Paper 2.0: Silage fermentation processes and their manipulation - Stefanie J.W.H. Oude Elferink, Frank Driehuis, Jan C. Gottschal, and Sierk F. Spoelstra

INTRODUCTION

Fresh forage crops, such as maize, grasses, legumes, wheat and lucerne, can be preserved by ensiling. In many countries, ensiled forages are highly valued as animal feed. In European countries, such as The Netherlands, Germany and Denmark, more than 90% of the forages locally produced are stored as silage. Even in countries with generally good weather conditions for hay making, such as France and Italy, about half of the forages are ensiled (Wilkinson et al., 1996). It is essential to have a good microbial fermentation process to produce high quality silage. A good fermentation process is not only dependent on the type and quality of the forage crop, but also on the harvesting and ensiling technique. In this paper, our current knowledge on general silage microbiology is reviewed with the aim of assisting in the choice of the best ensiling strategy to produce high quality silage.

THE ENSILING PROCESS

Ensiling is a forage preservation method based on a spontaneous lactic acid fermentation under anaerobic conditions. The epiphytic lactic acid bacteria (LAB) ferment the water-soluble carbohydrates (WSC) in the crop to lactic acid, and to a lesser extent to acetic acid. Due to the production of these acids, the pH of the ensiled material decreases and spoilage micro-organisms are inhibited. Once the fresh material has been stacked and covered to exclude air, the ensiling process can be divided into 4 stages (Weinberg and Muck, 1996; Merry et al., 1997).

Phase 1 - Aerobic phase. In this phase - normally taking only a few hours - the atmospheric oxygen present between the plant particles is reduced, due to the respiration of the plant material and aerobic and facultative aerobic micro-organisms such as yeasts and enterobacteria. Furthermore, plant enzymes such as proteases and carbohydrases are active during this phase, provided the pH is still within the normal range for fresh forage juice (pH 6.5-6.0).

Phase 2 - Fermentation phase. This phase starts when the silage becomes anaerobic, and it continues for between several days and several weeks, depending on the properties of the ensiled forage crop and the ensiling conditions. If the fermentation proceeds successfully, LAB develop and become the predominant population. Due to the production of lactic and other acids, the pH decreases to 3.8-5.0.

Phase 3 - Stable phase. For as long as air is prevented from entering the silo or container, relatively little occurs. Most micro-organisms of phase 2 slowly decrease in numbers. Some acid-tolerant micro-organisms survive this period in an almost inactive state; others, such as clostridia and bacilli, survive as spores. Only some acid-tolerant proteases and carbohydrases and some specialized micro-organisms, such as Lactobacillus buchneri, continue to be active at a low level. The activity of L. buchneri will be discussed in more detail later in this paper.

Phase 4 - Feed-out phase or aerobic spoilage phase. This phase starts as soon as the silage is exposed to air. During feed-out this is unavoidable, but it can start earlier due to damage to the silage covering (e.g. by rodents or birds). The process of spoilage can be divided into two stages. The primary spoilage stage is the onset of deterioration due to the degradation of preserving organic acids by yeasts and, occasionally, by acetic acid bacteria. This will cause a rise in pH, and thus the second spoilage stage is started, which is associated with increasing temperature, and activity of spoilage micro-organisms such as bacilli. The last stage also includes the activity of many other (facultative) aerobic micro-organisms, such as moulds and enterobacteria. Aerobic spoilage occurs in almost all silages that are opened and exposed to air. However, the rate of spoilage is highly dependent on the numbers and activity of the spoilage organisms in the silage. Spoilage losses of 1.5-4.5% DM loss per day can be observed in affected areas. These losses are in the same range as losses that can occur in airtight silos during several months of storage (Honig and Woolford, 1980).

