/water ratio. Among several compost/water ratios from 1:2 to 1:50 (v/v), St. Martin (2014) showed that the denser CTs were more effective towards different pathogens. In addition, Scheuerell and Mahaffee (2004) and Cayuela et al. (2008) found that dilution decreased disease suppression of Pythium dampingoff of cucumber and Pythium capsici, respectively. Further, Evans et al. (2013) found positive effects of a manure compost tea obtained at compost/water ratio of 1:3 after 48 h of fermentation on the pathogen Botrytis cinerea. About the fertilizing effect of CT, Wang et al. (2014) reported no effect on the yield of zucchini using a chicken manure based vermicompost tea (1:10, v/v, continuously aerated for 12 h) as fertilizer. Also Hewidy et al. (2015) showed no effect on broccoli yield of a CT obtained from the organic fraction of the municipal solid wastes and pruning residues (1:5, v/v, continuously aerated for 24 h).
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Table 4 Pearson correlation coefficient (r) of CWR, ET, SD and ST with CT quality parameters. Bold figures indicate significant P-values. Parameters CWR ETj SDk STl *** * NS a b c d e f g h i j k l
i
ECa
pH NS
0.098 0.056NS 0.741*** 0.149NS
TNb ***
0.976 0.110 NS 0.027NS 0.000NS
OCc ***
0.936 0.101NS 0.093NS 0.010NS
OMd ***
0.933 0.135NS 0.035NS 0.010NS
CNRe ***
0.929 0.134NS 0.040NS 0.015NS
BPDf NS
0.098 0.188NS 0.534*** 0.050NS
FPDg ***
0.363 0.024NS 0.656*** 0.190NS
APDh ***
0.396 0.184NS 0.595*** 0.231*
0.748*** 0.127NS 0.215* 0.035NS
P < 0.001. P < 0.05. P P 0.05. Electrical conductivity. Total nitrogen. Organic carbon. Organic matter. Carbon/nitrogen ratio. Bacteria population density. Fungal population density. Actinomycetes population density. Compost/water ratio. Extraction time. Storage duration. Storage temperature.
Significant decrease of total N, organic carbon and organic matter contents of compost tea were observed when the extraction time increased, together with a slight reduction of bacterial and fungal populations. Ingham and Alms (1999) reported that the optimum extraction time is usually between 18 and 36 h, when the microbial biomass activity is the highest. St. Martin and Brathwaite (2012) showed that brewing cycles of 18 h for aerated compost tea can be used to produce CT with optimal Pythium ultimum in vitro inhibitive properties using lawn clipping and banana leaf composts. It was observed that the storage duration did not significantly influence the chemical properties of compost tea, with the only exception of pH and total N. In contrast, fungi and bacteria populations appeared more influenced from the storage duration since they decreased with increasing the storage duration possibly because of the competition for nutrients and oxygen among microorganisms and the release of metabolic toxic molecules. The same trend, but at lesser extent, was observer for actinomycetes populations. Long storage times negatively impacts upon the diversity of microorganisms, as well as nutrients carried by the tea for plant use (Bess, 2000). Number and activity of organisms reduces significantly with storage and, although this reduction is acceptable for a soil application of compost tea, it is not acceptable for a foliar application of compost tea (Ingham, 2005). McQuilken et al. (1994) found that the age of CT had some effects on its subsequent activity against both germination and mycelial growth of B. cinerea. In particular, a 3–12 day old CT still reduced the conidial germination, whereas a >18 day old CT significantly declined its inhibition properties. In addition, a 3–5-day old CT showed the highest inhibition against the mycelia growth. The storage temperature influenced the fungal and bacterial populations and, at a lesser extent, the pH of compost tea. To our knowledge, no study has been done in relation to the compost tea storage temperature, even if the tea temperature is considered very important for enzyme activity. In particular, at temperatures below the optimum, enzymes do not catalyze any reaction and membranes solidify, whereas at temperatures above the optimum, denaturation of enzymes, transport carriers and proteins occurs (Madigan et al., 2015). Many interactions and individual effect of the studied four factors (compost/water ratio, extraction time, storage duration and storage temperature) on compost tea quality parameters were found significant and those were elaborated by regression equations. Significance tests of predicted model were performed in
comparison with actual data. Test of significance of individual factor or interaction factors at individual level revealed that many quality parameters of compost tea can be predicted by compost/ water ratio and extraction time. According to the results, the best combination between chemical and microbial properties of CT can be obtained using a compost/water ratio of 1:2.5 and an extraction time of two days. If the CT production is devoted to exploit solely one or more microbial features, no storage should be applied since the microbial populations of CT decline with time. Therefore, the CT should be prepared and immediately used in agriculture, otherwise a cool storage is necessary to preserve the microbial populations of CT. However, the sprinkler and drip irrigation systems clog when a very dense CT is used for fertigation. It would be better to utilize surface irrigation systems or, if possible, to adapt slurry spreaders for soil application, and wet somehow the aerial parts of crops for the foliar application. Finally, further studies are needed to expand the validity of the proposed equations for different compost source and different maturity level, and to evaluate the performance of extracted compost tea on field crops. Acknowledgements Authors are very grateful to Prof. Nicola Senesi for his help to improve the quality of the manuscript. Authors wish to thank Maria Giuseppina Borracci for her moral support. References Adl, S., Iron, D., Kolokolnikov, T., 2011. A threshold area ratio of organic to conventional agriculture causes recurrent pathogen outbreaks in organic agriculture. Sci. Total Environ. 409, 2192–2197. Al-Dahmani, J.H., Abbasi, P.A., Miller, S.A., Hoitink, H.A.J., 2003. Suppression of bacterial spot of tomato with foliar sprays of compost extracts under greenhouse and field conditions. Plant Dis. 87, 913–919. Badgly, C., Perfetto, I., 2007. Can organic agriculture feed the world? Renew. Agric. Food Syst. 22, 80–85. Bess, V.H., 2000. Understanding compost tea. Biocycle 41 (10), 71–72. Biao, X., Xiaorong, W., Zhuhong, D., Yaping, Y., 2003. Critical impact assessment of organic agriculture. J. Agric. Environ. Ethics 16, 297–311. Cantisano, A., 1998. Compost teas. Organic Ag Advisors Letter. Colfax, California. Casado, G.I.G., de Molina, M.G., 2009. Preindustrial agriculture versus organic agriculture: the land cost of sustainability. Land Use Policy 26, 502–510. Cayuela, M., Millner, P., Meyer, S., Roig, A., 2008. Potential of olive mill waste and compost as biobased pesticides against weeds, fungi, and nematodes. Sci. Total Environ. 399, 11–18.
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Please cite this article in press as: Islam, M.K., et al. Effects of the main extraction parameters on chemical and microbial characteristics of compost tea. Waste Management (2016), http://dx.doi.org/10.1016/j.wasman.2016.03.042