Received: 2 May 2023 Revised: 12 October 2023 Accepted: 18 October 2023

DOI: 10.1111/raq.12870

REVIEW

Emerging technologies revolutionising disease diagnosis and

monitoring in aquatic animal health

Kailash Bohara 1 | Pabitra Joshi 2 | Krishna Prasad Acharya 3 | Grace Ramena 1

1
Department of Aquaculture and Fisheries,
University of Arkansas at Pine Bluff, Pine Bluff, Abstract
Arkansas, USA
In recent years, aquaculture has seen tremendous growth worldwide due to
2
Department of Plant Science, University of
Idaho, Moscow, Idaho, USA technological advancements, leading to research and development of various
3
Animal Quarantine Office, Budhanilkantha, innovations. Aquaculture farmers prioritise early diagnosis for timely treatment to
Kathmandu, Nepal
achieve better productive and economic performance. Aquatic animal health
Correspondence experts still employ traditional diagnostic methods using visual diagnosis, cell cul-
Kailash Bohara, Department of Aquaculture
ture, media culture, histopathology and serology. However, the developments of
and Fisheries, University of Arkansas, Pine
Bluff, AR 71601, USA. technologies in aquamedicine, such as sequencing, biosensors and CRISPR, have
Email: [email protected] enabled rapid disease detection within minutes. Furthermore, integrating sensors,
Funding information drones, artificial intelligence and the internet in aquaculture farm monitoring has
U.S. Department of Agriculture helped farmers take decisive actions to improve production. Advancements in
[The copyright line for this article was changed diagnostic techniques have significantly enhanced the efficient detection of bac-
on 4 June 2024 after original online
terial, viral, parasitic and fungal diseases in aquatic animals. Moreover, monitoring
publication]
water quality, aquatic animal health and animal behaviour on farms has become
exceptionally streamlined with cutting-edge tools like drones, sensors and artifi-
cial intelligence. Summarising research and development in aquatic animal health
and monitoring aids efficient technology adoption in aquaculture. With these
advanced technologies' continued development and adoption in developed coun-
tries, the aquaculture industry is experiencing growth and increased efficiency,
benefiting farmers and consumers in these regions. However, farmers and educa-
tors in developing countries lack information about these technologies. Training
of agricultural educators and efficient dissemination of knowledge and technolo-
gies through advertising and publication in collaboration with companies is essen-
tial. This review delves into emerging technologies capable of replacing the
conventional diagnostic and monitoring methods utilised in aquaculture. We also
explore their strengths, limitations and potential future applications within aqua-
culture settings.

KEYWORDS
aquatic disease, early diagnosis, internet of things (IoT), smart aquaculture, UAVs

836 © 2024 John Wiley & Sons Australia, Ltd. wileyonlinelibrary.com/journal/raq Rev Aquac. 2024;16:836–854.
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
BOHARA ET AL. 837

1 | I N T RO DU CT I O N the pathogen causing the problem. Disease diagnostic is categorised