To avoid failures, it is important to control and optimize each phase of the ensiling process. In phase 1, good silo filling techniques will help to minimize the amount of oxygen present between the plant particles in the silo. Good harvesting techniques combined with good silo filling techniques will thus minimize WSC losses through aerobic respiration in the field and in the silo, and in turn will leave more WSC available for lactic acid fermentation in phase 2. During phases 2 and 3, the farmer cannot actively control the ensiling process. Methods to optimize phases 2 and 3 are therefore based on the use of silage additives applied at the time of ensiling, as will be discussed in the section on additives, below. Phase 4 will start as soon as oxygen is available. To minimize spoilage losses during storage, an airtight silo is required, and any damage to the silo covering should be repaired as soon as possible. During feed-out, spoilage by air ingress can be minimized by a sufficiently high feed-out rate. In addition, silage additives capable of decreasing spoilage losses can be applied at the time of ensiling.

THE SILAGE MICROFLORA

The silage microflora plays a key role in the successful outcome of the conservation process. The flora can basically be divided into two groups, namely the desirable and the undesirable micro-organisms. The desirable micro-organisms are LAB. The undesirable ones are the organisms that can cause anaerobic spoilage (e.g. clostridia and enterobacteria) or aerobic spoilage (e.g. yeasts, bacilli, Listeria and moulds). Many of these spoilage organisms not only decrease the feed value of the silage, but also have a detrimental effect on animal health or milk quality, or both (e.g. Listeria, clostridia, moulds and bacilli).

Desirable micro-organisms - Lactic acid bacteria

LAB belong to the epiphytic microflora of plant material. Often the population of LAB increases substantially between harvesting and ensiling. This is probably mainly due to the resuscitation of dormant and non-culturable cells, and not by inoculation by the harvesting machinery or growth of the indigenous population. Crop characteristics, including sugar content, DM content and sugar composition, combined with LAB properties such as acid- and osmo-tolerance, and substrate utilization will decisively influence the competitiveness of the LAB flora during silage fermentation (Woolford, 1984; McDonald et al., 1991).

LAB that are regularly associated with silage are members of the genera Lactobacillus, Pediococcus, Leuconostoc, Enterococcus, Lactococcus and Streptococcus. The majority of the silage LAB are mesophilic, i.e. they can grow at temperatures between 5° and 50°C, with an optimum between 25° and 40°C. They are able to decrease the silage pH to between 4 and 5, depending on the species and the type of forage crop. All LAB are facultative aerobes, but some have a preference for anaerobic conditions (Holzapfel and Schillinger 1992; Hammes et al., 1992; Devriese et al., 1992; Weiss, 1992; Teuber et al., 1992).

Based on their sugar metabolism LAB can be classified as obligate homofermenters, facultative heterofermenters or obligate heterofermenters. Obligate homofermenters produce more than 85% lactic acid from hexoses (C₆ sugars) such as glucose, but cannot degrade pentoses (C₅ sugars) such as xylose. Facultative heterofermenters also produce mainly lactic acid from hexoses, but in addition they also at least degrade some pentoses to lactic acid, and acetic acid and/or ethanol. Obligate heterofermenters degrade both hexoses and pentoses, but unlike homofermenters they degrade hexoses to equimolar mounts of lactic acid, CO₂ and acetic acid and/or ethanol (Hammes et al., 1992; Schleifer and Ludwig 1995). Obligate homofermenters are species such as Pediococcus damnosus and Lactobacillus ruminis. Facultative heterofermenters include Lactobacillus plantarum, L. pentosus, Pediococcus acidilactici, P. pentosaceus and Enterococcus faecium. To the obligate heterofermenters belong members of the genus Leuconostoc, and some Lactobacillus spp., such as Lactobacillus brevis and Lactobacillus buchneri (Devriese et al., 1992; Weiss, 1992; Holzapfel and Schillinger, 1992; Hammes et al., 1992).

Undesirable micro-organisms

Yeasts

Yeasts are eukaryotic, facultative anaerobic, heterotrophic micro-organisms. In silages, anaerobic as well as aerobic yeast activity is considered undesirable. Under anaerobic silage conditions, yeasts ferment sugars to ethanol and CO₂ (Schlegel, 1987; McDonald et al., 1991). This ethanol production in silage not only decreases the amount of sugar available for lactic acid fermentation, but it can also have a negative effect on milk taste (Randby et al., 1999). Under aerobic conditions, many yeast species degrade the lactic acid to CO₂ and H₂O. The degradation of lactic acid causes a rise in silage pH, which in turn triggers the growth of many other spoilage organisms (McDonald et al., 1991).

