Various reactor configurations, viz., suspended growth, biofilm/packed-bed/fixed bed, fluidized bed, expanded bed, upflow anaerobic sludge blanket (UASB), gran­ular sludge, membrane based systems, immobilized systems, etc., have been used successfully to produce H2 by fermentation processes. Biofilm/attached-growth sys­tems are generally robust to shock-loads compared to the corresponding suspended growth systems, with the biofilms acting as a buffer to reduce the effective concen­tration of toxic chemicals to which the organisms are exposed, protect the culture from predation, provide improved reaction potential due to the presence of high cell densities and provide resilience and resistance to change in the process parameters [26, 92-95]. Generally bacteria achieve maximum growth rates in biofilm resulting in improved reaction potential finally leading to stable and robust system which are well suited for treating highly variable wastewater. Cell-immobilization approaches and granular processes also showed good H2 production efficiency.

Various modes of reactor operation viz., batch, fed-batch, semi-batch/continuous, periodic discontinuous batch (sequencing batch operation) and continuous have been used to produce H2. About a 25% improvement in H2 production and sub­strate degradation efficiency was reported with batch mode operation compared to the corresponding continuous mode operation [92]. The efficacy observed in fed — batch mode operation might be attributed to the reduced accumulation of soluble metabolic intermediates formed during acidogenic fermentation due to fill-draw mode operation [24, 26, 31, 38, 92]. A fed-batch mode of operation with acidic pH showed highest H2 production [92]. Poor biomass retention/cell washout encoun­tered during continuous mode operation can be prevented to some extent with a batch mode operation [92, 96, 97]. Batch mode operation coupled with a biofilm configuration combines the operational advantages of both systems and helps to maintain stable and robust cultures suitable for treating highly variable wastewater [21-25, 98-100].

Morphologically similar bacteria were observed in the scanning electron microscopy (SEM) image [26] of the biofilm formed on the fixed-bed of bioreac­tor producing H2 from the treatment of chemical wastewater (Fig. 4a). The biofilm reactor was inoculated with selectively enriched H2-producing consortia and oper­ated under an acidic microenvironment for more than 300 days. SEM imaging visualized slightly bent, scattered and short chain rods (predominant) along with a relatively low frequency of cocci shaped bacteria of approximate length of 10 p, m. SEM images of isolated bacteria strains from a biofilm reactor (acidogenic mixed culture) (Fig. 4b, c) visualized slightly bent, rod shaped, thick fluorescent capsid bacteria with (~10 ^m in length). Images of both the isolated strain and mixed consortia showed comparatively similar morphology demonstrating the presence of related groups of bacteria proliferated in the bioreactor producing H2. Transmission electron microscopy (TEM) image showing sub-cellular structures of the isolated bacteria from an acidogenic mixed culture [26] (Fig. 4d). TEM image showing oval centralized spore formation with sub-terminal endospore development in rod shaped bacteria (1-7 ^m in length). Terminal bulging with granulose accumulation was not observed. Flagellum attached subapically to the bacterium (two times length of the cell body) was observed. Vegetative cell surrounded by thick membrane (peptido — glycan layer) with two layers (inner and outer forming fibrillar capsule structure) on the cell surface was also visualized (Fig. 4e).