Infectious diseases are one of the main constraints for the operation and expansion of the aquaculture industry. Aquaculture systems have been accused of causing many negative environmental impacts, including water pollution, destruction of mangrove forests, reduction in biodiversity, and salinisation of fresh water . Chemical disinfection is an effective treatment for many types of pathogens, including viruses, bacteria, fungi and protozoan parasites . Use of chlorination, ozone treatment or antibiotics generates potentially toxic by-products and can leave residues which not only affect fish condition but may also pose health risks to the human population .
Water quality is important in determining the success or failure of fish production in aquaculture systems , being one of the aspects that requires careful consideration . Many physical and chemical water quality variables are involved in fish health . These variables can be influenced by each other and by environmental and biological conditions . Therefore, this study investigates the impact of several aspects of water quality on the inactivation of the fish pathogen Aeromonas hydrophila using a solar photocatalytic system under full sunlight. This study reports on the extent of oxygen-sensitive cell injury occurring in a thin-film fixed-bed reactor (TFFBR) with solar photocatalytic disinfection for several important water quality variables. This study also investigates and compares the levels of inactivation of A. hydrophila in filtered and unfiltered aquaculture pond water, to compare results using synthesised and natural waters.
To assess the viability of bacteria during solar disinfection, the conventional approach is to enumerate samples using plate counts on a suitable agar-based growth medium after exposure to sunlight using standard aerobic conditions (e.g. 24 h incubation at a suitable temperature). However, previous studies have demonstrated that ROS, derived mainly from aerobic respiration during the enumeration process, may inactivate sub lethally damaged bacteria and prevent their growth and enumeration under conventional aerobic incubation . Tandon et al. also demonstrated that due to oxygen sensitivity, the enumeration of Enterococcus faecalis on selective media under aerobic condition is not sufficient to count injured bacteria . Two main reasons for oxidative stress during enumeration are: (a) The presence of reactive components in the growth media which occurs either due to oxidation of nutrients during autoclaving or due to photo-oxidation of growth media components after autoclaving. (b) The cellular respiratory process of the growing bacteria on exposure to light. Due to cellular respiration “cell destruction” or “cell suicide” can occur in sub-lethally injured bacteria as their protective mechanism against oxidative stress are damaged and they are incapable of coping with the oxidative burst when they are rapidly growing on nutrient rich medium . Such cells cannot be counted under standard aerobic conditions, but can be cultured under conditions where reactive oxygen species are neutralised (ROS-neutralised conditions), e.g., in growth medium supplemented with the peroxide scavenger sodium pyruvate and incubated under anaerobic conditions to prevent cellular respiration [8, 11]. The significance of this was shown in our recent study using a solar photocatalytic reactor under different flow rates with low sunlight and high flow rates showing substantial sub-lethal injury of A. hydrophila.
pH is a major variable in aquaculture systems; it influences the survival and growth of fish in culture and affects the physiological condition of the end product . Lower pH generally decreases the survival and reproductive maturity of fish, while high pH can cause toxic ammonia imbalance within an aquaculture system . The acceptable pH range for water used in aquaculture production is typically from 6.5 to 9 . In solar photocatalysis, pH is also one of the main variables affecting the process. At higher pH levels, TiO2 surfaces are negatively charged and repulse anionic compounds in water . In contrast, at low pH the density of positively charged catalyst increases which can then form an electrostatic link with the negatively charged surfaces of bacteria, resulting in a higher rate of microbial photo-disinfection . Herrera Melian and his co-workers showed higher bacterial inactivation at pH 5 than at pH 7.8 which is consistent with such proposals . However, Rincon and Pulgarin did not find any differences in bacterial inactivation at pH 4–9 . Consequently, this research investigated microbial inactivation at pH levels of 5, 7 and 9 using the TFFBR system, thereby covering the typical pH range of aquaculture systems .
The salinity of aquaculture pond water is an influential factor for fish survival and growth . Selven and Philip stated that salinity can cause negative effects in aquaculture species, linked to the growth and production of toxins by pathogens . They showed that salinity variation increased the virulence characteristics of Vibrio harveyi in aquaculture systems, reducing the immune response in the shrimp hosts and causing heavy mortality. Wang and Chen showed that 2.5% NaCl significantly increased the growth rate of Photobacterium spp. and that addition of the same amount of NaCl to the growth medium (Tripticase soy broth) also increased the virulence of this pathogen towards shrimps . Seawater has a typical salinity of 3.5% . Therefore, this study investigates the effect of salinity (with and without NaCl and sea salt at 3.5%) on the photocatalytic inactivation of A.hydrophila through the TFFBR system.