Yeast populations can reach up to 10⁷ colony forming units per gram during the first weeks of ensiling; prolonged storage will lead to a gradual decrease in yeast numbers (Jonsson and Pahlow, 1984; Middelhoven and van Baalen, 1988; Driehuis and van Wikselaar, 1996). Factors that affect the survival of yeasts during storage are the degree of anaerobiosis and the concentrations of organic acids. The presence of oxygen enhances survival and growth of yeasts during storage (Jonsson and Pahlow, 1984; Donald et al., 1995), whereas high levels of formic or acetic acid reduce survival during storage (Driehuis and van Wikselaar, 1996; Oude Elferink et al., 1999). Initial yeast activity appears to be enhanced in crops with a low initial pH (<5), for example, due to the addition of acid additives, and in crops with a high sugar content, such as potato, orange peel or sugar beet. These crops often result in silages high in ethanol and low in lactic acid (Henderson et al., 1972; Ashbell et al., 1987; Weinberg et al., 1988; Driehuis and van Wikselaar, 1996). Silage additives developed to inhibit yeast activity are described in the section on additives, below.

Enterobacteria.

Enterobacteria are facultatively anaerobic. Most silage enterobacteria are considered to be non-pathogenic. Nevertheless, their growth in silage is undesirable because they compete with the LAB for the available sugars, and in addition they can degrade protein. This protein degradation not only causes a reduction in feeding value, but also leads to the production of toxic compounds such as biogenic amines and branched fatty acids. Biogenic amines are known to have a negative effect on silage palatability (Woolford, 1984; McDonald et al., 1991; van Os and Dulphy, 1996), especially in animals that are not yet accustomed to the taste (van Os et al., 1997). Moreover, the ammonia formed through proteolysis increases the buffer capacity of the ensiled crop, thus counteracting any rapid decrease in silage pH. A special characteristic of enterobacteria is their capability to reduce nitrate (NO₃) to nitrite (NO₂) under silage conditions. In silage, nitrite can be degraded by enterobacteria to ammonia and nitrous oxide (N₂O), but it can also be chemically degraded to NO and nitrate (Spoelstra, 1985, 1987). With air, NO is oxidized into a mixture of gaseous, yellow-brown nitrogen oxides (NO₂, N₂O₃, N₂O₄). Gaseous NO and NO₂ have a damaging effect on lung tissue and can cause a disease with pneumonia-like symptoms known as “silo filler’s disease” (Woolford, 1984). To prevent animals from being in contact with gaseous nitrogen oxides, they should not be housed in buildings adjoining silos during silo filling or the first week of silage storage (O'Kiely et al., 1999). Despite the above-mentioned problems, a little nitrite reduction is considered positive for silage quality, because the nitrite and NO formed are very effective inhibitors of clostridia (Woods et al., 1981; Spoelstra, 1985).

Enterobacteria will not proliferate at low pH. Ensiling methods that induce a rapid and sufficient drop in silage pH will therefore help to decrease enterobacterial growth (McDonald et al., 1991).

Clostridia

Clostridia are endospore-forming anaerobic bacteria. Many clostridia ferment carbohydrates as well as proteins, thus causing problems such as reduction in feeding value and the production of biogenic amines, similarly to that described for enterobacteria. Furthermore, clostridia in silage impair milk quality. This is due to the fact that clostridial spores can survive the passage through the alimentary tract of a dairy cow. Clostridial spores present in silage are transferred to milk via faeces and faecal contamination of the udder. The acid-tolerant Clostridium tyrobutyricum is the most relevant species for the dairy industry. In addition to carbohydrate fermentation, C. tyrobutyricum can degrade lactic acid to butyric acid, H₂ and CO₂ according to the following overall reaction:

2 lactic acid Þ 1 butyric acid + 2 H₂ + 2 CO₂

This butyric acid fermentation not only counteracts lactic acid fermentation in silage and cheeses, but it is also responsible for significant gas production, causing a cheese defect called “late blowing” in hard and semi-hard cheeses such as Emmental, Grana, Gouda and Parmesan (Gibson, 1965; Goudkov and Sharpe, 1965; Klijn et al., 1995).

