Effects of a Delayed Harvest and Additives on the Fermentation Quality of Corn Stalk Silage

: This study investigated the effects of harvest time, cellulase, lactic acid bacteria, and ensiling with alfalfa hay on the chemical parameters, bacterial community composition, and fermentation of corn stalk silage. Corn stalks were ensiled after the ear harvest at 0 days (D0) and 20 days (D20). Each harvest was treated with alfalfa hay [AL, 5% of fresh matter (FM)], Lactobacillus plantarum (Lp) (1 × 10 6 CFU/g of FM), Trichoderma cellulase (TC) (100 mg/kg of FM), or both Lp and TC (Lp+TC). The silos were opened after 60 days of fermentation. The dry matter (DM) content of raw materials increased from 18.79% to 28.32% with the harvest time. The acid detergent fiber (ADF) content also significantly increased ( p < 0.05), while water-soluble carbohydrates (WSCs) and in vitro dry matter digestibility (IVDMD) significantly decreased ( p < 0.05). After 60 days, the neutral detergent fiber (NDF) and ADF contents increased, and all chemical composition and fermentation state parameters of silages had significant differences ( p < 0.05) with the harvest time. Compared with the control group, the AL treatment resulted in higher ( p < 0.05) DM and crude protein (CP) contents, and the Lp+TC treatment resulted in a lower NDF ( p < 0.05) content. For D0, the TC and Lp+TC treatments resulted in a higher WSC content and lower NDF and ADF contents than in D20. Meanwhile, the ammonia-nitrogen (NH 3 -N) content in each treatment was less than 10% of the DM. The pH values were all approximately 3.70. The AL treatment significantly increased the lactic acid (LA) content and lactic/acetic acid ratio (LA/AA). Additionally, the silages had a similar microbial environment before and after fermentation. We recommend harvesting corn stalks for silage immediately after the corn ear harvest. Adding cellulase improved the corn stalk feed value, and ensiling with alfalfa hay improved its fermentation quality.


Introduction
Corn is considered a crucial forage grain crop with diverse applications, ranging from consuming corn ears to utilizing stalks as animal fodder and producing derivative products through extensive processing.Worldwide corn production has been abundant, reaching 1151.36 million metric tons by the year 2022 [1].This substantial corn yield results in a significant generation of corn stalks.Unfortunately, many farmers opt to burn corn stalks, leading to resource wastage, air pollution, and exacerbating environmental challenges [2].
While corn stalks are abundant, their complex structure, composed of cellulose, lignin, and hemicellulose tightly bound by covalent bonds [3], presents challenges in using them as feed [4].This results in a rough texture, poor palatability, and low digestibility.Enhancing the effective utilization of corn stalks as feed becomes crucial, particularly in the winter when forage for herbivores becomes scarce, and animals compete with humans for grain for food [5].
Ensiling is an effective preservation method that can improve forage palatability, digestibility, and nutrient retention for long-term storage [6].However, leaving corn stalks in the field, a common practice, leads to nutrient loss [7].Previous studies have explored the use of additives, such as cellulase and lactic acid bacteria (LAB), to enhance corn stalk silage quality [8,9].Cellulase has become a popular additive for silage in recent years [10].It can convert cellulose to glucose, which is better for silage fermentation and can be more easily digested.Lactic acid bacteria (LAB) have also been used as additives in many studies [11][12][13].They can convert carbohydrates in raw materials into lactic acid, lower the pH, and inhibit the growth of harmful microorganisms, thereby achieving long-term preservation and maintaining nutrients.At the same time, the activity of microorganisms can improve palatability.Alfalfa hay is frequently used in mixed ensiling due to its rich protein content and good palatability.It can improve the moisture of raw materials, which is an important issue in silage making.
While many studies have focused on additives and mixed ensiling, the impact of harvest time and ensiling duration on corn stalk silage quality remains underexplored.Compared to whole-plant corn silage, corn stalks, often considered waste, present an opportunity to improve feed selectivity and environmental sustainability.Identifying the optimal time for harvest, silage-making, and storage is crucial for farmers [14].Therefore, our study aims to investigate the effects of harvest time, cellulase, lactic acid bacteria, and mixed ensiling with alfalfa on the chemical parameters, bacterial community compositions, and fermentation characteristics of corn stalk silage.
Approximately 400 g of chopped corn stalks treated with additives were placed into plastic bags and sealed with a vacuum packer (QV-100, BlueTec Co., Ltd., Ansong City, Gyeonggi-do, Republic of Korea).Treatments were replicated three times.The D0 and D20 silages were opened after 60 days.Samples of each silage were then taken to analyze the quality of the chemical parameters, bacterial community composition, and fermentation characteristics.

