Influences of Northern Leaf Blight on corn silage fermentation quality, nutritive value and feed intake by sheep
ABSTRACT
This study examined the differences between non-inoculated (control) corn and Northern Leaf Blight (NLB)-damaged corn (inoculated corn); dry matter (DM) yield, silage fermentation quality, nutritive value and feed intake by sheep were compared. Leaf, stem and grain dry weights and gross yield of inoculated corn were significantly (P < 0.05) decreased compared with control corn. The contents of water-soluble carbohydrate and nitrogen-free extract (NFE) were decreased in inoculated corn compared with control corn. Silage made from both inoculated and control corn showed good fermentation quality. The digestibility of DM, organic matter, ether extract, NFE, and energy of silage made from inoculated corn were significantly (P < 0.05) lower, and contents of total digestible nutrients (TDN) and digestible energy (DE) were also significantly (P < 0.05) lower compared with silage made from the control corn. DM intake showed no significant discrepancy between the two types of silage. TDN and DE intakes from inoculated silage were significantly (P < 0.05) lower compared with control silage. From the above results it was shown that NLB caused a decrease in DM yield and NFE content in corn and a decrease in the nutritive value and feed intake of silage.
INTRODUCTION
In Hokkaido the yield of whole-crop corn (Zea mays L.) was 202.6 million tons in 2007 (MAFF 2008), which was 44.6% of the total yield in Japan. Corn silage is commonly used because of high-quality fermentation, its taste and high nutritive value.
In the course of planting corn, Northern Leaf Blight (NLB) (Exserohilum turcicum Pass.; teleomorph: Setosphaeria turcicum) is a major plant diseases of corn alongside insect pests. According to a Hokkaido Plant Protection Office investigation, NLB has started to spread rapidly and Hokkaido's NLB rate in 2000 was 15.3%. NLB induced by NLB fungus, has been acknowledged in many countries. The disease shows as spots on the leaves of young corn, which grow as the leaves grow and appear as spindle or lentoid spots when they become bigger (Raymundo & Hooker 1981; Levy 1984; Perkins & Pederson 1987; Lipps et al. 1997; Pataky et al. 1998). It was reported that corn production can be reduced by 30–50% due to NLB (Ullstrup & Miles 1957; Perkins & Pederson 1987; Carson & Van Dyke 1994). Although the reduction in corn production is influenced by the infection rate, it was also reported that there is cellular content loss of infected corn leaves, and acid detergent fiber (ADF) and lignin content increase (Yamakawa & Isawa 1986).
Northern Leaf Blight can be prevented by the use of resistant corn species. Therefore, many studies on field resistance to NLB have been reported (Perkins & Pederson 1987; Lipps et al. 1997; Carson 2006). Except for seriously infected corn, the use of NLB infected corn is equal to use of non-infected corn for ensiling. The effects of NLB on the yield and component of corn have been investigated, but there are very few studies on fermentation quality, nutritive value and feed intake of NLB-infected corn silage. To determine the feed value of NLB-infected corn it is necessary that resistant corn species have not become too widely spread, even though the use is generally expected.
The purpose of this study was to determine the effect of NLB on corn silage fermentation quality, digestibility, nutritive value and feed intake by sheep.
MATERIALS AND METHODS
Plant material
This experiment used whole-crop corn grown at the PIONEER HI-BRED Co., Ltd. farm (Kitami Tanno, Hokkaido, Japan, latitude 43°51′N, longitude 143°56′W). The planting density was 9000/10 are (= 100 sq m), planted on June 1, 1999. NLB fungus was inoculated on July 15, and corn was harvested on October 1. This experimental treatment was performed with a non-inoculated (control) treatment (54 m × 5 m) and an inoculated treatment (54 m × 5 m). NLB fungus was collected from infected leaves, dried and inoculated. For inoculation, crushed leaf samples (3 g dry matter (DM)) were scattered at the grassy crown center in five roots of corn with about 10 leaves. The usefulness of this method in a preliminary experiment was previously confirmed. Resistant species were planted between the two experimental treatments as a barrier (54 m × 5 m) to prevent infection of the control treatment.
Yield of each plant part of corn
On the date of harvest, the catch degree of corn in the two experimental treatments was evaluated by the disease index of NLB (Elliott & Jenkins 1946). Then, to determine the fresh weight of the leaves, stems and grains, these plants were collected from five randomly selected blocks (1 m × 1 m) from each treatment.
Silage preparation
Silage from each of the treatments was ensiled on October 1, 1999. The control and inoculated materials were cut into approximately 1–4 cm lengths before ensiling in 220 L plastic silos. The filling weight was 125–130 kg of fresh matter. The silos were fermented for 120 days. Five replicate silos were prepared for feeding and digestion trials.
