[This corrects the article DOI: 10.1016/j.aquaculture.2023.739932.].

Open net pen saltwater aquaculture faces criticism due to the potential transmission of pathogens between fish farms and to wild stocks. To address this issue and improve the sustainability and growth of net pen farming, closed containment farms have been suggested, but the cost and feasibility of disinfecting large volumes of water in these types of farms is problematic. We explored the potential for using electrolysis to disinfect saltwater in a flow-through system with water flow velocities between 47 and 105 cm/s. This was the first step to investigating whether this technology could be applied to saltwater flow-through closed containment systems. Various voltage levels (3.3-9.0 V) were applied to generate chlorine from saltwater. We found the disinfection properties of the system varied with wattage (i.e., voltage x ampere), velocity of water flow over the electrodes, salinity of water, and residual chlorine contact time. Wattage was highly correlated with the production of chlorine, and this relationship was dependent on water flow (p = 0.0398). A slower flow velocity led to higher chlorine concentration, and the effect was more pronounced at higher wattages. Using a zero-inflated negative binomial regression model, we found the probability of full disinfection was increased by increasing wattage (p < 0.001) and the residual chlorine contact time (p < 0.001). The level of disinfection (count model) suggested the number of bacteria in the treated samples was determined by the interaction between wattage and flow (p = 0.0056) and the interaction between wattage and salinity (p < 0.001). The bacterial count was also associated with residual chlorine contact time (p < 0.001). The results of this study, although preliminary and limited in their scale, offering a potential solution for disinfecting large volumes of seawater, which could make closed containment fish farming in the ocean viable for reducing bacterial transmission within a farm and to wild fish stocks.

Non-neoplastic thyroid hyperplasia is common in terrestrial animals, secondary to nutritional imbalances or other goitrogenic compounds. Thyroid hyperplasia is relatively common in teleost fish; however, malignant thyroid neoplasia is rarely reported. We diagnosed cases of thyroid neoplasia in a population of jade perch (Scortum barcoo). The 3,000 affected fish had grossly apparent, bilateral pharyngeal swellings. Histologic examination confirmed proliferative thyroid lesions ranging from hyperplasia to well-differentiated follicular cell carcinoma. In addition, the younger population of animals on the farm also had bacterial septicemia and mild Dactylogyrus sp. gill infections. Feed analysis revealed a severe deficiency of iodine and vitamin C in the homemade fish diet used on the farm. The concentrations of other minerals, such as zinc, were also on the lower end of the recommended requirements for freshwater fish. The farm was using surface water in its recirculating aquaculture system. We recommended a switch to a commercial aquaculture diet, as well as to use well water rather than surface water to avoid any contaminants, and to treat the younger fish with an antibiotic for bacterial septicemia. Our case provides evidence of progression from nutritional-associated thyroid hyperplasia to neoplasia in farmed teleost fish.

Context. Farming black soldier fly larvae (BSFL) has become an emerging agricultural sector for upcycling food waste into high-quality protein and oil biomass. Depending on the chemical composition of the food waste, the oil extracted from BSFL can reach high levels of lauric acid, a natural antimicrobial compound. Aims. We aimed to evaluate whether feeding a bakery waste-based substrate can enhance the lauric acid synthesis of BSFL. In addition, we aimed to test the antibacterial activity of these BSFL oil extracts against E. coli in vitro. Methods In a 12-day-long feeding trial, we reared BSFL either on a ground corn-based control substrate (n = 6) or a substrate based on bread waste sourced from a local bakery in Hong Kong (n = 6). We studied the growth performance, crude fat and lauric acid content, and agar well-diffusion assay-based antibacterial activity of oil samples extracted from the larvae. Key results. The BSFL reared on the bread waste had a higher average weight (P < 0.001) and average daily gain (P < 0.001), similar crude fat content (P = 0.17), and higher lauric acid content (P < 0.001) than did the control larvae. Both oil samples moderately inhibited the growth of E. coli in vitro; the inhibition zones were 1.92 +/- 1.0 mm and 1.25 +/- 0.75 mm (means +/- s.d.) big in the bread waste-based and the control oil samples respectively (P = 0.28). Conclusions. Our results indicated that locally sourced bread waste is a suitable substrate for farming BSFL, providing a sustainable alternative for waste management. Implications. The inclusion of locally produced BSFL oil as a feed ingredient for the local livestock farming sector might be an avenue not only to achieve circular economy, but also to reduce antimicrobial drug use. However, the effects on animal health and productive performance warrant further in vivo experiments.