Chemical and Microbiological Analysis
The dry matter (DM) content of fresh samples and silages was determined by drying in an oven at 65 • C for 72 h.Then, the dried samples were passed through a 0.20 mm sieve (Thomas Scientific, Inc., Swedesboro, NJ, USA) and ground into a powder for chemical analysis.Water soluble carbohydrate (WSC) content was determined by modifying the anthrone method [17].Crude protein (CP) content was analyzed according to the Association of Official Agricultural Chemists (AOAC) method [18].Acid detergent fiber (ADF) and neutral detergent fiber (NDF) contents were determined using an Ankom2000 fiber analyzer (Ankom Technologies, Inc., Fairport, NY, USA) following the method of Van Soest et al. [19].In vitro DM digestibility (IVDMD) was determined by placing the nylon sample filter bags (50 × 55 mm, ANKOM F57, ANKOM Tech., Fairport, NY, USA, 0.5 ± 0.09 g/sample) into bottles with buffer solutions and ruminal gastric juice from cows, and keeping them in a digester at 39 • C for 48 h.Samples were then analyzed using the same method as that used for the NDF analysis [20].
The forage nutritive value was expressed as the total digestible nutrient (TDN) and relative feed value (RFV), which were calculated as follows [21]: TDN: TDN% = 88.9 − 0.79 × ADF% RFV: DDM% = 88.9 − 0.779 × ADF% DMI% = 120/NDF% RFV = (DMI% × DDM%)/1.29 Next, 10 g of fresh silage samples were homogenized in 90 mL sterile saline (0.85% NaCl solution), shaken for 1 h in a mechanical shaker, and refrigerated for 24 h.Samples were then filtered through a filter paper (Whatman No. 1, AVANTE C, Toyo Roshi Kaisha, Ltd., Tokyo, Japan).The extract was serially diluted 10 2 −10 5 times with 0.85% NaCl, and 10 µL of each extract was evenly spread on an agar surface using a plastic spreader followed by incubation.The LAB number was counted after growing on De Man, Rogosa, and Sharpe agar (MRS) medium incubated at 37 • C for 24−48 h.Total microorganisms (TMs) were counted on plate count agar (PCA) medium incubated at 37 • C for 48−72 h.Mold and yeast were counted on potato dextrose agar (PDA) medium incubated at 25 • C for 48−72 h.After incubating, the colony-forming units per gram (cfu/g) of microorganisms were counted on the agar plates, calculated according to the dilution factor, and converted to a log 10 value.

Statistical Analysis
This study had one control and four treatment groups, which were analyzed using statistical methods.The data were analyzed as a 5 × 3 factorial arrangement in a completely randomized design.Further analysis was conducted using a general linear model (GLM) in Statistical Product and Service Solution (SPSS) version 26.The fixed effects of harvest time and additives were analyzed via one-way ANOVA, and their interaction was analyzed via univariate two-way analysis.The differences among treatments were determined using a least significant difference (LSD) test with a significant difference at p < 0.05.All figures were produced using GraphPad Prism 8 version 8.0.2 (263) software.