Feeding and digestion trial
Four male cross Corriedale sheep (79.3 ± 10.0 kg) were used in the feeding and digestion trials. The trial used total feces collection method by metabolic cage between January 25 and February 27 2000. The experiments used a method with one treatment to four sheep. Feces samples were collected for 5 days after a 10 day adaptation period. Blood was collected from the sheep before feeding early in the morning on day 6. Water and mineral blocks were accessible at all times. The management of animals in this experiment conformed with the Guideline for Animal Experimentation (JALAS 1987). In the digestion trial and on blood collection, experimenters gave consideration to minimize the animals' discomfort and pain, while observing their physical state every day.
Chemical analysis
The chemical composition of the corn, silage and feces were determined using ground samples that had been oven-dried at 60°C for 48 h. The fermentation quality of the silage was determined using water extracts prepared from macerated materials. Fifty grams of silage was macerated with 100 mL of water using a blender. DM content was determined by oven-drying at 135°C for 2 h. Crude protein (CP), ether extracts (EE), and gross energy (GE) contents were determined by AOAC (1990) methods. The ADF and neutral detergent fiber (NDF) contents were analyzed using the procedures described by Van Soest et al. (1991). Water-soluble carbohydrate (WSC) content was determined using the Somogi-Nelson method (McDonald & Henderson 1964). The pH value was determined using a pH meter. Lactic acid content was determined using the colorimetric method of Barker and Summerson (Barker & Summerson 1941). Volatile fatty acids (VFA) were measured using gas chromatography (GC-12A; Shimadzu Co., Ltd. Kyoto, Japan) method (Kageyama et al. 1973). Ammonia nitrogen (NH3-N) was measured using steam distillation (Robinson et al. 1986). V-score was calculated to evaluate the fermentative quality based on the values of organic acids and NH3-N content (JGFFSA 1994).
Using a vacuum tube for blood collection, blood was collected through the jugular vein. Blood samples were analyzed at the East-Hokkaido Clinical Examination Center. The white blood cell (WBC) count, red blood cell (RBC) count, platelet (PLT) count, and hematocrit (Ht) value were measured using the electronic resistance method. The hemoglobin (Hb) level was determined using the sodium dodecyl sulfate method. The levels of alkaline phosphatase (ALP), glutamic oxaloacetic transaminase (AST), and alanine transaminase (ALT) were measured using the modified kind-king method. The urease indophenol, glucose oxidase, and modified Jaffe tests were measured urea nitrogen (UN), glucose (Glu), and creatinine (CRE), respectively.
Statistical analysis
Statistical analysis of differences between mean values of the control and inoculated treatment was performed using Student's t-test.
RESULTS
Disease index of NLB and chemical composition of corn
The disease index of NLB and chemical composition of corn are presented in Table 1. The disease index of NLB was 4.7 and 8.0 for the control and inoculated corn, respectively. The content of WSC was decreased by 18.6 g/kg DM in the inoculated corn compared with the control corn. The contents of ADF and NDF were increased by 40–50 g/kg DM in the inoculated corn compared with control corn.
| Control | Inoculate | |
|---|---|---|
| Growth stage | 1/4 milk line | 1/4 milk line |
| Date of cutting | 1 Oct. 1999 | 1 Oct. 1999 |
| Disease index | 4.7 | 8.0 |
| Chemical composition | ||
| DM (g/kg) | 360.9 | 306.0 |
| OM (g/kg DM) | 940.2 | 931.1 |
| CP (g/kg DM) | 61.7 | 62.6 |
| EE (g/kg DM) | 22.5 | 19.5 |
| WSC (g/kg DM) | 65.3 | 46.7 |
| NFE (g/kg DM) | 646.7 | 603.2 |
| ADF (g/kg DM) | 285.3 | 326.5 |
| NDF (g/kg DM) | 489.3 | 541.9 |
| GE (MJ/kg DM) | 18.5 | 18.2 |
- ADF, acid detergent fiber; CP, crude protein; DM, dry matter; EE, ether extract; GE, gross energy; NDF, neutral detergent fiber; NFE, nitrogen free extract; OM, organic matter; WSC, water soluble carbohydrate.