Given the close contact between animals, animal products, and consumers in wet markets, fresh meat products are considered a potential source and disseminator of antimicrobial-resistant (AMR) bacteria near the end of the food chain. This cross-sectional study was conducted to estimate the prevalence of select AMR-E. coli in fresh chicken meat collected from wet markets in Hong Kong and to determine target genes associated with the observed resistance phenotypes. Following a stratified random sampling design, 180 fresh half-chickens were purchased from 29 wet markets across Hong Kong in 2022 and immediately processed. After incubation, selective isolation was performed for extended-spectrum β-lactamase producing (ESBL), carbapenem-resistant (CRE), and colistin-resistant (CSR) E. coli. The bacterial isolates were identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). Disc Diffusion was used to determine the susceptibility of ESBL- and CRE-E. coli isolates. The broth microdilution method was used to determine the minimum inhibitory concentration of CSR-E. coli. Targeted resistance genes were then detected by PCR. The prevalence of ESBL-E. coli and CSR-E. coli were estimated at 88.8% (95% CI: 83.4–93.1%) and 6.7% (95% CI: 3.5–11.4%), respectively. No CRE-E. coli isolate was detected. The blaCTX-M-1 gene was the most common β-lactamase group in isolated E. coli (80%), followed by blaTEM (63.7%); no blaSHV gene was detected. Forty-five percent of the isolates had blaTEM and blaCTX-M-1 simultaneously. The mcr-1 gene was detected in all 12 CSR isolates. Of 180 meat samples, 59 were from Mainland China, and 121 were locally sourced. There was no statistically significant difference in the prevalence of ESBL- and CSR-E. coli between the two sources. Our findings can be used to inform food safety risk assessments and set the stage for adopting targeted control and mitigation measures tailored to the local wet markets.

Immersion vaccination, albeit easier to administer than immunization by injection, sometimes has challenges with antigen uptake, resulting in sub-optimal protection. In this research, a new strategy to enhance antigen uptake of a heat-inactivated Vibrio harveyi vaccine in Asian seabass (Lates calcarifer) using oxygen nanobubbleenriched water (ONB) and positively charged chitosan (CS) was explored. Antigen uptake in fish gills was assessed, as was the antibody response and vaccine efficacy of four different combinations of vaccine with ONB and CS, and two control groups. Pre-mixing of ONB and CS before introducing the vaccine, referred to as (ONB + CS) + Vac, resulted in superior antigen uptake and anti-V. harveyi antibody (IgM) production in both serum and mucus compared to other formulas. The integration of an oral booster (4.22 x 108 CFU/g, at day 21-25) within a vaccine trial experiment set out to further evaluate how survival rates post exposure to V. harveyi might be improved. Antibody responses were measured over 42 days, and vaccine efficacy was assessed through an experimental challenge with V. harveyi. The expression of immune-related genes IL1 beta, TNF alpha, CD4, CD8, IgT and antibody levels were assessed at 1, 3, and 7-day(s) post challenge (dpc). The results revealed that antibody levels in the group (ONB + CS) + Vac were consistently higher than the other groups post immersion immunization and oral booster, along with elevated expression of immune-related genes after challenge with V. harveyi. Ultimately, this group demonstrated a significantly higher relative percent survival (RPS) of 63 % +/- 10.5 %, showcasing the potential of the ONB-CS-Vac complex as a promising immersion vaccination strategy for enhancing antigen uptake, stimulating immunological responses, and improving survival of Asian seabass against vibriosis.