Microbial Counts and Chemical Composition of Raw Materials
The microorganism counts and chemical composition of corn stalks are presented in Tables 1 and 2. The LAB, TM, and yeast and mold (YM) counts were in the ranges of 5.47 to 5.39, 5.73 to 5.41, and 3.50 to 3.65 log 10 cfu/g FM, respectively.The LAB and YM counts were not significantly affected by harvest time (p > 0.05), while the TM count in D0 was significantly higher than that of D20 (p < 0.05).Harvest time had effects on DM, WSC, ADF, and IVDMD contents significantly (p < 0.05).The DM content was averaged from 18.79% to 28.32%, and the WSC content of corn stalks was greater than 13.0% of DM on average at D0 and greater than 12.0% of DM on average at D20.All corn stalks had a CP content greater than 6.0% of DM, while the AL treatment had the highest CP content.The NDF content was from 47.18~50.35% of DM at D0 and from 47.74~51.43% of DM at D20. ADF content was higher at D20 than at D0, which was 28.72% of DM on average.Meanwhile, D0 had higher IVDMD content than D20 (averaged from 72.44% of DM to 66.65% of DM).

Chemical Composition of Corn Stalk Silage
The chemical compositions of corn stalk silages treated with additives for 60 days are shown in Table 3.After 60 days of ensiling, there were significant differences (p < 0.05) in all chemical parameters due to the harvest time, additives, and their interactions, except the ADF contents; the additives had no significant effect(p = 0.05) on it, which averages 32.99 at D0 and 33.30 at D20.Additionally, also the CP content showed no significant differences due to interactions (p > 0.05).The DM, NDF, and ADF contents increased significantly with harvest time (p < 0.001), while the WSC, CP, and IVDMD contents decreased significantly (p < 0.001).Compared to the raw materials, the DM content was reduced (average from 18.79% to 17.17% at D0 and from 28.32% to 26.09% at D20), the WSC content declined sharply, more than 85.0%, and the CP content also decreased.Moreover, the digestibility of the feed deteriorated, with an increase in the NDF and ADF contents and a decrease in the IVDMD content.After the ensiling process, the effects of additives on the chemical compositions of D0 and D20 displayed similar changes.Compared with the control group, the AL treatment had a higher CP content, which was 7.56 at D0 and 6.77 at D20, the TC treatment had a higher DM content, and the Lp+TC treatment had a higher WSC content and lower NDF and ADF contents.The TDN content and RFV of corn stalk silage are shown in Figures 1 and 2, respectively.D0 had higher TND and RFV values than D20, while the additives resulted in no significant difference compared to the control group.The fermentation indicators, pH, and the NH 3 -N and organic acid contents are shown in Table 4.There were significant differences in the NH 3 -N, LA, and LA/AA contents with a delayed harvest time, and there were significant differences in pH and NH 3 -N, LA, and LA/AA contents when additives were supplied.There were also significant differences in pH and the NH 3 -N and LA contents due to the interactions between delayed harvest time and additives.The additive treatments had a similar pH at D0 and D20, which were all around 3.70.The NH 3 -N content was significantly increased with harvest time (p < 0.001), with average 4.50% of total N at D0 and 7.37% of total N at D20, while the LA and LA/AA contents decreased (p < 0.001).The NH 3 -N content was lower in the TC treatment than in other treatments (p > 0.05).The effect of additives on LA and the LA/AA ratio was similar at D0 and D20, with the AL treatment having the highest content (3.64% of DM, 6.28 at D0 and 2.98% of DM, 4.89 at D20, respectively) but the differences with the other treatments were not significant.

Microbial Population of Corn Stalk Silage
The microbial composition of corn stalk silage on D0 and D20 is shown in Figure 3.At the same harvest time, there were no significant differences in the microorganism populations under the different additive treatments.There were significant differences in the LAB counts for the AL, Lp, and Lp+TC treatments at D0 and D20, with larger LAB populations in the AL, Lp, and Lp+TC treatments at D0 than at D20.Neither additives nor harvest time had a significant effect on the YM populations.For the TM count, only the Lp treatment had a significant difference in D0 and D20, with a higher population at D20.

