observations. At the initial development stage, the biomass was composed more of loosely clumped sludge, which can easily break up into pieces under vigorous shaking. Within a week, the anaerobic seed granules underwent morphological changes from spherical in shape and black in color with average diameter of 1 mm into smaller grey granules due to exposure to the shear force during the aerobic react phase. On day 30, two different types of granules were clearly observed in the reactor as shown in Figure 4.

Figure 4a shows mainly irregular-shaped with yellow colored biogranules (Type A) that are solely developed from the activated sludge. In Figure 4b, the anaerobic granules that have fragmented into smaller pieces have formed different sizes of biogranules (Type B) containing pieces of anaerobic granules. The outer layer of the latter were yellow in color indicating the domination of aerobic or facultative microorganisms while the darker spots within the granules indicate the presence of anaerobic fragments originated from the anaerobic granules. The formation of Type A biogranules can be elucidated by the mechanisms explained by Beun et al. (1999). The development was initiated from the mycelial pellets that were retained in the reactor due to high settling velocity. These mycelial pellets eventually become the support matrix for the bacterial growth. Bacteria that were able to attach to this matrix were retained and suppressed the growth of filamentous microorganisms and became the dominant species in the reactor.

The formation of Type B granules has been discussed by Linlin et al. (2005). These biogranules were formed through a series of physical and morphological changes. The anaerobic granules initially disintegrated into smaller size flocs and debris when exposed to aeration forces in the reactor column. Some of the granules and debris that were too small were washed out with the effluent while the heavier ones were retained in the column and acted as nuclei for the formation of new granules. Having these combinations of aerobic and anaerobic portions within the biogranules will increase the possibility of complete degradation through the anaerobic/aerobic degradation process. Figure 5 shows the obvious morphological differences between sludge particles with average sludge particles of 0.02 ± 0.01 mm (Figure 5a) during the initial stage of the experiment and matured biogranules (Figure 5b) at the final stage with average diameter of 2.3 ± 1.0 mm.

The microstructure of the biogranules was examined using SEM (Figure 6). The SEM observation of the mature biogranules shows the domination of non-filamentous coccoid bacteria. The bacteria are tightly linked and embedded to one another and form a rounded shape on the surface of the biogranule and covered with extracellular polysaccharides substances (EPS) (Figure 6a). Figure 6b shows the presence of cavities between the clumped bacteria. These cavities are anticipated to be responsible to allow a smooth mass transfer of substrates or metabolite products into and out of the granules (Tay et al., 2003 and Toh et al., 2003).