Water-borne viral diseases are a significant concern for public health. In particular, they threaten the health of people and animals in countries that lack proper water treatment facilities. Novel water treatment technology may efficiently improve water quality and prevent the spread of waterborne viral pathogens. Laser-induced graphene (LIG) has been shown to inactivate viruses and bacteria with its photothermal properties, electrochemical reaction, and rough surface texture. However, LIG's activity to prevent virus transmission via contaminated water has not been fully explored. Here, we demonstrated that enveloped and non-enveloped viruses in seawater could be rapidly inactivated by LIG technology. After being activated by 3 V of electricity, the LIG electrodes inactivated both types of viruses spiked in water within 30 min. In addition, the electrolyzed seawater exhibited virucidal effects even after the cessation of the electrical charge. The generation of different oxidants, such as chlorine, chlorine dioxide, and hydrogen peroxide, may play an essential role in the antiviral mechanism of the LIG electrodes. Furthermore, after 10 min of electrolysis, the pH of the seawater dropped from approximately 8–5, which may also have contributed to the virucidal effects of the LIG technology. The virucidal activity of LIG technology highlighted its potential for preventing the spread of viral infections via seawater systems which may have public health implications in areas where seawater is used in the sewage system. It may also have applications in aquaculture, where viral diseases do not have treatments and can cause high fish mortality.

The rapid growth and intensification of aquaculture industries have led to an increased use of antibiotics. Consequently, growing concerns have mounted over the environmental contamination of these drugs from medicated feeds and the risk that this poses for antimicrobial resistance. To circumvent environmental leaching, farmers topcoat medicated feeds with oil; however, this only partially addresses the issue. This study investigated the potential of food-grade pregelatinized corn starch (PGS) as a second top-coating agent to reduce oxytetracycline (OTC) leaching from the hand-mixed medicated feed. We immersed top-coated medicated feeds for different periods of time and measured the concentration of OTC in the water to determine leaching. We found a significantly lower level of OTC in water samples collected from the PGS-coated medicated feed compared to the non-PGS-coated medicated feed, with concentrations of OTC approximately 4 and 2.6 times the latter after 5 min and 2 h of water immersion, respectively. We also fed PGS-coated antibiotic feed to jade perch to determine if fish accepted the top-coating and whether they absorbed the OTC. Results from a feeding trial suggested no difference in palatability between PGS and non-PGS-coated medicated feed. We also found that muscle tissue from fish fed with the aforementioned diets had similar levels of OTC concentrations, suggesting that PGS coating does not alter the gastrointestinal absorption of this medication. From our experiment, we conclude that PGS is potentially a new top-coating agent to reduce leaching in hand-mixed OTC medicated feed.

The microbial composition and diversity in aquaculture pond ecosystems are closely related to animal productivity and disease outbreaks. Interventions that alter the bacteria found in pond water can therefore affect the productivity of these systems. Ozone nanobubbles have recently been shown to reduce pathogens, improve dissolved oxygen, and influence fish innate immunity. However, little is known about the effect of nanobubble treatment on the microbial community of aquaculture ponds. This study investigated the impact of ozone macrobubbles (O3MB) and nanobubbles (O3NB) on the microbial ecology of pond water and fish health. We successfully eliminated between 90.9 and 99.4% of the heterotrophic bacteria and 90.1 to 95.2% of the bacterial DNA in our small pond water ecosystems after treatment with 0.15 mg/L ozone. According to the shotgun metagenomic sequencing, ozone macro- and nanobbuble treatments reduced the relative abundance of all bacteria in our water sample, including the dominant bacterial species, as well as Cyanobacteria. The top ten bacterial species in the community changed and were more evenly distributed within the water sample. The bacterial richness of the ozone-treated water samples declined slightly, but over 6000 species were still identified 24 h after the treatment. We also observed a rebound in the bacterial community 24 h after the ozone treatments. The advantage of the nanobubble delivery of ozone over macrobubble delivery of this gas was that the former took significantly less time to deliver the desired quantity of gas while it also greatly increased the dissolved oxygen in the water. Further, we assessed the impact of ozone nanobubbles on jade perch, and no effects were found on the fish at an exposure dose of 0.15 mg/L. This study provides preliminary information on potential applications of nanobubble technology for “resetting” microbial communities, which may be useful during disease outbreaks.