Microbial Population of Corn Stalk Silage
The microbial composition of corn stalk silage on D0 and D20 is shown in Figure 3.At the same harvest time, there were no significant differences in the microorganism populations under the different additive treatments.There were significant differences in the LAB counts for the AL, Lp, and Lp+TC treatments at D0 and D20, with larger LAB populations in the AL, Lp, and Lp+TC treatments at D0 than at D20.Neither additives nor harvest time had a significant effect on the YM populations.For the TM count, only the Lp treatment had a significant difference in D0 and D20, with a higher population at D20.

Microbial Population of Corn Stalk Silage
The microbial composition of corn stalk silage on D0 and D20 is shown in Figure 3.At the same harvest time, there were no significant differences in the microorganism populations under the different additive treatments.There were significant differences in the LAB counts for the AL, Lp, and Lp+TC treatments at D0 and D20, with larger LAB populations in the AL, Lp, and Lp+TC treatments at D0 than at D20.Neither additives nor harvest time had a significant effect on the YM populations.For the TM count, only the Lp treatment had a significant difference in D0 and D20, with a higher population at D20.

Chemical Composition, Digestion Index, and Microbial Population of Corn Stalks and Their Silages
Increasing the development and use of non-grain feed resources, such as grass and stalks, is an effective measure to resolve the conflict between the development of animal husbandry and human competition for grain [24].Corn stalks are the by-products of corn after the ears are consumed.If they can be used effectively, both economic and environmental benefits could be achieved.Farmers usually harvest the corn ears before they are fully mature (more reproductive) to ensure the best flavor, but the harvest date

Chemical Composition, Digestion Index, and Microbial Population of Corn Stalks and Their Silages
Increasing the development and use of non-grain feed resources, such as grass and stalks, is an effective measure to resolve the conflict between the development of animal husbandry and human competition for grain [24].Corn stalks are the by-products of corn after the ears are consumed.If they can be used effectively, both economic and environmental benefits could be achieved.Farmers usually harvest the corn ears before they are fully mature (more reproductive) to ensure the best flavor, but the harvest date for the remaining stalks is not certain.It is, therefore, important to determine the optimal stalk harvest date to produce good silage and to establish methods that can improve the quality of stalk silage.
Previous studies have shown that leaving stalks in the field after the ear harvest could decrease the CP content, increase the DM and fiber contents of the stalks, and reduce the dry matter digestibility [16,[25][26][27].The DM content in stalks left in the field has been shown to increase by more than 50%, which may be due to the ears of corn harvested at an earlier stage being more vegetative and the corn stalks still growing to some extent after the ears were harvested [28].The CP content decreased after 60 days ensiling.In the fresh stalk (D0) silage, it decreased by nearly 5.0%, while in the D20 silage, it was nearly 16.0% lower.This was consistent with the results of previous studies.It has been reported that the CP content decreases after ensiling [29,30], although in some good-quality silages, the CP content can be maintained without significant changes [31].In this study, the silages from the D0 treatment groups maintained their CP content well, while there was an obvious loss in the D20 treatment groups, indicating that ensiling fresh stalks immediately could maintain their nutritional content and produce higher quality silage.In previous studies, the water loss of corn stalks and the proportion of DM increased significantly after field exposure; the NDF and ADF contents also increased sharply [28].Our results were similar.Compared with the D0 stalks, the D20 stalks had higher NDF and ADF contents and a lower IVDMD content.
The LAB and WSC contents play important roles in the fermentation process before ensiling.The LAB counts in this study ranged from 5.39 to 5.47 log 10 cfu/g FM.Guo et al. (2021) [25] also reported similar counts, indicating that harvest time could reduce the fermentation quality of corn stalk silage.This was also evidenced by the TM population, which decreased significantly in D20 stalks.The WSC content, which ranged from 6.0% to 8.0% of DM, was more likely to produce high-quality silage [32].In this study, the average WSC content was 11.96 to 13.09% of DM and, therefore, contributed to the production of good silage.After ensiling, the LAB population only increased in the D20 silages, while the YM population decreased, but it was similar in D0 and D20.Furthermore, additives had no significant effect on the various bacterial populations in the silages.These were similar to the results of Sun et al. (2019) [33].It may be due to LAB dominating the microbial population of raw materials.The raw materials contained high WSC and LAB contents, enabling strong fermentation effects to be achieved even without additives.LAB produce a large amount of lactic acid at the beginning of fermentation, creating an acidic environment, thereby inhibiting the growth of mold, yeast and other harmful bacteria.If the epiphytic LAB of corn stalks are homofermentative LAB, it will promote the dominance of homolactic fermentation and achieve a better silage effect [34].
Using alfalfa hay as an additive to mix with corn stalks for ensiling can not only increase the CP content of corn stalks but also realize the ensiling possibility of alfalfa.In terms of chemical compositions, the alfalfa treatment significantly increased the CP content, but there were no significant differences with the control group for the other parameters.