into two types in aquaculture. Presumptive diagnosis involves visual
In the 21st century, technological advancements have transformed observation of symptoms expressed by the animal using microscopy,
the agriculture industry. The integration of advanced technologies, drones and machine learning approaches. Confirmatory diagnosis
such as robots,1 sensors,2 GPS3 and aerial imaging,4 has led to the involves pinpointing the cause with almost 100% accuracy. Confir-
adoption of precision agriculture, optimising yields and improved man- matory diagnosis is done using biochemical tests, serology and
5
agement decisions more accurately and efficiently. It has had a signif- molecular techniques. Pathogen identification plays a crucial role in
icant impact on improving efficiency, profitability and sustainability. implementing containment strategies promptly. Surveillance
Remote sensing technology maps soil conditions, identifies suitable involves a structured approach to collecting data on the prevalence
crop species and monitors crops for insect or disease problems.6,7 The of significant diseases and pathogens, intending to generate infor-
integration of these technologies has led to more profitable farming mative reports on the disease situation within a specific area. At the
conditions and enhanced food security. “The State of World Fisheries same time, monitoring focuses on overall observation, assessment
and Aquaculture 2022” indicates that the aquaculture production for and collection of data throughout the area with no specific inten-
human consumption was 157 million tons, and it is predicted to tion.34 Moreover, researchers face an ongoing challenge in identify-
increase by 15% by 2030 to meet the growing demand of the increas- ing and monitoring novel pathogens and strains that impact various
ing population.8 However, this increased production must be more aquatic species.35 However, sequencing technologies such as nano-
environmentally sustainable to preserve natural resources for future pore sequencing and other next-generation sequencing methods
generations. Pollution, climate change, farming practices, regulatory can also be used to characterise novel pathogens in aquaculture.
9–11
issues, water quality and diseases affect aquaculture production. On-farm monitoring can be conducted using water quality data,
However, the transformation in technology in aquaculture has led to pathogen loads in water and routine testing of fish species from dif-
more efficient and sustainable production.12 Advanced technology is ferent ponds. Current diagnostics in aquaculture are conducted
being applied in various aspects of aquaculture, including nutrition, through visual observation, biochemical analysis, cell culture and
13
genetics, health and marketing. molecular techniques, which can take days for bacterial diseases while
Breeding techniques that utilise quantitative genetics, such as taking months in case of viral infections, leading to significant losses.36
14 15
genome-wide association mapping and genomic selection, can New diagnostic tools are needed to rapidly and accurately detect
help farmers improve critical traits such as disease resistance, fillet pathogens, preventing further losses. Portable sequencing machines
quality and growth, leading to increased production. In addition, there can sequence DNA or RNA in real time, identifying diseases within
is significant progress in aquaculture nutrition where alternative feed hours, even in remote areas without laboratory facilities.37 Clustered
ingredient meals successfully replace fish meals in diets without Regularly Interspaced Short Palindromic Repeats (CRISPR)-based38
affecting the quality and quantity of fish production.16,17 Live feeds, diagnostic tools and microfluidic chips39,40 are newly developed
probiotics, prebiotics, algae, rotifers and other feed supplements are tools used for rapid and efficient detection of fish diseases, allowing
also used to maintain health and increase production.18 Vaccines for detecting multiple pathogens with a single test and reducing
against disease,19 alternatives to antibiotics,20 medical imaging diagnosis time and effort. Various diagnostic tools have been devel-
techniques,21,22 diagnostic databases23 and gene editing24 are exten- oped since the inception of serology techniques for disease identifi-
sively utilised for fish health research. With the improvement in com- cation (Figure 1.).
puterised models,12 remote sensing,25 artificial intelligence (AI),25,26 Regular health checks are crucial to maintain fish quality and
machine learning27,28 and the Internet of Things (IoT),29–31 freshwater reduce losses due to disease. Additionally, aquatic disease monitoring
and marine aquaculture is revolutionising worldwide.1 However, these and diagnosis are essential for policymakers to develop efficient plans
technologies are mostly limited to high-profit aquaculture industries for management strategies and biosecurity. With evolving protocols
such as salmon, shrimp and tilapia. Farmers and researchers use these and technologies, numerous novel techniques have been developed.
tools to collect, monitor and analyse the data in real time to make However, only a few are employed in commercial aquaculture. The
decisions. main challenges to their adoption include a lack of know-how, eco-
Aquatic ecosystems are subject to frequent fluctuations in water nomic perspectives, technical challenges, regulatory issues and disad-
quality, temperature, sunlight, dissolved oxygen, pH levels and plank- vantages of these technologies. Therefore, keeping up with the latest
ton density. Farmers use sensors to monitor the water quality param- disease diagnostics and monitoring technology is vital for researchers
eters in real time, and any deviation from optimal conditions can be and farmers.41 There are several limitations to making diagnostic tech-
2
detected using cell phones. These changes in water quality parame- nology more efficient and productive.
ters can make aquatic animals more vulnerable to diseases. Fish dis- This comprehensive review highlights the emerging technologies
eases significantly cause economic losses in the global aquaculture in aquaculture disease diagnosis and health monitoring, offering
industry, with annual losses of billions.32 Researchers actively seek readers a profound insight into the transformative potential of these
25
ways to detect and treat fish diseases at early stages while devel- innovations. Readers will also gain a detailed understanding of how
oping strategies to enhance disease resistance by vaccine cutting-edge tools like nanopore sequencing, biosensors, CRISPR,
development.19 Disease diagnostics refers to the identification of environmental DNA/RNA (eDNA/eRNA) analysis, Matrix-assisted
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
838 BOHARA ET AL.