Imbalance in an aquaculture pond ecosystems can change the water transparency, due to additional suspended solids . Turbidity refers to the indicator of cloudiness of a water sample due to the presence of suspended materials in the water that affect light transmission ; this can adversely affect fish health. Clays are one of the most common suspended materials present in aquatic systems . Reduced phytoplankton production and increased growth of heterotrophic bacteria in aquatic systems have often been attributed to high clay turbidity levels and low light transmission levels [24, 25]. In relation to solar disinfection, highly turbid water samples at 300 Nephelometric Turbidity Units (NTU), showed reduced microbial inactivation compared to less turbid or non-turbid samples, which may be due to shielding of microbes from sunlight by suspended materials . In batch system solar disinfection, Joyce et al. found that, less than 1% of the total solar UV light would reach a depth of 2 cm in water with a turbidity of 200 NTU . Therefore, EAWAG, the Swiss Federal Institute of Aquatic Sciences and Technology, recommended that water for solar disinfection batch systems need to be not more than 10 cm in depth and a turbidity level of not more than 30 NTU . Rincon and Pulgarin observed that water turbidity negatively affected the photocatalytic inactivation of microbes and resulted in bacterial re-growth, supported by nutrients associated with the suspended particles . They also stated that suspended particles absorb heat from sunlight and warm the water. Warmer water holds less oxygen and consequently affects microbial respiration and photocatalysis. Suspended particles also reduce light penetration capacity by their scattering effect. One recent research study used a batch sequential CPC reactor to eliminate water pathogens, with reduced exposure time and minimal user input compared to other systemsn . However, most of the previous studies of turbidity in solar disinfection have been in batch reactors with TiO2 suspensions, rather than immobilized systems. Another recent investigation has developed a CFD (computational fluid dynamics) model for water disinfection through a CPC pilot-plant reactor . However, no laboratory experiments were evaluated in that study to evaluate its practical efficiency. In contrast to batch reactors and CPC reactor systems, the TFFBR system evaluated in the present study is a single-pass system. The reaction on the surface of the TFFBR reactor is different, as water is not in a static condition. Therefore, this study reports for the first time the use of a single-pass flow-through TFFBR system to investigate the elimination of an aquaculture pathogen from water of different turbidities.
Suspended particles are not the only obstacle to light penetration; dissolved coloured materials also absorb sunlight of different wavelengths . Natural organic matter is present in all surface water; humic acids are major component in natural waters which are brown in colour . Humic acids are generally present in most surface waters at levels of up to 10 mg L-1. These substances may act as photosensitisers under the influence of solar radiation [34, 35]. This can cause oxidative damage to the cell membrane  and also may influence solar photocatalytic degradation via TiO2. Doll and Frimmel showed a reduction in photocatalytic degradation of several chemicals (carbamazepine, clofibric acids and iomeprol) with 2 commercially available TiO2 preparations, in the presence of humic acids, with these substances competing for active sites and causing surface deactivation of the catalyst by adsorption . In contrast, humic acids can also negatively affect solar disinfection by absorbing the radiation that passes through the water and this can decrease solar UV transmission  and reduce cell inactivation [34, 36, 37, 39]. As humic acids have an attraction towards aqueous metal cations, they may be able to interact with microbial surfaces and protect them from solar UV disinfection . Therefore, this study has investigated the use of the TFFBR system to disinfect aquaculture bacteria from water samples containing added humic acids.
Temperature and dissolved oxygen (DO) levels are two important variables in aquatic systems which also influence microbial solar disinfection [29, 34, 40]. However, in this study, the TFFBR is an open system where the temperature of the thin layer of the water cannot be readily controlled and will rapidly change during passage across the reactor plate in full sunlight. During the experiments, the ambient temperature of that day was noted and the reservoir water temperature was maintained. As experiments were performed through a 2 year time period in different seasons, further control of water temperature was not considered. Similarly, water samples used in this research were fully oxygenated due to a combination of (i) mixing [flow/agitation] and (ii) the thinness of the film of water across the photoreactor, at <0.3 mm. Photo-oxidation happens on the TFFBR reactor plate and while residual reactive oxygen species are present in the treated water, they are extremely short-lived with half-lives measured in milliseconds. Therefore, DO levels have not been considered further in this study.