pH and Organic Acid and NH 3 -N Contents of Corn Stalk Silage
Among the different fermentation states, the pH of all the treatments was similar, but all pH values were less than 4.2.As reported by Guo et al. (2021) [25], successful silage requires a pH lower than 4.2.Guan et al. [35] said that the rapid decrease in pH value is caused by the proliferation and fermentation of lactic acid bacteria to produce more lactic acid, thereby inhibiting the growth of harmful bacteria and forming a highquality fermentation environment.The high WSC content in the raw materials can provide enough nutrition for LAB so that lactic acid fermentation can dominate at the beginning of fermentation and drop the pH rapidly.Therefore, the microbial community and WSC content in the raw materials will have an impact on pH [36].If the fermentation microbial communities in different treatment groups are similar, similar pH changes will result.In this experiment, there were similar counts of microorganisms before ensiling, and the WSC content in raw materials was high, which may be the reason why the pH values of each treatment group were similar.McDonald (1991) [37] reported that the NH 3 -N content was less than 10% of the total N, indicating that the silage preserved the N content, and NH 3 -N is an iconic parameter for indicating protein degradation.In this study, the AL treatment had the lowest NH 3 -N content in both the D0 and D20 silages, indicating their proteolysis was relatively low.The NH 3 -N content in the D20 silage was nearly twice that of the D0 silage, which indicated that the fermentation effect of D0 was better than that of D20.This was also confirmed using the LA content, which was higher in D0 silage.
Both LA and AA are important indicators of the quality of silage.During the fermentation process, LA contributes most to the decrease in pH.Kilic (1986) [38] reported that an LA content higher than 2.0% on a DM basis was required to produce high-quality silage.The higher the LA content, the better the quality of silage, while AA has an inhibitory effect on fungi and a positive effect on aerobic stability [37].The LA/AA is an indication of homolactic dominant fermentation.If the LA/AA content is more than 3, the fermentation process is homolactic dominant fermentation [39].In this study, the LA content and LA/AA ratio in the AL treatment were significantly higher than in the control group, indicating that the AL treatment group had better fermentation than the control group.This may be due to the addition of alfalfa hay, which controlled the moisture content of the raw material, allowing it to achieve better silage results [40].
Silage inoculated with LAB is considered an effective method to improve the quality of silage fermentation [15].Epiphytic LAB occurs naturally in forage crops, participates in silage fermentation, and has an impact on silage quality.The LAB count determines if microbial additions to raw materials are required [33].The homofermentative LAB in corn silage uses glucose during fermentation to produce LA and some AA, formic acid (FA), and PA, and therefore, the pH drops rapidly to approximately 4.0, thereby inhibiting the activities of other microorganisms [41].When the pH drops to 3.8, the growth of LAB is also inhibited, and the biochemical processes in the silage are stopped.
If LAB cannot quickly become the dominant flora during fermentation, a large number of spoilage microorganisms will grow rapidly and affect the quality of silage [42].Therefore, adding LAB to silaged raw materials can increase the LAB population in silage raw materials, enabling LAB to quickly become the dominant flora and thereby improving the nutritional value and quality of silage [15].In this study, the pH value of the LAB treatment was less than 4.2, the NH 3 -N content was less than 10%, the LA content was greater than 2% on a DM basis, and the LA/AA ratio was higher than 3.This indicated that the silage quality resulting from the LAB treatment was good.However, compared with the control group, it did not significantly improve the fermentation effect, nor was it significantly different from the control group in terms of chemical composition.This was similar to the results of Khota et al. (2018) [43]; this may be because the effect of the natural epiphytic LAB population on the raw materials was large enough to allow homolactic fermentation to occur at the beginning of the fermentation process, thereby achieving a strong fermentation effect.Thus, for the raw materials in our study, LAB inoculation cannot improve silage quality, and it is therefore not necessary to add an LAB inoculation when making silage.
Cellulase is a frequently used silage additive.Li et al. (2018) [44] reported that the addition of cellulase can increase the amount of LAB in the substrate, and the mixeduse of cellulase and LAB could be an effective way to enhance fermentation.Cellulase acts on the β-1,4-glucosidic bonds in cellulose macromolecules to decompose them into monosaccharides or disaccharides, which are directly absorbed by animals and can improve feed utilization [45].In addition, more sugars can be released for fermentation by LAB via the action of enzymes that help break down carbohydrates into sugars [46].Our results showed that the NDF and ADF contents in the TC and Lp+TC treatments were lower than those in the control group, while the WSC content was much higher.This shows that the addition of cellulase decomposes the cellulose in corn stalks into WSCs, which can degrade fiber and improve the quality of silage.This is also supported by other studies [47].The NH 3 -N content in the TC treatment was lower than that in the control group (<10% DM), which indicated that the addition of cellulase was beneficial to the fermentation of silage, limited the decomposition of proteins, and better-preserved nutrients.Furthermore, the LA content (>2% DM) and LA/AA ratio (>3.0) in the TC and Lp+TC treatments reached the standard required for good silage, but their contents were not improved compared with the control group.The inability of these treatments to improve the fermentative properties of silage may be due to the inability of the inoculum to compete with the epiphytic microbial population in the raw material [48].

