Pressed sugar beet pulp (PBP) silage represents a highly nutritive component for dairy cow rations in many European countries, but the hygienic quality often renders the silages produced from it unfit for feeding due to fungal spoilage. For this reason, farmers are hesitant to use this valuable dietary component in summer periods when the susceptibility to aerobic deterioration is most obvious. Verhülsdonk et al. (2008) monitored fungal counts in farm silos which had been filled with PBP from the same sugar factory in 2006. Sampling in March/April of the following year revealed lower yeast counts than were detected in the same silos in May/June. In both testing periods the level of yeast infestation was lower in PBP samples taken from 2 m behind the cutting face than found in those from the cutting face. In a farm survey in Northwest Germany, Kalzendorf (2007) reported that 75% of all tested PBP silages contained yeasts in excess of 106 cfu/g. In addition, 75% of these silages were also infested with lactate-assimilating yeasts exceeding the proposed threshold value of 105 cfu/g (Jonsson and Pahlow, 1984), above which there is a high risk for aerobic instability (heating) of silages. It was found that many silos were improperly sealed or were opened too early after less than six weeks of fermentation. This was confirmed by Potthast et al. (2014a) who monitored PBP silage use on 31 farms in the German States of Hesse and Rhineland-Palatinate. Visibly moulded feed was detected on nine farms (29%), and in 35% heating was observed. The weekly feed-out rate ranged between 0.7 and 3.6 m and was frequently found to be below the recommended values of 1.4 m in winter and 2.8 m in summer.
The level of fungal contamination of fresh PBP tends to increase with progressing length of the sugar beet harvesting and processing season. According to Weber et al. (2006a) who evaluated PBP from three different sugar plants located in the German States of Saxony-Anhalt and Saxony, yeasts and mould counts were higher at the end of the campaign in December when compared to the start in October. Concurrently, aerobic stability (ASTA) of PBP silages stored in plastic bags decreased from 6.4 days to 4.3 days.
The bagging technology has attracted significant attention and is now widely used for the production of PBP silage because it enables the adaptation of the feed-out rate based to daily demand by using bags of different diameters. However, ASTA is often a matter of concern, especially when storage length is short (Weber et al., 2006b) and ambient temperatures are high. In order to enhance ASTA, different silages additives have been tested but the number of studies is still very limited, especially for storage of PBP in plastic bags. Weber et al. (2006b) evaluated biological and chemical silage additives at laboratory scale. Air ingress was allowed for 24 hours on day 28 and 49 of fermentation by removing rubber stoppers, which closed a hole of 6 mm diameter in the lid and in the glass jar. The applied homo- and heterofermentative lactic acid bacteria (LAB) products were outperformed by all tested chemical additives containing antimycotic ingredients, which is in agreement with data by Wyss and Fivian (1999). This can be explained by the inability of LAB to survive and grow at high temperatures (around 60°C) at which PBP leave the sugar factory (Verhülsdonk et al., 2008). Weber et al. (2006b) also ensiled PBP in plastic bags after treatment with graded doses (2.5 and 5.0 L/tonne) of a chemical additive (mixture of sodium benzoate and sodium propionate) and showed the effect of storage length on the required additive dosage. The application of the low rate (2.5 L/tonne) was sufficient to markedly improve ASTA in silages which were stored for 183 days, whereas the high dosage (5.0 L/tonne) was efficient even after a very short storage period of 14 days. The beneficial effects of chemical additives in reducing fungal contamination and improving ASTA of PBP stored in plastic bags were substantiated by Potthast et al. (2014b), who tested three different chemical additives of different composition.
As there is still little information available on the effects of chemical additives on fungal counts and ASTA, this study was designed to gain more knowledge on the behavior of PBP silages stored in plastic bags and exposed to air under challenging testing conditions of high temperature in summer and long exposure time of 7 days.
2. MATERIALS AND METHODS
The study was performed at the State Institute for Agriculture, Forestry and Horticulture Saxony-Anhalt, Iden, Germany, on December 12, 2012. Fresh sugar beet pulp was obtained from the sugar factory of the company Pfeifer&Langen in Könnern, Saxony-Anhalt, Germany and transported for about 4 hours by truck to the research facility. Each bag (diameter: 3 m, length 6-8 m) of the four treatments – control and three different chemical additives – was filled with about 50 tonnes of fresh PBP by using a M7000 bagging machine (Ag-Bag, Kleinbautzen, Germany). The following additives were used: KOFASIL STABIL (KS, ADDCON EUROPE GmbH, Bitterfeld-Wolfen, Germany), containing sodium benzoate and potassium sorbate: 1.5 L/tonne; SILOSTAR LIQUID HD (SS, Schaumann GmbH, Pinneberg, Germany), composed of sodium benzoate, potassium sorbate and sodium acetate: 2.0 L/tonne and NOVIBAC (NB, INNOV AD, Essen, Belgium), containing formic acid, propionic acid, acetic acid, salts of fatty acids and essential oils: 2.7 L/tonne. The additives were applied by an applicator mounted on the bagging machine above the rotor which feeds the bag. All bags were placed outdoors in west-east direction (Fig. 1) and opened at the east end on June 25, 2013, after 195 days of storage.