Some clostridia can cause serious health problems. One extremely toxic species is Clostridium botulinum. This organism can cause botulism, which can be deadly for cattle. Fortunately, C. botulinum has a limited acid tolerance, and does not grow in well-fermented silage. Incidences of animal botulism caused by silage contaminated with C. botulinum could nearly always be attributed to the presence of a cadaver (e.g. mouse, bird, etc.) in the silage (Kehler and Scholz, 1996).

A typical “clostridial silage” is characterized by a high butyric acid content of more than 5 g/kg DM, a high pH (over pH 5 in low DM silages), and a high ammonia and amine content (Voss, 1966; McPherson and Violante, 1966). Ensiling methods that cause a rapid and sufficient drop in silage pH will help to prevent the development of such clostridial silage, because, as for enterobacteria, clostridia are inhibited at low pH. Furthermore, clostridia are more susceptible to a low availability of water (i.e. a low water activity (a_w)) than LAB (Kleter et al., 1982, 1984; Huchet et al., 1995). For this reason, decreasing the aw-value of a crop, such as by wilting to a higher DM content, can be a way of selectively inhibiting clostridia (Wieringa, 1958). Finally, clostridia will also be inhibited by nitrite and NO, or compounds that are degraded in silage to nitrite and NO (Spoelstra, 1983, 1985).

Acetic acid bacteria

Acetic acid bacteria are obligate aerobic, acid-tolerant bacteria. Thus far, all acetic acid bacteria that have been isolated from silage belong to the genus Acetobacter (Spoelstra et al., 1988). The activity of Acetobacter spp. in silage is undesirable because they can initiate aerobic deterioration, as they are able to oxidize lactate and acetate to CO₂ and water. Generally, yeasts are the main initiators of aerobic spoilage, and acetic acid bacteria are absent, or play only a minor role. However, for whole-crop corn silages there is evidence that acetic acid bacteria alone can initiate aerobic deterioration (Spoelstra et al., 1988). Furthermore, selective inhibition of yeast can also increase proliferation of acetic acid bacteria in silage (Driehuis and van Wikselaar, 1996).

Bacilli

Bacilli are like clostridia: endospore-forming, rod-shaped bacteria. Nevertheless, they can easily be distinguished from clostridia as they are (facultative) aerobes, whereas all clostridia are obligate anaerobes (Claus and Berkeley, 1986; Cato et al., 1986). Facultative aerobic bacilli ferment a wide range of carbohydrates to compounds such as organic acids (e.g. acetate, lactate and butyrate) or ethanol, 2,3-butanediol and glycerol (Claus and Berkely, 1986). Some specific Bacillus spp. are able to produce antifungal substances, and have been used to inhibit aerobic spoilage of silage (Phillip and Fellner, 1992; Moran et al., 1993). Except for these specific strains, the proliferation of bacilli in silage is generally considered undesirable. Not only are bacilli less efficient lactic and acetic acid producers than LAB (McDonald et al., 1991), they can also enhance (later stages of) aerobic deterioration (Lindgren et al. 1985; Vreman et al., in press). Furthermore, high numbers of bacillus spores in raw milk have been associated with high spore numbers in fresh cow faeces (Waes, 1987; te Giffel et al., 1995). It seems very plausible that bacillus spores are transferred from silage to milk via faeces, as occurs with clostridial spores (Vreman et al., in press). Psychrotrophic Bacillus cereus spores are considered to be the most important spoilage organism of pasteurized milk (te Giffel, 1997). High numbers of these (psychrotrophic) B. cereus spores have been found in silages (Labots et al., 1965; te Giffel et al., 1995).

To decrease bacillus growth in silage, storage temperatures should not be too high (Gibson et al. 1958) and air ingress should be minimized (Vreman et al., in press). In addition, initial contamination of fresh plant material with soil or manure should be prevented (McDonald et al., 1991; Rammer et al. 1994).