Yield of each plant part of corn
The yield of each plant part of corn is presented in Table 2. The DM yields of the leaf, stem, grain and gross of corn yield in the inoculated corn were significantly (P < 0.05) decreased by 6.9%, 10.6%, 9.1%, and 9.4% compared with the control corn.
| Control | Inoculate | Significance | |
|---|---|---|---|
| (n = 5) | (n = 5) | ||
| Leaf (kg/10are DM) | 159 ± 10† | 148 ± 11 | P < 0.05 |
| Stem (kg/10are DM) | 471 ± 22 | 421 ± 24 | P < 0.05 |
| Leaf + Stem (kg/10are DM) | 630 ± 39 | 569 ± 28 | P < 0.05 |
| Grain (kg/10are DM) | 961 ± 45 | 874 ± 33 | P < 0.05 |
| Gross (kg/10are DM) | 1591 ± 76 | 1442 ± 74 | P < 0.01 |
- † Mean ± standard error. Comparison of P-values of the control group versus the inoculate group using unpaired t-test. DM, dry matter.
Fermentation quality and chemical composition of silage
The fermentation quality and chemical composition of silage are presented in Table 3. As for fermentation quality of silages, the pH values were 3.98 and 4.01, respectively, in the control and inoculated silages. The contents of butyric acid and the proportion of NH3-N to total N were also low in both the silages. The lactic acid content of the inoculated silage was significantly (P < 0.05) higher than that of the control silage. The V-score in the control and inoculated silage were high, 87.8 and 89.3, respectively, and showed good fermentation quality. With regard to the chemical composition of the silages, the DM, EE, and nitrogen free extract (NFE) content of the inoculated silage was significantly (P < 0.05) lower, and the contents of ADF and NDF were significantly (P < 0.05) higher than that of the control silage.
| Control | Inoculate | Significance | |
|---|---|---|---|
| (n = 5) | (n = 5) | ||
| Fermentation quality | |||
| pH | 3.98 ± 0.03† | 4.01 ± 0.02 | NS |
| Lactic acid (g/kg DM) | 43.3 ± 6.9 | 51.4 ± 5.7 | P < 0.05 |
| Acetic acid (g/kg DM) | 8.5 ± 1.7 | 8.4 ± 2.7 | NS |
| Propionic acid (g/kg DM) | 0.0 ± 0.0 | 0.0 ± 0.0 | NS |
| Butyric acid (g/kg DM) | 0.1 ± 0.0 | 0.1 ± 0.0 | NS |
| Valeric acid (g/kg DM) | 0.0 ± 0.0 | 0.2 ± 0.2 | NS |
| Caproic acid (g/kg DM) | 0.6 ± 1.3 | 0.0 ± 0.0 | NS |
| Total acid (g/kg DM) | 52.5 ± 6.3 | 60.2 ± 5.0 | P < 0.05 |
| Ammonia-N (% TN) | 5.6 ± 1.5 | 4.7 ± 0.9 | NS |
| V-score | 87.8 ± 5.2 | 89.3 ± 1.6 | NS |
| Chemical composition | |||
| DM (g/kg) | 358.2 ± 25.9 | 326.9 ± 21.4 | P < 0.05 |
| OM (g/kg DM) | 952.6 ± 7.4 | 942.7 ± 10.2 | P < 0.05 |
| CP (g/kg DM) | 67.3 ± 9.1 | 71.2 ± 2.7 | NS |
| EE (g/kg DM) | 33.1 ± 2.9 | 27.3 ± 4.7 | P < 0.05 |
| NFE (g/kg DM) | 693.5 ± 28.7 | 647.9 ± 33.8 | P < 0.05 |
| ADF (g/kg DM) | 217.0 ± 30.3 | 263.0 ± 32.5 | P < 0.05 |
| NDF (g/kg DM) | 392.1 ± 32.1 | 449.9 ± 40.1 | P < 0.05 |
| GE (MJ/kg DM) | 18.7 ± 0.2 | 18.6 ± 0.1 | NS |
- †Mean ± standard error. Comparison of P-values of the control group versus the inoculate group using unpaired t-test. ADF, acid detergent fiber; CP, crude protein; DM, dry matter; EE, ether extract; GE, gross energy; NDF, neutral detergent fiber; NFE, nitrogen free extract; NS, not significant; OM, organic matter; TN, total nitrogen.