Figure 1: Position of the silage bags in the trial
Fresh PBP was sampled manually from the conveyer belt during filling of the bags. On the day of bag opening, three replicate samples were taken by using a hollow drill (diameter: 13 cm; length: 33 cm) from each bag from pre-assigned locations in the core (points 1-3) and the upper layer (points 4-6) (Fig. 2).
Figure 2. Sampling points in the bags
The surfaces of the opened bags were covered with a bird protection net and exposed to air for 7 days. The same surface was sampled again (“old”), followed by removal of 1 m of PBP silage and repeated sampling from the new surface (“fresh”). This procedure was repeated twice at weekly intervals. The trial was terminated 4 weeks after bag opening when only old surfaces were left for sampling.
Fresh PBP was analyzed for dry matter (DM) and nutrient fractions (VDLUFA, 2011). Energy content was calculated according to the German Feed Evaluation System (GfE, 1995). The DM of the silages was corrected for the loss of volatiles during drying (Weissbach and Strubelt, 2008). Fermentation pattern was determined in frozen samples by gas-chromatography (propionic, butyric and valeric acids, methanol) and high-pressure liquid chromatography (lactic and acetic acids, ethanol), and pH measured by using an electrode. Samples for the determination of yeasts and moulds according to ISO 7954:1987 were kept cool at 4°C until analysis within 4 hours after sampling. Aerobic stability of the silages was assessed by employing the temperature method as described by Honig (1990) using thermologgers. Silages were considered unstable once the temperature of the silage had reached 3-°C above ambient (20-22°C). Statistical analysis was performed by using the ANOVA procedure of the computer program SPSS (IBM Statistics 19.0). Fungal counts were transformed into log units prior to statistical evaluation. Values below the limit of detection were set at log 1.7 cfu/g, which is half the detection limit of 102 cfu/g. A significance level of P<0.05 was employed and trends declared at P<0.10. Differences among means were separated by Tukey´s test. The correlations between fungal populations and aerobic stability were characterized by means of Spearman and Pearson correlation coefficients (SPSS, IBM Statististics 19.0), and PROC NLIN (SAS, 9.4) was used to describe the relationship between total fungal count and aerobic stability.
3. RESULTS AND DISCUSSION
3.1 Chemical and microbiological characterization of fresh pressed sugar beet pulp prior to bagging
The results in Table 1 on nutrient and energy concentration of fresh PBP prior to ensiling support previous findings by Weber et al. (2006a). However, fungal counts were lower than those reported by these authors for PBP ensiled in the late campaign (December) reflecting the high, and unavoidable, variation between production batches. The low pH of 4.4 indicated that a fermentation process had already started during intermediate storage in the sugar plant and transport as pH values of around 6 are normally measured in unfermented PBP at factory level (Weber et al., 2006a).
Table 1: Mean energy and nutrient concentrations, fungal counts and pH of fresh pressed sugar beet pulp (n=12, values in brackets represent standard deviation)
3.2 Fermentation characteristics of pressed sugar beet pulp silage on the day of bag opening
Generally, PBP silages were well fermented. With the exception of lactic acid, sampling site did not affect fermentation characteristics (data not given). Silages from the upper layer contained less lactic acid than those from the core of the bags (57.0 g/kg DM vs. 65.6 g/kg DM, P<0.05). Weber et al. (2006a) observed no differences between sampling sites with regard to lactic acid but found lower acetic acid contents in upper layer samples. Ethanol was also unaffected by sampling site, supporting the results of this study.
Regardless of the sampling site, the concentrations of lactic, acetic and butyric acids and pH were not influenced by the used chemical additives (Tables 2 and 3). Propionic acid content was highest in treatment NB, which is attributable to the composition of the product by which propionic acid was added to fresh PBP. The most prominent effect of additive treatment was observed for the parameter ethanol. All additives decreased the level of this end-product of yeast metabolism in silages from the upper layer of the bags, whereas in core samples only product SS reduced it. Methanol, which is formed by acidic degradation of pectins, remained unaffected by additive use in core samples, whereas a decrease was found by treatment SS in silages from the upper layer. The reason for this finding remains to be elucidated as product SS does not contain acids which may have increased the extent of pectin degradation.