Moulds

Moulds are eukaryotic micro-organisms. A mould-infested silage is usually easily identified by the large filamentous structures and coloured spores that many species produce. Moulds develop in parts of the silage where (a trace of) oxygen is present. During storage, this is usually only in the surface layers of the silage, but during aerobic spoilage (phase 4) the whole silage can become mouldy. Mould species that regularly have been isolated from silage belong to the genera Penicillium, Fusarium, Aspergillus, Mucor, Byssochlamys, Absidia, Arthrinium, Geotrichum, Monascus, Scopulariopsis and Trichoderma (Pelhate, 1977; Woolford, 1984; Frevel et al., 1985; Jonsson et al., 1990; Nout et al., 1993). Moulds not only cause a reduction in feed value and palatability of the silage, but can also have a negative effect on human and animal health. Mould spores are associated with lung damage and allergenic reactions (May, 1993). Other health problems are associated with mycotoxins that can be produced by moulds (Oldenburg, 1991; Auerbach, 1996). Depending on the type and amounts of toxin present in the silage, health problems can range from minor digestive upsets, small fertility problems and reduced immune function, to serious liver or kidney damage, and abortions (Scudamore and Livesey, 1998). Some important mycotoxin-producing mould species are Aspergillus fumigatus, Penicillium roqueforti, and Byssochlamys nivea. P. roqueforti, a species which is acid tolerant and can grow at low levels of oxygen and high levels of CO₂, has been especially detected as the predominant species in different types of silages (Lacey, 1989; Nout et al., 1993; Auerbach et al., 1998; Auerbach, 1996). There is still uncertainty concerning the conditions under which mycotoxins are formed in silage. A heavily infested silage does not necessarily contain high levels of mycotoxins, and not all types of mycotoxins that a mould species can produce are necessarily present in one silage lot (Nout et al., 1993; Auerbach, 1996). For Aflatoxin B1, a mycotoxin of Aspergillus flavus, it is known that it can be transferred from animal feed to milk. However, so far it is unknown if a similar transfer can occur with mycotoxins from P. roqueforti or A. fumigatus (Scudamore and Livesey, 1998).

Ensiling methods that minimize air ingress (e.g. good compaction and covering of the silo), and additives that prevent initiation of aerobic spoilage, will help to prevent or limit mould growth.

Listeria

Members of the genus Listeria are aerobes or facultative anaerobes. Regarding silage quality, the most important species is the facultative anaerobe L. monocytogenes, because this species is pathogenic to various animals and man. Animals with a suppressed immune system (e.g. pregnant females and neonates) are especially susceptible to L. monocytogenes infections (Jones and Seeliger, 1992). Silage contaminated with L. monocytogenes has been associated with fatal cases of listeriosis in sheep and goats (Vazquez-Boland et al., 1992; Wiedmann et al., 1994). In addition, Sanaa et al. (1993) have identified poor quality silage as one of the main sources of contamination of raw milk by L. monocytogenes. Growth and survival of Listeria spp. in silage are determined by the degree of anaerobiosis, and the silage pH. L. monocytogenes can tolerate a low pH of 3.8-4.2 for long periods if oxygen is present, even if only at low levels. Under strictly anaerobic conditions, it is rapidly killed at low pH (Donald et al., 1995). Silages that have a higher chance of aerobic surface spoilage, such as big bale silages, seem to be particular liable to Listeria contamination (Fenlon et al., 1989). L. monocytogenes generally does not develop in well-fermented silages with a low pH. So far, the most effective method to prevent growth of L. monocytogenes is to keep the silage anaerobic (McDonald et al., 1991).

SILAGE ADDITIVES

In the 1990s, it became increasingly common to use silage additives to improve the ensiling process. The choice of additives appears to be almost limitless if one looks at the large number of chemical and biological silage additives that are commercially available. The UKASTA Forage Approval Scheme of the UK, for example, listed more than 80 products (Rider, 1997). Fortunately, the choice of a suitable additive is less complicated than it seems, because the modes of action of most additives fall into a few categories (Table 1).