Digestibility, nutritive value and feed intake of silage
The digestibility, nutritive value and feed intake of silage are presented in Table 4. The digestibility of DM, OM, EE, NFE and GE were significantly (P < 0.05) decreased in the inoculated silage compared with the control silage. The contents of total digestible nutrients (TDN) and digestible energy (DE) were significantly (P < 0.05) decreased by 10.5% and 10.6%, respectively, for the inoculated silage compared with control silage. The intakes of TDN and DE were significantly (P < 0.05, P < 0.01) decreased by 24.5% and 24.2%, respectively, for inoculated compared with control silage. However, the DM intake was not significantly different between the two silage sources.
| Control | Inoculate | Significance | |
|---|---|---|---|
| (n = 4) | (n = 4) | ||
| Digestibility | |||
| DM | 0.725 ± 0.012† | 0.665 ± 0.029 | P < 0.05 |
| OM | 0.747 ± 0.011 | 0.683 ± 0.029 | P < 0.05 |
| CP | 0.450 ± 0.077 | 0.374 ± 0.058 | P < 0.05 |
| EE | 0.788 ± 0.037 | 0.633 ± 0.084 | P < 0.01 |
| NFE | 0.811 ± 0.011 | 0.747 ± 0.034 | P < 0.05 |
| ADF | 0.581 ± 0.028 | 0.600 ± 0.030 | NS |
| NDF | 0.580 ± 0.023 | 0.557 ± 0.022 | NS |
| GE | 0.719 ± 0.012 | 0.648 ± 0.033 | P < 0.05 |
| Nutritive value | |||
| TDN (g/kg DM) | 744.1 ± 11.2 | 665.6 ± 29.2 | P < 0.05 |
| DE (MJ/kg DM) | 13.5 ± 0.2 | 12.1 ± 0.6 | P < 0.01 |
| Feed intake | |||
| DM intake (g/kg BW0.75/day) | 40.9 ± 4.4 | 34.6 ± 4.1 | NS |
| TDN intake (g/kg BW0.75/day) | 30.5 ± 3.6 | 23.0 ± 2.1 | P < 0.05 |
| DE intake (MJ/kg BW0.75/day) | 0.55 ± 0.07 | 0.42 ± 0.04 | P < 0.05 |
- †Mean ± standard error. Comparison of P-values of the control group versus the inoculate group using unpaired t-test. ADF, acid detergent fiber; BW, body weight; CP, crude protein; DE, digestible energy; DM, dry matter; EE, ether extract; GE, gross energy; NDF, neutral detergent fiber; NFE, nitrogen free extract; NS, not significant; OM, organic matter; TDN, total digestible nutrients.
Biochemical composition of blood in sheep
The concentrations of blood biochemical composition are shown in Table 5. In the inoculated treatment, the WBC was significantly higher than in the control treatment (P < 0.05). However, there were no marked differences in the other parameters between the two treatments.
| Control | Inoculate | Significance | Normal ranges | |
|---|---|---|---|---|
| (n = 4) | (n = 4) | |||
| WBC (103/µL) | 5.8 ± 1.8† | 6.6 ± 2.5 | P < 0.05 | 8.6–16.0‡ |
| RBC (106/µL) | 9.8 ± 0.4 | 10.1 ± 0.2 | NS | 5.0–10.0‡ |
| PLT (104/µL) | 32.8 ± 10.0 | 35.7 ± 9.8 | NS | 25–75‡ |
| Hb (g/dL) | 12.4 ± 0.4 | 12.6 ± 0.2 | NS | 9–16‡ |
| Hct (%) | 39.6 ± 1.3 | 40.7 ± 1.8 | NS | 24–50‡ |
| ALP (IU/L) | 161.8 ± 62.3 | 188.3 ± 64.3 | NS | 35–234‡ |
| AST (IU/L) | 79.8 ± 10.3 | 84.5 ± 7.0 | NS | − |
| ALT (IU/L) | 14.8 ± 4.5 | 12.8 ± 3.8 | NS | − |
| UN (mg/dL) | 7.9 ± 2.0 | 10.0 ± 4.1 | NS | 8–20§ |
| CRE (mg/dL) | 1.2 ± 0.3 | 1.1 ± 0.2 | NS | 1.2–1.9§ |
| Glu (mg/dL) | 56.0 ± 9.8 | 59.3 ± 11.8 | NS | 50–80‡ |
- †Mean ± standard error. Normal ranges of the biochemical composition ( ‡ Kaneko 1989; § Nakano 1985). Comparison of P-values of the control group versus the inoculate group using unpaired t-test. ALP, alkaline phosphatase; ALT, alanine transaminase; AST, glutamic oxaloacetic transaminase; CRE, creatine; Glu, glucose; Hb, hemoglobin; Hct, hematocrit; NS, not significant; PLT, blood platelet; RBC, red blood cell; UN, urea nitrogen; WBC, white blood cell.
DISCUSSION
Northern Leaf Blight is generated by representative ascomycetous fungus in cold regions with fleck features. Infections occur before and after the flowering stage with lesions with purplish-brown rims and off-white centers, forming spindly spots of 2–15 cm in length and 1–3 cm in width. Later the spots grow larger, and the leaves turn purple, and then begin to wither (George 1988).