Table 2. Effects of additives on fermentation characteristics in pressed sugar beet silages taken from the core of the bags (n=12, data given in g/kg DM unless otherwise stated)
Table 3. Effects of additives on fermentation characteristics in pressed sugar beet silages taken from the upper layer of the bags (n=12, data given in g/kg DM unless otherwise stated)
3.3 Fungal counts and aerobic stability
3.3.1 Effect of sampling site on fungal counts and aerobic stability in pressed sugar beet pulp silages
The effects of the sampling site on fungal count and ASTA are summarized in Tables 4 and 5. There were observed trends for lower yeast count in core samples taken from fresh surfaces and for moulds in old surfaces, which had been exposed to air for 7 days. However, if only untreated silages were evaluated (9 observations per sampling site), core samples contained lower mould (lg 2.2 vs lg 4.4, P<0.05) and yeast (lg 2.2 vs lg 3.3, P=0.06) counts than determined in the upper layer of the bags. Concurrently, ASTA was enhanced (103 hours vs 59 hours, P<0.05). These results substantiate data by Weber et al. (2006a) who found numerically higher yeast numbers and lower ASTA in samples taken from the mid upper layer of bags than were detected in the core. This can be explained by decreased density in the upper layers, enabling larger air volume to penetrate deeper into the bag behind the cutting face. Weber et al. (2006) showed density to be increased by 15-20% in the core of the bags.
Table 4: Effect of sampling site in bags on fungal count and aerobic stability of pressed sugar beet silages taken from the fresh surface
Table 5: Effect of sampling site in bags on fungal count and aerobic stability of pressed sugar beet silages taken from surfaces exposed to air for one week
3.4 Effects of additives on fungal counts and aerobic stability of pressed sugar beet pulp silages
Across treatments, additives increased ASTA of PBP silages taken from the fresh surface from 81 hours in the control to 248 hours (P<0.001) with no differences between chemical additives. This can be explained by the reduction in yeast (lg 2.8 vs log 1.8, P<0.001) and mould (lg 3.3 vs lg 2.1, P<0.001) counts (Table 6). The magnitude of the effect of the additives was higher than the previously reported improvement in ASTA by approximately 2 days in PBP silages stored in bags (Weber et al., 2006b). Also Wyss and Fivian (1999) observed enhanced ASTA by the use of chemical additives when tested at laboratory scale in 1.5 L glass jars, and no effect of biological additives was observed.
Table 6: Effects of additives on fungal counts and aerobic stability in samples taken from the fresh surface of pressed sugar beet pulp silages stored in bags
Exposure to air for one week resulted in reduced aerobic stability (Table 7) when compared to samples taken from fresh surfaces. Untreated PBP silage deteriorated already after 13 hours. Regardless of additive type, there was an increase in ASTA by 100 hours (P<0.001). This can be attributed to lower fungal counts in treated PBP silages when compared with controls (P<0.001). These findings are in line with those by Potthast et al. (2014b) using a similar experimental design of removing 2 m of PBP silage per week and subsequently exposing the surface of the bagged PBP silages to air for one week. The tested combination of sodium benzoate and sodium propionate showed the lowest fungal counts, and in turn the highest ASTA during the entire experimental period.
Table 7: Effects of additives on fungal counts and aerobic stability in samples taken from the surface of pressed sugar beet pulp silages stored in bags after one week of exposure to air
3.5 Relationship between fungal counts and aerobic stability
Fungal counts and ASTA were negatively correlated whereas a positive relationship was detected between yeast and mould numbers (Table 8). The relationship between total fungal count and ASTA was best fitted by an exponential curve (Fig. 3).
Table 8: Correlation coefficients between fungal counts and ASTA
Figure 3: Relationship between total fungal count and aerobic stability in PBP silages (n=72)
Strong negative relationships between yeast counts and ASTA have been described earlier by Wyss (2002) for PBP silages (logarithmic, r2=0.78), but also for other silage types, e.g. green rye (Auerbach et al., 2014; linear, r2=0.85; P<0.0001) and corn (Auerbach and Nadeau, 2013, linear, r2=0.87; P<0.001). This clearly indicates that yeasts are the most likely causal microorganisms for aerobic deterioration of silages.
It is concluded that PBP silages stored in plastic bags are prone to aerobic deterioration caused by yeasts and moulds and, if untreated, samples from upper layers show lower aerobic stability than those from the core. Chemical silage additives reduce the level of fungal infestation thereby effectively enhancing aerobic stability. Therefore, their use in PBP silage is strongly advised especially when PBP silage is to be fed during the summer period in order to maintain good hygienic quality of this valuable feed component.
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This paper was presented at the 1st International Conference of Grain Storage in Silo Bag, Argentina, 2014.