1. Introduction
As number of livestock has increased all around the world, the amount of animal manure has increased simultaneously. From 2006 to 2016, the number of total livestock increased from 293.48 to 374.99 million in Bangladesh [
1]. In China, the livestock population is estimated to be 9.03 million cattle and buffaloes, 0.61 million horses and donkeys, 44.06 million goats and sheep, and 74.66 million hogs in 2017 [
2]. However, the increasing amounts of the livestock wastes, such as feces and urine, were difficulty in disposing of as it induced extensive environmental pollution [
3,
4]. Besides, economical reason is also important for avoiding over-fertilization. Holm-Nielsen et al. [
5] have suggested that increasing animal production areas need suitable manure management and to optimize their recycling. For sustainable development, it is necessary to find out a cost-effective way to dispose of these wastes.
Anaerobic digestion is one of the effective solutions for livestock wastes and satisfies growing concerns on energy supply [
6,
7,
8]. Among the alternative energy generation technologies, anaerobic digestion is an efficient way to transform waste into resources and it benefits by improving fertilizer qualities, reducing odors and pathogens of livestock wastes. The main product of anaerobic digestion is biogas, which is an important and clean energy source. The biogas could be used for human activity like cooking or lighting, thereby replacing other fuels. Several research studies are reported on the utilization of farm waste as feedstock to produce biogas. In Malaysia, it was found that the amount of biogas could be generated for the year 2012 (4589.49 million m
3) from animal farm waste [
9]. In Thailand, approximately 620 million m
3 biogas can be generated from that animal waste [
10]. Similar kinds of studies were also conducted in Laos and Myanmar [
11]. Besides, biogas could be sent to a biogas upgrading system for the production of bio-methane further, which plays an important role in agriculture development [
12]. Meanwhile, the by-product biogas slurry is also an environmentally friendly organic fertilizer which could be used in agricultural production [
5,
13]. Recently, the use of biogas slurry has been recognized as an important kind of fertilizer owing to its high nutrient components, environmentally friendly and lower costs than chemical fertilizers [
14,
15]. It was reported that more than 450 million tons of biogas slurry have been used in China each year [
16] to popularize ecological circular agriculture, recycle agricultural wastes, reduce chemical fertilizer input, and protect the environment. However, the increasing popularity of anaerobic digestion has created another challenge of how to deal with large quantities of biogas slurry generated during this process. An 800 m
3 volume biogas system needs to discharge 15 tons of biogas slurry daily. Therefore, this condition will become an environmental issue if improperly managed.
As a kind of bio fertilizer, biogas slurry has plenty of nitrogen (N), phosphorus (P), potassium (K), and other trace elements. Further, biogas slurry could use as a biological pesticide due to its high levels of amino acids, as well as growth hormones, and antibiotics that promote plant growth [
17,
18,
19,
20,
21,
22]. It was reported that dry slurry contained lower levels of nitrogen (less than 0.5%), while the wet slurry contained more than 1.6% nitrogen as readily available nutrients. Based on the fermentation process, the concentration of carbon (C) from the dry matter reduced, while the ammonium ion (NH
4+) content of the biogas slurry increased, which induced the C/N ratio decreased [
23,
24]. Furthermore, biogas slurry supplies more plant readily available nitrogen than common chemical fertilizers [
25]. Generally, the available forms of N contained nitrate (NO
3), ammonium (NH
4+), and simple structured organic partly from the degradation of organic matter, which could directly be absorbed by plants. Besides, biogas slurry also supplied varied nutrients, including some micronutrients, auximones, vitamins, and humic acid, which could not only retain water as well as regulate soil fertility, soil environment, gas and heat, but were also beneficial to plant growth [
26,
27]. It had been reported that biogas slurry could efficiently be utilized for crops, such as
Triticum aestivum [
28],
Hordeum vulgare [
23],
Oryza sativa [
20,
29],
Allium porrum [
30],
Zea mays [
31],
Arachis hypogaea [
32], and
Brassica rapa [
33]. However, different standards are required for various crops based on plant nutrient utilization efficiency, soil conditions, as well as environmental release.
Italian ryegrass (lolium multiflorum L.) is one of the most important annual forage grasses worldwide and it has been cultivated for hay, grazing, or silage owing to its easy to establish, high productivity and palatability, excellent resistance to mow, and adapted to a wide range of soil types [
34,
35,
36]. Italian ryegrass was always planted and mixed with small grains and clovers (
Trifolium sp.) to provide forage for animals during winter and spring. Additionally, it is widely used for the cereal-forage rotation system or economic crop intercropping in south of China. The forage quality could directly influence the animal performance, while various factors directly or indirectly decided forage quality. Among them, the most key factors are forage species and stage of maturity at harvest. Followed, soil fertility, fertilization, and temperatures during forage growth period are all important [
37]. For grasses species, it is well-known that nitrogen fertilization directly contributes to the quantity and quality of forage production. However, inappropriate or excessive use of chemical fertilizers induced adverse effect on the soil, causing a decrease in organic carbon, as well as environment pollution [
38]. Additionally, Italian ryegrass can promote the purification of domestic sewage, aquaculture wastewater, and other sewage, and has good environmental, ecological, and economic effects [
39], which makes it become a good plant with absorbing biogas slurry to reduce environmental pollution.