Conclusions
In this study, we found that all treatments improved the chemical composition value and resulted in good fermentation effects.However, delaying the harvest time of corn stalks resulted in a decrease in their chemical composition value and fermentation effects.Among the fresh stalk silage treatments, the TC and Lp+TC additives produced a better chemical composition value, while the AL treatment resulted in a better fermentation effect.Therefore, harvesting fresh corn stalks for silage immediately after a corn ear harvest could result in better chemical composition value and fermentation effects.In terms of the materials suitable for making silage, the additives tested here may not produce significant effects.However, if additives are recommended, they would be most effective for raw materials with a low nutritional content, such as the addition of cellulase or LAB and cellulase.For high-moisture raw materials, it is recommended to add hay additives, such as alfalfa hay.

Figure 1 .
Figure 1.Total digestible nutrients (TDN) of corn stalk silages treated with additives after 60 days.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; SEM, standard error of the mean.D0 and D20 indicated corn stalks at 0 and 20 days after ear harvest, respectively; bars labeled with * are significantly different (p < 0.05).

Figure 2 .
Figure 2. Relative feed value (RFV) of corn stalk silages treated with additives after 60 days.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; SEM, standard error of the mean.D0 and D20 indicated corn stalks at 0 and 20 days after ear harvest, respectively; bars labeled with * are significantly different (p < 0.05).

Figure 1 .
Figure 1.Total digestible nutrients (TDN) of corn stalk silages treated with additives after 60 days.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; SEM, standard error of the mean.D0 and D20 indicated corn stalks at 0 and 20 days after ear harvest, respectively; bars labeled with * are significantly different (p < 0.05); NS means no significant.

Figure 1 .
Figure 1.Total digestible nutrients (TDN) of corn stalk silages treated with additives after 60 days.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; SEM, standard error of the mean.D0 and D20 indicated corn stalks at 0 and 20 days after ear harvest, respectively; bars labeled with * are significantly different (p < 0.05).