The treatment of Nile tilapia with ozone nanobubbles (ONb) prior to vaccination with an immersible heat-killed Streptococcus agalactiae ( Sa ) vaccine has been reported to modulate and enhance both innate and specific immunity. The efficacy of this novel vaccination strategy is explored further in field trials. This strategy involved a short-term treatment of ONb to activate the fish’s immunity prior to immersion vaccination during their transportation in oxygenated plastic bags (VAC in BAG), followed by two oral boosters during the grow-out stage mixing vaccine in feed (VAC in FEED). The field trial was conducted over 112 days in open cages, comprising four groups: normal aeration control (AC), normal aeration + vaccine (AV), ONb control (NC), and ONb + vaccine (NV). The efficacy of the vaccine was evaluated by measuring specific antibodies for S. agalactiae , monitoring expressions of IgM and IgT transcripts in the gills and head kidney every two weeks, and a laboratory pathogen challenge. Results found that fish in the NV group had significant increases in anti- S. agalactiae antibodies after the primary dose, whereas fish in the AV group required an oral booster dose to produce significant anti- S. agalactiae antibodies. In the vaccinated groups (AV and NV), only IgM was observed to be upregulated at 14 days post-immersion (dpi), while this gene was upregulated in both gills and head kidney in the NC group. No statistically significant upregulation of IgT was recorded in any group at any time point. Despite a decline in the levels of specific antibodies among the vaccinated groups at the time of challenge (88 dpi), the NV and AV groups demonstrated a relative percent survival (RPS) of 50% and 46.7%, respectively, following a relatively high injection dose of S. agalactiae injection (0.1 mL of 10 8 CFU/mL). In summary, this ONb, VAC in BAG and VAC in FEED vaccination strategy represents a promising alternative to the undesirable handling and costly injection approach used within the Nile tilapia industry.

Bacterial infections account for one of the major causes of disease losses in aquaculture. Antibiotics are the most common method used to mitigate these infections, but over the last few decades this has given rise to antimicrobial resistance so finding alternatives to these treatments is imperative. Here we report a drug-free cost-effective method for rapid and safe water disinfection. A pair of laser-induced graphene (LIG) electrodes charged with low voltage (2 V) rapidly inactivated Escherichia coli and Bacillus subtilis in various circulating saltwater systems without significant changes in water quality parameters (pH, dissolved oxygen, and temperature). Bacterial inactivation was enhanced with increasing water salinity during electrochemical disinfection using LIG electrodes. Meanwhile, the concentrations of oxidants such as hydrogen peroxide and ozone were generally low regardless of water salinities, and chlorine was not detectable during the treatments. No health impacts were observed in Japanese medaka exposed to eight days of 1 h LIG electrochemical treatments applied twice a day. Our findings suggest that rapid disinfection of saltwater could be achieved using LIG electrodes without negative health impacts on fish, providing potentially an efficient and safe method for controlling bacteria in saltwater systems.

Laser-induced graphene (LIG) has gained popularity for electrochemical water disinfection due to its efficient antimicrobial activity when activated with low voltages. However, the antimicrobial mechanism of LIG electrodes is not yet fully understood. This study demonstrated an array of mechanisms working synergistically to inactivate bacteria during electrochemical treatment using LIG electrodes, including the generation of oxidants, changes in pH-specifically high alkalinity associated with the cathode, and electro-adsorption on the electrodes. All these mechanisms may contribute to the disinfection process when bacteria are close to the surface of the electrodes where inactivation was independent of the reactive chlorine species (RCS); however, RCS was likely responsible for the predominant cause of antibacterial effects in the bulk solution (i.e., ≥100 mL in our study). Furthermore, the concentration and diffusion kinetics of RCS in solution was voltage-dependent. At 6 V, RCS achieved a high concentration in water, while at 3 V, RCS was highly localized on the LIG surface but not measurable in water. Despite this, the LIG electrodes activated by 3 V achieved a 5.5-log reduction in Escherichia coli (E.coli) after 120-min electrolysis without detectable chlorine, chlorate, or perchlorate in the water, suggesting a promising system for efficient, energy-saving, and safe electro-disinfection.

