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Small-scale laboratory silos are accepted as a suitable model for farm-scale ensiling. They are used for rapid, cost-effective evaluation of the effects of numerous experimental treatments on silage fermentations. The ideal ensiling conditions that lab silos provide are difficult to achieve in practice, however these conditions allow controlled repeatable comparisons between silage fermentations to be made.

A range of laboratory and pilot scale silo types have been used, with capacities ranging from 50g to several tonnes (McDonald et al., 1991). These include fixed volume vessels such as boiling tubes, glass jars, and plastic pipes, all fitted with sealing devices. Historically, the glass tube has been most widely used laboratory silo (McDonald et al., 1991), however vacuum packing in plastic bags has become a popular method in recent times (Johnson et al., 2005). The basic criteria for any laboratory silo is the exclusion of air, and thus maintenance of anaerobic conditions. Other desirable features of a laboratory silo that mimic the conditions of a farm silo may include; high herbage density, light exclusion, and a means of escape for effluent or gasses produced in the silo. There are inherent problems with any type of laboratory silo, and caution is advised in extrapolating data from the lab scale to farm scale silages (Cherney et al., 2004). Only a few studies have compared fermentations in different types of laboratory silo, or compared laboratory silo fermentations with pilot scale or farm scale silo fermentations. Wilson and Wilkins (1972), compared 100g test tube silages with 6 kg and 1000 kg plastic bag silages of 18 grass and 8 legume species. They found good replication between the 100g test tubes, and close correlation in fermentation parameters between the test tube and plastic bag silages. O'Kiely and Wilson (1991), observed that ryegrass plastic pipe silos fitted with effluent and gas release devices were a better model than test tube silos for farm-scale clamp silos. The general process for ensiling was the same between silo types, however they discovered the potential for significant fermentation x silo type interactions when comparing treatments. Such interactions were observed recently in a comparison of vacuum packed and glass jar silages (Hoedtke & Zeyner, 2011).

2.7.1 Vacuum packing versus fixed volume vessel silages

There are several drawbacks to using fixed volume vessels as small model silages. Packing and unpacking vessels is labour and time consuming. Depending on the operator, variability in packing densities can arise within a vessel and between replicates, which affects the speed of fermentation and pH decline. In a

laboratory setting, vessel packing aims to achieve densities within the range of those found in a farm silo stack (0.32-0.64g cm−3_{) (Johnson et al., 2005). Even packing of samples becomes particularly difficult with} heterogeneous plant material (e.g. containing stems or large structures). Chopping can help in this regard, however fine chopping may not reflect typical on-farm practices.

Two recent studies have explored the use of vacuum bags as model silo vessels and have confirmed that vacuum packing herbage in plastic bags is a convenient ensiling method that avoids the shortcomings of fixed volume vessels. Johnson et al. (2005), ensiled chopped, wilted PRG and red clover (Trifolium pratense), with a DM of 28.0% and 25.0%, respectively. They applied commercial inoculant to half of the plant material before ensiling 100g replicates (wilted material) in heat sealed polyethylene vacuum bags or boiling tubes fitted with rubber bungs. Desired packing densities were achieved by varying the length of time spent under vacuum. A roughly comparable herbage density to that in the boiling tubes (0.625g cm−3_{) was identified and used for comparison to the boiling tubes. LA accumulation and pH decline} was more rapid at higher packing densities, but was very similar for glass tube and vacuum packed silos. After 35 days, LA and pH were similar for both silo types at all packing density treatments. Effluent production was greater at higher vacuum settings, due to crushing of the plant material.

Differences exist between conditions in sealed fixed volume vessels and vacuum packed bags. Vacuum packed bags leave very little residual oxygen and so anaerobic conditions are achieved more rapidly. While it is easier to achieve similar packing densities between vacuum packed silo replicates at the beginning of ensiling, the volume of vacuum bags can increase significantly during the fermentation phase, due to fermentation gas production. The resulting loosening of the herbage is not observed in rigid walled vessels such as glass tube. In the above experiment, the gradual loosening of herbage did not affect fermentation parameters (Johnson et al., 2005). Vacuum bags may differ in their permeability to oxygen and carbon dioxide, however Ashbell et al. (2001) confirmed that there were no significant differences in fermentation and nutritional parameters in silages made with a number of different vacuum bag types with different permeability to these gases.