In this experiment, the disease index in the inoculated corn was 8.0. The proportion of infected leaves reached 80% (Elliott & Jenkins 1946). However, this index in the control corn was 4.7; 47% of the leaves were infected. As described above, NLB infection is frequent in Hokkaido. In the experimental year, it was also detected in the control corn. In the present experiment, the WSC, NFE and EE contents of inoculated corn decreased more than in control corn. These decreases may be due to cellular content loses caused by fungus attacking the leaf cells. Therefore, it was inferred that the fiber composition, which is difficult to digest, such as ADF and NDF increase relatively (Yamakawa & Isawa 1986).
It is generally supposed that the influence of NLB on yield is related to the period of attack of the disease, and the damage is more severe if the disease occults earlier (Tosawa 1981). Moreover, the grain yield decreased as the severity of the infection increased (Ullstrup & Miles 1957; Hirose & Toda 1970). Ivanova (1983) reported that the grain yield of infected corn decreased by 11.4% at the 6–8 leaf stage. The present experiment with inoculation at the 10 leaf stage, in which the DM yield of grains decreased by 9.1% and that of leaves and stems decreased as well, implied that the poor growth of the leaves and stems affects the growth of grains.
Generally, because of the high DM content, low buffering capacity, and rich WSC content, corn exhibits good fermentation quality, regardless of its species, date of harvesting, or the use of additives (McAllan & Phipps 1977; McDonald et al. 1991; Aksu et al. 2004; Wang et al. 2009). In this experiment the WSC content of inoculated corn was to 46.7 g/kg DM lower, yet its silage fermentation quality was as good as with control corn, which implied that NLB has no influence on fermentation quality.
Isawa et al. (1974) reported that in crown rust-sickened Italian ryegrass, its cellular content decreased and cellulose levels increased as the disease developed, resulting in reduced digestibility of the fiber component. In the present experiment, NLB-damaged silage showed increased ADF and NDF contents and decreased EE content, and the digestibility of DM, OM, EE and NFE was reduced.
The contents of DM, EE and NFE of corn and silage decreased because of NLB; therefore, it seems that the change in the contents of DM, EE and NFE in corn resulted in changes in these compositions of the silage and influenced the digestibility of the same composition of the silage. Wang et al. (2009) reported that in corn silage experiments contents of CP, EE and NFE decreased, and digestibility of the three components also decreased.
On the other hand, Braman et al. (1973) indicated that the CP content should be established at 11% or more of the dry weight to increase microbial activity in the rumen fermentation of cow. As corn silage alone was given to the sheep in this study, the CP content was approximately 6%. Efficient microbial metabolism and proliferation in the rumen may have been inhibited (Russell & Strobel 1987) decreasing the digestibility of CP and fiber components (Eck et al. 1988; Forbes 1995). However, in this study, there was no difference in the CP content between the two materials; therefore, there may be no influence when comparing the digestibility between the two silages.
There was no discrepancy in the energy content between the inoculated and control silage, and yet the DE content reduced for a decrease of the energy digestibility in inoculated silage. The decrease in the TDN content resulted in the reduction of yield and percentage of grain in whole-crop corn (Woody et al. 1983; Miaki et al. 1989; Inoue & Kasuga 1991). It was thought that the decrease in TDN in NLB-damaged silage was related to the decreasing DM yield of grains on NLB-damaged corn, which equalled 9.1%.
El-shazly et al. (1961) and Forbes (1995) reported that the DM intake decreased with an increase in the fiber component content and decrease in the digestibility of fiber components. In this experiment, an increase in the withering leaf area and a decrease in the yield of grain were observed in the inoculated corn. However, the digestibility of fiber components was not reduced, suggesting that there was no influence on the DM intake of silage. The intakes of TDN and DE were reduced by 24%, and this might be attributed to the decrease in TDN and DE contents.
Hematological examination did not show any marked differences in the levels of components reflecting liver function (ALP, AST, and ALT) between the control and inoculated treatments. On a digestion test, neither diarrhea nor loose stools were observed. Therefore, inoculation did not influence the sheep's health status in this experiment.
This study showed that NLB caused DM harvesting losses and decreased contents of decomposable elements such as NFE and WSC. NLB had no influence on silage fermentation quality but it caused decreased digestibility and contents of TDN and DE. The DM intake from NLB-damaged silage did not decrease significantly, yet TDN and DE intakes reduced sharply according to the decrease in TDN and DE contents.
ACKNOWLEDGMENTS
This study was supported in part by a Grant-in-Aid (No.11660274) from the Ministry of Education, Culture, Sports, Science and Technology of Japan.