Oxygen nanobubble (NB-O2) technology has been introduced to the aquaculture industry in recent years. This treatment usually results in a tremendously high level of dissolved oxygen (DO) in the water. However, little is known about the possible negative effects of hyperoxia due to NB-O2 treatment (hyper-NB-O2) on farmed fish. Here, we investigated i) the effect of short-term hyper-NB-O2 exposure (single treatment) on the innate immunity in Nile tilapia, Oreochromis niloticus, and ii) the effect of long-term hyper-NB-O2 exposure (26-day treatments) on survival, growth performance, gill histology, and gut microbiome in Nile tilapia. A single treatment with NB-O2 for 10 min in 50 L of water resulted in 24.2 +/- 0.04 mg/L DO (approximately 2-3 x 107 nanoscale oxygen bubbles/mL). This treatment did not result in differences in expression of several immune-related genes (e.g., TNF-alpha, LYZ and HPS70) in various tissues (e.g., gill, head kidney, and spleen) compared to the non-treated control. Over a 26-day period of exposure, no significant differences were observed in survival and growth performance of the fish, but minor histological changes were occasionally noted on the gills. Analysis of the gut microbiome revealed a significant increase in the genera Bosea, Exiguobacterium, Hyphomicrobium, and Singuli-sphaera in the group receiving NB-O2. Moreover, no signs of "gas bubble disease" were observed in the fish throughout the duration of the experiment. Overall, these results suggest that both short-and long-term hyper-NB-O2 exposure appears to be benign and has no obvious adverse effects on fish.

A carp disease with clinical signs of acute gill rot (AGR) has frequently occurred in Henan province of China since 2013, leading to substantial economic losses. However, the causative agent of virus-related carp AGR (CAGR) disease has been controversial. Our study showed that carp edema virus (CEV), specifically the sub-genogroup IIa, might be the primary agent causing virus-related CAGR that is endemic to Henan, China. This conclusion is based on a cohabitation experiment using isolates from an outbreak in 2019 and a retrospective analysis of CAGR samples collected between 2013 and 2015 in Henan, China. In the cohabitation experiment, the clinical development and pathological changes of CEV disease were observed. The pathological alterations in the fish brain were first noticed to be strongly associated with clinical signs of being sleepy, suggesting that pathological alterations in the fish brain may be a critical differential characteristic of CEV. Another important finding in our study was that the mortality in common carp and koi carp was strongly associated by a temperature change and less correlated to the viral loads in organs. Considering the frequent outbreaks of virus-related CAGR disease in Henan (China), effective prevention, good environmental stress management, and efficient diagnosis are needed. Based on comparing the qPCR and the nested PCR designed at the CEFAS Weymouth Laboratory, we recommend using the qPCR method on gills to identify this pathogen because of the assay's sensitivity and the viral load in gill tissues.

Microcystis sp. is a harmful cyanobacterial species commonly seen in earthen ponds. The overgrowth of these algae can lead to fluctuations in water parameters, including DO and pH. Also, the microcystins produced by these algae are toxic to aquatic animals. This study applied hydrogen peroxide (7 mg/L) to treat Microcystis sp. in a laboratory setting and in three earthen pond trials. In the lab we observed a 64.7% decline in Microcystis sp. And in our earthen pond field experiments we measured, on average, 43% reductions in Microcystis sp. cell counts within one hour. The treatment was found to eliminate specifically Microcystis sp. and did not reduce the cell count of the other algae species in the pond. A shift of the algae community towards the beneficial algae was also found post-treatment. Lastly, during the pond trials, the gill status of Tilapia and Giant tiger prawn were not affected by the H2O2 treatment suggesting this may be a good mitigation strategy for reducing cyanobacteria in pond aquaculture.

East and Southeast Asia have the largest share of mariculture production in Asia, and most of this is cage farming. An increase in marine fish consumption in the region has led to an expansion of marine cage culture over the last decade, which has resulted in an increase in the incidence of infectious diseases within this sector. Knowledge about pathogens, their distribution and clinical signs can help establish appropriate preventive and control strategies. We systematically reviewed the literature for infectious diseases affecting fish in marine cage culture in East and Southeast Asia, using two scientific databases, Scopus and Aquaculture Compendium, as well as two government-based databases, quarterly aquatic animal disease reports published by the Network of Aquaculture Centres in Asia-Pacific, and the fish health data published by the World Animal Health Information System (OIE). We also discussed the transmission routes of pathogens and potential environmental triggers for outbreaks when the information was available. Our review revealed that Vibrio spp., followed by parasitic monogeneans, trematodes and copepods are the most commonly reported pathogens in the literature. There were a few reports of viral pathogens, such as iridoviruses and nervous necrosis virus, but these were more commonly reported in the government surveillance documents. The results of this review can help inform surveillance programmes, improve the early detection of disease on farms and identify pathogens of concern for developing preventive and control strategies.