Hoedtke and Zeyner (2011), compared glass jar and vacuum packed silages for both fresh and wilted PRG (15.1 and 28.6% DM respectively) and remoistened coarsely ground rye grain, using the silage fermentation profile and aerobic stability as criteria of fermentation quality. Polyethylene bags were air- evacuated in order to achieve the desired packing density of the plant material (equivalent to the glass jar). An attempt was made to constrain herbage within vacuum bags, by first placing plant material in an inner bag, wrapping adhesive tape tightly around this bag, puncturing it with a disinfected needle, and

then placing into larger vacuum bags which were air-evacuated. This meant that the plant materials were held at high packing density for the duration of their fermentation. The authors argued that these silos more accurately modelled the high density storage conditions of a crop in a farm-scale silo. As judged by fermentation parameters, the two ensiling methods were broadly comparable (Hoedtke & Zeyner, 2011). Mean pH, AA, propionic acid and ethanol content across the three crops (with or without a commercial inoculant) did not differ significantly, while LA content was higher and BA content lower, in the vacuum bag silages (Hoedtke & Zeyner, 2011). These results suggested that the material in the double bagged vacuum packed silages may have undergone a better fermentation than the material in the glass jars. Regardless of ensiling method, all of the non-inoculated ryegrass silages failed to reach the pH required for successful preservation, as indicated by a rise in pH and decrease in LA between day 8 and 49 and 90 of the fermentation.

2.7.2 Silage sample size

Few studies have addressed how much plant material is sufficient in order for small-scale modelling exercises to be useful. A theoretical lower limit will depend on how closely the researcher wishes to mimic a certain farm scale silo, and on experimental design. It will also depend on which measurements are made in order to evaluate the success of ensiling. Smaller quantities may be adequate if factors important to the fermentation process are controlled as accurately as possible.

An early silage experiment using glass tubes reported using 25g quantities of finely chopped, well mixed ryegrass (Allen et al., 1937). They found that with rare exceptions, there was consistency between replicates in chemical and bacteriological data. It is frequently quoted that 50g is the lowest quantity used for laboratory silages (McDonald et al., 1991). Vacuum packing as an ensiling method offers greater flexibility in modifying sample size, but this method has not yet been employed for quantities less than 100g to the author’s knowledge. Techniques for further ‘scaling down’ of the fermentation process have also been described by Johnson et al. (2004).

The effect of silage sample size on corn (Zea mays) fermentation in vacuum packed bags was studied by (Cherney et al., 2004), who concluded that 200g is the smallest useful sample size. In this study, 50g corn silos differed in their fermentation profile from 200-600g silage sample sizes, with higher LA, AA and NH3:N, and were deemed non-representative of typical corn silage (Cherney et al., 2004). Fermentation in the 100g sample size also differed from the larger sample sizes, but to a lesser extent. The reason for this was not apparent, but the authors suggest that it was unlikely due to sub-sampling from the fresh

corn samples. Other reports indicate that achieving a representative ratio of plant organs (ear:stover ratio) is difficult with corn. Interestingly, pH and the LA to AA ratio were consistent across all silage sample sizes.

Due to their lower WSC:BC ratio, perennial grasses are more difficult to ensile than corn. However, it is easier to obtain small representative samples of grasses due to their small stems and tillers, and lack of significant grain component. Hoedtke and Zeyner (2011), advocated a minimum 400g sample size for double bagged vacuum packed grasses. Using non-wilted tall fescue (Festuca arundinacea), Cherney et al. (2006) found no difference in pH after 30 days of fermentation between ensiled 250g and 500g samples across two different harvest and chopping treatments. A quantity of 100g has been used for vacuum packed grass, with low variation among replicates (Johnson et al., 2005), although no comparison was made between these silages and larger sample sizes.

Fermentation analyses and silage size

An important question related to the discussion of adequate silage sample size is ‘which measurements are being made in order to assess the type of fermentation that has occurred?’ When sampling from the face of a silage stack, large variation exists in, for example, DM and LA concentration, compared to pH and total nitrogen. Minimum sample numbers vary depending on the silage constituents chosen for measurement (Haslemore & Holland, 1981). It logically follows that when analyzing laboratory silages, smaller silage sample sizes may be satisfactory to measure may be sufficient for measuring certain parameters, such as pH.