Recently, more and more studies have focused on the effects of biogas slurry on crop biomass yield, soil properties, and fertility. Balance of the useage of biogas slurry is not only avoiding environmental risk of over-fertilization, but also for optimum plant growth. However, environmental factors were selected only as their treatment, and the integrated effects that related to crop growth, yield, and quality by applying biogas slurry are rarely investigated. Additionally, previous studies mainly focused on cereal crops or conventional vegetables, research on Italian ryegrass (lolium multiflorum) were reported rarely, which account for a large proportion of the total cool-season forages planting area [
34,
36]. In brief, there is little information available related to dealing with the use of biogas slurry and synthetic chemical fertilizer as an alternative to only synthetic fertilizer application in Italian ryegrass production. The objective of the study was to evaluate the effects of biogas slurry in combination with synthetic chemical fertilizer on biomass yield production and nutritional status of Italian ryegrass, and find the optimal ratio of synthetic fertilizer and biogas slurry to dissolve baogass slurry and reduce the synthetic fertilizer application. On the other hand, this study can provide basic information about that the balance of Italian ryegrass biomas yield and biogas slurry assimilating capacity.
4. Discussion
With the growing energy demand and inadequate natural resource availability, biogas technology has been considered a new renewable energy alternative, obtained by using cattle dung, poultry litter, and other wastes [
43]. However, with the synthesis of biogas through anaerobic digestion, solids and other wastes are released as slurry, which is commonly considered a waste product and potential caused environmental pollution. An optimized level of BS fertilizer is a good source of plant nutrients and can significantly improve soil properties and crop productivities [
44], providing an opportunity to treat and reutilize various organic wastes and minimize greenhouse gas emissions [
14,
45].
Analysis of variance (ANOVA) is the most efficient method available for the analysis of experimental data, and it is a technique for analyzing the way in which the mean of a variable is affected by different types and combinations of factors. One-way analysis of variance is the simplest form. It is an extension of the independent samples t-test [
46] and can be used to compare any number of groups or treatments. This method is widely used in agricultural research. Previous study showed that ANOVA was used to compare changes in the phenological properties, yield, and phytochemical composition of guar (
Cyamopsis tetragonoloba L.) landaraces under various irrigation regimes and planting dates [
47]. In the present study, we designed a randomized block experiment to compare the effects of CSF + BS on the biomass yield and forage quality of Italian ryegrass with ANOVA, which could have statistical meaning for agriculture application.
It has been proved that the application of biogas slurry as fertilizer could effectively stimulate the growth of plants and is the ideal substitute of chemical synthetic fertilizer. As a kind of biofertilizer with abundant nutrients, bioslurry has been reported to have significantly increased yields in
S. tuberosum L. [
48],
Spinacia oleracea L. [
49],
Capsicum annuum L. [
49],
Abelmoschus esculentus (L.) Moench [
50], and
Lactuca sativa L. [
51] compared with only the application of synthetic fertilizer. However, in order to obtain optimized growth and be environment friendly, the suitable application rate of biogas slurry should be study. Based on this study, approximately 37.5 kg/ha CSF + 100.5 t/ha BS is the optimum level for Italian ryegrass growth, and the yield greater than the yield of traditional farmer practice (only used the chemical synthetic fertilizer), which was consistent with previous study in tomato by Ferdous et al. [
52]. Marino et al. [
53] reported that the nitrogen fertilizer significantly increased the forage dry matter and N accumulation. Although Wentzel and Joergensen [
54] reported the positive effect of biogas slurry fertilizer on Italian ryegrass plant biomass, the biomass yield was non-significantly increased with the excess use of biogas slurry. Similarly, in order to obtain the maximum growth of maize fodder, an optimal level of biogas slurry (70 kg of slurry N ha
−1) was suggested to be applied, while excessive use beyond this optimal level might induce adverse effects [
27]. Other factors like genetic makeup of plant, soil fertility, and environment conditions were all important to plant height of forage.