Between products of a particular category, differences exist in product properties, such as general effectiveness, suitability for certain crop type, and ease of handling and application. These factors, together with price and availability, will determine what product will be the most appropriate for a specific silage. A drawback of some of the chemical additives is that they can be corrosive to the equipment used, and can be dangerous to handle. The biological additives are non-corrosive and safe to handle, but they can be costly. Furthermore, their effectiveness can be less reliable, since it is based on the activity of living organisms. Proper storage of these biological additives by the manufacturer, retailer and farmer is of vital importance. Despite these disadvantages, in Europe and the USA bacterial inoculants have nowadays become the most commonly used additives for maize, and grasses and legumes that can be wilted to above 300 g DM/kg (Bolsen and Heidker, 1985; Pahlow and Honig, 1986; Bolsen et al., 1995; Kung, 1996; Weinberg and Muck, 1996). In the Netherlands, the absolute as well as the relative amount of silages treated with bacterial inoculants has increased between 1995 and 1998, and in 1998, 13.7% of all grass silage in the Netherlands was ensiled with an additive, of which 31% was treated with an inoculant, 37% with molasses and 29% with fermentation inhibitors (Hogenkamp, 1999).

Table 1. Categories of silage additives (adapted from McDonald et al., 1991)

Notes: *or corresponding salt

Additives improving silage fermentation

Assuming good harvesting and ensiling techniques, initial silage fermentation (phase 2) can still be sub-optimal. This can be due to a lack of sufficient numbers of suitable LAB or a lack of sufficient amounts of suitable WSC, or both.

The amount of WSC necessary to obtain sufficient fermentation depends on the DM content and the buffer capacity of the crop. Weissbach and Honig (1996) characterized the relation between these factors as follows,

FC = DM (%) + 8 WSC/BC

Where:

FC = fermentation coefficient
DM = dry matter content
WSC = water-soluble carbohydrates
BC = buffer capacity.

Forages with insufficient fermentable substrate or too low a DM content have an FC <35. In these forages, sufficient fermentation can only be achieved if the sugar content of the material is increased, either by adding sugars directly (e.g. molasses) or by adding enzymes that release extra sugars from the crop. In forages with an FC of 35 or more, sufficient fermentable substrate is available. Also, adding suitable LAB can accelerate and improve the ensiling process. In high DM silages with reduced water availability, the presence of suitable, osmo-tolerant LAB could become a limiting factor in the ensiling process. It has been shown that these bacteria represent only a small percentage of the indigenous microflora on forage crops (Pahlow and Weissbach, 1996). Forages with a DM content above 50% are considered difficult to ensile (Staudacher et al., 1999).

The formula of Weissbach and Honig (1996) does not apply for crops with a low nitrate content, such as extensively managed grasses and immature whole-crop cereals, because these crops are more liable to clostridial fermentations than crops with a moderate nitrate content (Spoelstra, 1983, 1985). Inoculants that increase lactic acid fermentation might be useful to inhibit clostridial activity. The minimum number of LAB required to inhibit clostridial activity was found to be at least 100 000 colony-forming units per gram of fresh crop (Weissbach and Honig, 1996; Kaiser and Weiss, 1997).

Additives inhibiting silage fermentation

Fermentation inhibitors could in theory be used for all types of forages. However, in practice they are generally only used in wet crops with a low WSC content and/or high buffer capacity (McDonald et al., 1991). In the Netherlands, salts of acids have become the most popular fermentation inhibitors (Hogenkamp, 1999). An advantage of these salts is that they are easier and safer to handle than the corresponding acids.

Silage additives inhibiting silage fermentation can reduce clostridial spore counts. In wilted grass silages, a decrease in spore counts by a factor 5 to 20 has been observed. A similar decrease in spore counts could be obtained by adding molasses, a fermentation stimulant. To inhibit clostridial growth, the most effective fermentation inhibitors appear to be additives based on formic acid, hexamethylene and nitrite (Hengeveld, 1983; Corporaal et al., 1989; van Schooten et al., 1989; Jonsson et al., 1990; Lattemae and Lingvall, 1996).