Figure 2 .
Figure 2. Relative feed value (RFV) of corn stalk silages treated with additives after 60 days.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; SEM, standard error of the mean.D0 and D20 indicated corn stalks at 0 and 20 days after ear harvest, respectively; bars labeled with * are significantly different (p < 0.05).

Figure 2 .
Figure 2. Relative feed value (RFV) of corn stalk silages treated with additives after 60 days.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; SEM, standard error of the mean.D0 and D20 indicated corn stalks at 0 and 20 days after ear harvest, respectively; bars labeled with * are significantly different (p < 0.05).

Figure 3 .
Figure 3. Microbial composition of corn stalk silages treated with additives after 60 days.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; TM, total microorganisms; YM, yeasts and molds; C = control; AL = alfalfa; LP = Lactobacillus plantarum; TC = Trichoderma cellulase; LP+TC = L. plantarum + T. cellulase; and SEM, standard error of the mean.D0 and D20 indicate corn stalks at 0 and 20 days after ear harvest; bars labeled with * are significantly different (p < 0.05); bars labeled with ** are significantly different (p < 0.001); NS means no significant; and different superscripts in different letters indicate that additives differ (p < 0.05).

Figure 3 .
Figure 3. Microbial composition of corn stalk silages treated with additives after 60 days.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; TM, total microorganisms; YM, yeasts and molds; C = control; AL = alfalfa; LP = Lactobacillus plantarum; TC = Trichoderma cellulase; LP+TC = L. plantarum + T. cellulase; and SEM, standard error of the mean.D0 and D20 indicate corn stalks at 0 and 20 days after ear harvest; bars labeled with * are significantly different (p < 0.05); bars labeled with ** are significantly different (p < 0.001); NS means no significant; and different superscripts in different letters indicate that additives differ (p < 0.05).

Table 1 .
Microorganism population of corn stalks before ensiling., corn stalks 0 and 20 days after the ear harvest, respectively; LAB, lactic acid bacteria; TM, total microorganisms; YM, yeasts, and molds; and SEM, standard error of the mean.Different superscripts in lower-case letters indicate that harvest time differs (p < 0.05).

Table 2 .
Chemical compositions of corn stalks treated with additives before ensiling.
H = harvest time; A = additives; H × A = harvest time × additives.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; DM, dry matter; WSCs, water-soluble carbohydrates; CP, crude protein; ADF, acid detergent fiber; NDF, neutral detergent fiber; IVDMD, in vitro dry matter digestibility; and SEM, standard error of the mean.D0 and D20, corn stalks 0 and 20 days after the ear harvest, respectively.Different superscripts in capital letters indicate that harvest time differs.

Table 3 .
Chemical compositions of corn stalks silage treated with additives after 60 days.
H = harvest time; A = additives; H × A = harvest time × additives.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; DM, dry matter; WSCs, water-soluble carbohydrates; CP, crude protein; ADF, acid detergent fiber; NDF, neutral detergent fiber; IVDMD, in vitro dry matter digestibility; and SEM, standard error of the mean.D0 and D20, corn stalks 0 and 20 days after the ear harvest, respectively.Different superscripts in capital letters indicate that harvest time differs.Different superscripts in lower-case letters indicate that additives differ (p < 0.05).

Table 4 .
pH and NH 3 -N and fermentation acid contents in corn stalk silages treated with additives after 60 days.
H = harvest time; A = additives; H × A = harvest time × additives.AL, corn stalk co-ensiling with alfalfa hay; Lp, corn stalk with the addition of lactic acid bacteria (LAB); TC, corn stalk with the addition of cellulase; Lp+TC, corn stalk with the addition of LAB and cellulase; NH 3 -N, ammonia nitrogen; LA, lactic acid, AA, acetic acid, PA, propionic acid, BA, butyric acid; ND, not detected and SEM, standard of error mean.D0 and 20 indicated corn stalks after 0 and 20 days of ear harvest.A,B within a column, different superscripts in capital letters indicate that harvest time differs; a-c those in lower-case letters indicate that additives differ (p < 0.05).Agriculture 2024, 14, x FOR PEER REVIEW 8 of 13