The objectives of this study were to assess the serological response to rabies vaccination in Hong Kong macaques and provide evidence-based recommendations for the vaccination interval implemented by the Government of Hong Kong. An inactivated rabies vaccine was administered subcutaneously to captured macaques under a mass sterilization program in Hong Kong. Blood samples from the animals were collected in a 2015 field survey and stored in −80°C freezer. In 2021, the frozen sera from vaccinated animals were prepared and tested for antibodies against the rabies virus using a commercial blocking enzyme-linked immunosorbent assay (ELISA) test. Sixty-five samples were available from the vaccinated macaques that had received at least one dose of the vaccine between 2008 and 2015. The interval between the first vaccination and blood sampling ranged from 21 to 2,779 days (median: 990). Only five macaques had a second vaccination record at the sampling time, all with high antibody levels. Among the remaining macaques, 77% (46/60) were positive for rabies antibodies. No specific association was observed between the post-vaccination period and the antibody titer of these macaques, and no adverse reactions were reported. Although the precise level of protection against a potential challenge with the rabies virus cannot be ascertained, the vaccination elicited rabies antibodies in 87% (21/24) of the macaques tested within 2.5 years of their first vaccination. Our findings indicate the potential benefits of the current vaccination strategy to protect the population from rabies and consequential mandatory culling of all macaques if a natural infection occurs.

Injection of ozone nanobubbles into water reduces bacterial load, improves dissolved oxygen, and modulates the fish innate immune system. Little is known about the effect that nanobubble treatment has on the concentration of viruses in water. This study, investigated the disinfection impact of oxygen and ozone nanobubbles (NB-O-2 and NB-O-3) on an Aeromonas hydrophila-specific phage, pAh6.2TG, a virus lab model. After 5-, 10-and 15-min treatments with NB-O-2, the concentration of phage remained the same, while the same treatment with NB-O-3 eradicated 99.99 to 100% of the phage in the water. Since this phage has been shown to control bacterial in-fections in fish, we further investigated whether NB-O-2 improved the adherence of the phage to the body surface of the fish (i.e. skin mucus, and gills) and phage penetration into fish internal organs, specifically the liver. Nile tilapia, Oreochromis niloticus were used as experimental fish in this study. The results indicated that the number of phages adhered to the skin mucus and gills in NB-O-2 treatment group was 1.07 to 15.0 times higher than in the untreated control group without gas nanobubbles. The phage uptake into fish liver after NB-O-2 treatment increased 1.29 to 4.75 fold compared to untreated control. These findings suggested a plausible application of NB-O-2 treatment for improving efficacy of phage therapy in aquaculture. On the other hand, NB-O-3 application may be useful for disinfection of harmful viruses in culture water, but the application would need to be omitted during phage treatment. This study provides preliminary information on potential applications of nanobubble technology in aquaculture to reduce viral load in the water.

The discharge of untreated effluent containing excessive pharmaceutical chemicals (PCs) from aquaculture farms has caused several negative effects on the aquatic ecosystems. Herein, we used a hybrid system based on two emerging technologies, namely forward osmosis (FO) and nanobubbles (NBs), as an energy-efficient, sustainable, and effective alternative to conventional processes for the treatment and reuse of aquaculture wastewater. The combination of NB technology with FO served as a single-step treatment process for the removal of aquaculture pharmaceutical contaminants. In the hybrid system, the FO membrane removed organic matter, dissolved solids, and pharmaceutical chemical residues from aquaculture effluents with a high efficiency (~98%), whereas NBs functioned as a physical membrane-cleaning agent that enhanced the performance and longevity of the FO membrane. Notably, the results revealed minimal contribution of NBs for the direct degradation of the tested PC i.e. oxytetracycline (OTC), where air and ozone NBs could only oxidize nearly 11% and 30% of the OTC in the water respectively. The relatively higher OTC degradation by ozone NBs was attributed to the ozone NBs-induced reactive hydroxyl radicals (HO & BULL;) that react with OTC for its oxidative decomposition. We believe that the tested hybrid system offers a sustainable solution for aquaculture wastewater treatment as well as the recovery of antibiotics from the wastewater and will play a vital role in the sustainable development of the fisheries industry.