Additives inhibiting aerobic spoilage

It is clear that to inhibit aerobic spoilage, spoilage organisms, in particularly the ones causing the onset of deterioration (i.e. yeasts and acetic acid bacteria) have to be inhibited in their activity and growth. Some additives that have proven to be effective in this respect include chemical additives based on volatile fatty acids such as propionic and acetic acid, and biological additives based on bacteriocin-producing micro-organisms such as lactobacilli and bacilli (Woolford, 1975a; McDonald et al., 1991; Phillip and Fellner, 1992; Moran et al., 1993; Weinberg and Muck, 1996).

Furthermore, it is known that sorbic acid and benzoic acid have a strong antimycotic activity (Woolford, 1975b; McDonald et al., 1991). Recently, it was discovered that Lactobacillus buchneri is a very effective inhibitor of aerobic spoilage. The inhibition of spoilage appears mainly due to the capability of L. buchneri to anaerobically degrade lactic acid to acetic acid and 1,2-propanediol, which in turn causes a significant reduction in yeast numbers (Driehuis et al., 1997, 1999; Oude Elferink et al., 1999). This reduction in yeast numbers is in agreement with the finding that volatile fatty acids such as propionic acid and acetic acid are much better inhibitors of yeasts than is lactic acid, and that mixtures of lactic acid and propionic or acetic acid have a synergistic inhibitory effect (Moon, 1983). The results of Moon (1983) also explain why, in most cases, biological inoculants that promote homofermentative lactic acid fermentation do not improve, and may even decrease, aerobic stability (Weinberg and Muck, 1996; Oude Elferink et al., 1997).

Biological additives based on the propionate-producing propionibacteria appear to be less suitable for the improvement of silage aerobic stability, due to the fact that these bacteria are only able to proliferate and produce propionate if the silage pH remains relatively high (Weinberg and Muck, 1996).

Additives used as nutrients or absorbents

Certain crops are deficient in dietary components essential for ruminants. The nutritional quality of these crops can be improved by supplementation with specific additives at the time of ensiling. Additives that have been used in this respect are ammonia and urea to increase the crude and true protein content of the silage, and limestone and MgSO₄ to increase the calcium and magnesium contents. The above mentioned additives generally have no beneficial effect on silage fermentation, but urea and ammonia can improve the aerobic stability of silage (Glewen and Young, 1982; McDonald et al., 1991).

Absorbents are used in crops with a low DM content to prevent excessive effluent losses. Good results have been obtained with dried pulps such as sugar beet pulp and citrus pulp. Straw can also be utilized, but has a negative effect on the nutritive value of the silage (McDonald et al., 1991).

Combined additives

Most commercial additives contain more than one active ingredient in order to enhance efficacy and have a broad range of applicability. Very popular are, for example, combinations of inoculants stimulating homofermentative lactic acid fermentation together with sugar releasing enzymes, or combinations of fermentation and aerobic deterioration inhibiting chemicals such as formic acid, sulphite salts and propionic acid (Rider, 1997; Anon., 1999). New additives are currently being developed that decrease the negative effect of homofermentative lactic acid fermentation on aerobic stability. Promising results have been obtained by combining homofermentative or facultative heterofermentative LAB with chemicals such as ammonium formate and sodium benzoate (Kalzendorf, 1992; Bader, 1997), or by combining facultative heterofermentative LAB with the obligate heterofermentative L. buchneri.

SILAGE FERMENTATION IN TROPICAL SILAGES

Ensiling of forage crops or industry by-products could make an important contribution to the optimization of tropical and sub-tropical animal production systems, but thus far it has not yet been widely applied (Wilkins et al., 1999). This is due not only to the low prices for animal products, the low levels of mechanization and the high costs of silo sealing materials, but also to a lack of ensiling experience. More research is needed to address the specific problems associated with tropical silages. Tropical grasses and legumes have, for example, a relatively high concentration of cell wall components and a low level of fermentable carbohydrates compared to temperate forage crops (Catchpoole and Henzell, 1971; Jarrige et al., 1982). Furthermore, on average, storage temperatures in tropical climates are higher than in temperate climates, which might give bacilli a competitive advantage over LAB (Gibson et al., 1958). In addition, it has to be taken into account that some silo sealing materials cannot withstand intense sunlight, and this might impair the aerobic stability of the silage. Nevertheless, it seems likely that ensiling technologies from temperate climates can be modified for tropical conditions.

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