It has been demonstrated that BS might increase the availability of macro- and micronutrients, particularly nitrogen, which in turn promotes meristematic growth, leading to higher bioactive compounds like anthocyanin and vitamin. BS is the rich source of nitrogen which holds prime importance since it is the key element in all proteins including enzymes which regulate all the metabolic processes in plants [
27]. Nitrogen also has a direct correlation with the important metabolic pathways such as growth, photosynthesis, and consequently affects the light energy efficiency and yield because it is the main element of leaf chlorophyll content [
55,
56]. The plant photosynthetic rate may be linear in correlation to leaf nitrogen content but the continuous lack of nitrogen reduces photosynthesis rate, soluble protein, enzymes, and chlorophyll [
57]. The observed increase in forage yield in response to slurry nitrogen might be owing to its positive effect on cell elongation, cell division, and increasing photosynthetic surface, resulting in more production and accumulation of photosynthetic compounds.
Usually, crops can directly use soil-available NH4 + -N, P, and K in biogas slurry [
8,
58]. Further, it has been reported that the use of biogas slurry in combination with chemical fertilizer could effectively increase soil organic matter, nutrient concentrations, and N use efficiency [
59]. The increase in nitrogen use efficiency could promote root and shoot growth, and increase the crop biomass yield [
52]. Katuwal and Bohara [
60] reported an increase in vegetable crop yields among 68% of farming households, along with an increase in revenue among 42% households after use of biogas slurry. Muhmood et al. [
49] reported that synthetic fertilizer in combination with biogas slurry applied could increase economic returns from spinach (
Spinacia oleracea) and chilli (
Capsicum frutescens) production. In this study, the results showed that the optimum level of biogas slurry for Italian ryegrass could benefit plant growth, and the yield, which was similar to those reported by Daudén and QuíLez [
61], who conducted a maize yield experiment using different levels of slurry. However, these studies mainly explored the effect of BS on yield, rather than both the yield and quality of productions.
Forage DM content is also an indicator of its quality according to variety, stage of forage maturity, season, temperature, and environment conditions. In the current study, due to soil fertilizer residues in the 1st growth season and the improvement of soil structure by biogas slurry, dry matter production in the 2nd growth season was higher than that of the previous year [
62]. Interestingly, the DM content of the second and third cutting were higher than other treatments results where the DM production of T4 and T5 in 2017 were higher than others. When the Italian ryegrass suffered drought stress during the second and third growth, the treatment of T4 and T5 can provide enough water for Italian ryegrass to grow. Besides, the CP and CF content of forage is the most important indicator contributing to its forage quality. Generally speaking, it has a direct effect on milk production in dairy animals and body growth in cattle. Sarker [
63] reported that the increased use of nitrogen fertilizer could effectively increase the CP content in Zamboo grass (
Hybrid jowar). The previous study of mixed sown pasture in North Wales indicated that application of digestate leads to a similar response in pasture yield as when compound inorganic fertilizer is applied, and better than straight N organic fertilizer [
64]. In this study, application of 37.5 kg/ha chemical synthetic fertilizer and 100.5 t/ha biogas slurry significantly increased the CP content. In addition, the CF content of Italian ryegrass decreased when the biogas slurry was used in combination with chemical fertilizer. The reason may be that biogas slurry had the potential to increase Italian ryegrass yield and improve the forage quality. Additionally, use of chemical fertilizer with biogas slurry could be a viable input for production of high yield and high quality forage. Similar results were also reported in switchgrass by the University of Life Sciences in Lublin where the biogas digestate can increase the biomass yield, the content of protein, P, and Mg of switchgrass [
65]. The present study provides guidance for the use of BS to grow Italian ryegrass, especially in the dairy farm and the developed area of animal husbandry. Italian ryegrass can provide feed for cow, and the cow dung is used to produce biogas to provide energy, and BS as a by-product can provide nutrients for Itanlian ryegrass. Due to an 800 m
3 volume, the biogas system needs to discharge 15 tons of biogas slurry daily; this circulation model can be applied on a large scale to reduce environmental pollution. The biogas slurry produced by a scale farm can be supplied to Italian ryegrass planting for animal feed. However, the biogas slurry transportation seems troublesome, and it is best to have pipeline laying as a cost-effective way, which sometimes requires government financial supply.
Biogas slurry used in crop cultivations is an agricultural eco-circulatory system for the reuse of livestock and poultry waste and reduction of chemical syntheic fertilizer input. Although this study explored the appropriate amount of biogas slurry to improve the yield of Italian ryegrass, there is much more to be learned. Uncertain chemical characteristics of biogas slurry including liquidity, high pH, high ammonium (NH4
+) content, and potentially high heavy metal content [
20,
29,
31], which are very different with traditional mineral fertilizers, could induce biology security risks. Thus, necessary evaluating studies must be performed to elucidate the effects of biogas slurry on crop development and environment before its extensive application. Especially, the accumulation of heavy metals in plants and soil to prevent them from affecting human health through the food chain.