FIGURE 1 A chronological overview of disease diagnostic tool development.

laser desorption ionisation-time of flight mass spectrometry (MALDI- due to high dissolved oxygen in different aquaculture species.43,44
TOF MS), sensors, artificial intelligence (AI) and drones are poised to Additionally, the requirement of pH, ammonia nitrogen, temperature
reshape disease diagnosis and monitoring in aquaculture. This review and light varies with fish species, making it challenging for farmers to
helps connect the ideas from research to their real-life use, showing optimise the parameters according to the species reared. Changes in
how these technologies can make aquaculture more sustainable, effi- these parameters can be easily caused by environmental factors such
cient and profitable. This review also assesses these technologies' as algal bloom, rain, snow, sunlight and human factors. The water
strengths, limitations and future potential in improving disease diag- quality parameters need to be monitored regularly to maximise aqua-
nosis and monitoring. Ultimately, this review acts as a guiding tool for culture production.
researchers, practitioners and policymakers, pointing them towards a Traditionally, water quality monitoring and testing is done in labo-
future where aquatic animal monitoring and diagnosis will be techni- ratories after collecting the samples using the titration method.45
cally and economically feasible. However, with advanced technology, portable and on-farm sensors
can monitor water quality in real time, reducing time and effort.
Contact-based temperature sensors use resistance temperature
2 | EARLY DIAGNOSIS AND MONITORING devices or thermocouples to sense the temperature, while infrared
METHODS thermal imaging utilises the change in electromagnetic radiation to
measure the temperature. Dissolved oxygen sensors are more compli-
2.1 | Sensors cated and use oxygen-permeable membranes to quantify the dis-
solved oxygen in water. Other sensors, such as sensors to measure
Aquaculture, like other forms of agriculture, is susceptible to diseases turbidity, use light reflectance; salinity sensors use water density, light
caused by environmental stress factors that affect the production of refraction and electrical conductivity.
fish, crustaceans and molluscs. Rapid diagnosis is a key factor in sus- Moreover, sensors with wireless technology simplify parameter
tainable aquaculture management. Rapid diagnosis of diseases and monitoring via mobile phones or computers. This technology has been
abnormal water quality can reduce treatment costs and maintain the employed in monitoring the rivers for pollution pathways46,47 and
economic viability of the farm. Additionally, it can also improve animal ocean monitoring.48 In aquaculture, multiparameter sensors with
welfare, enhance product quality, prevent the transmission of diseases remote monitoring are extensively used to gather information on tem-
to wild populations, ultimately protecting the environment and sup- perature, pH and dissolved oxygen.49–52 A water quality monitoring
porting sustainable production. system leveraging Narrow-Band Internet of Things (NB-IoT) technol-
Water quality is a paramount concern for inland and off-shore ogy has been developed to centralise the oversight of ponds, facilitat-
aquaculture farmers. In aquaculture, dissolved oxygen, pH, ammonia ing the efficient and timely detection of any deviations in water
nitrogen, temperature, salinity and light are critical parameters for quality parameters with a relative error rate ranging from 0.15% to
maintaining optimum water quality. These parameters play a crucial 2%.50 Innovasea (https://www.innovasea.com/) has also pioneered
role in determining fish behaviour, health and production as a whole. the development of wireless sensors for assessing dissolved oxygen,
Environmental factors such as abnormal water quality is one of the turbidity, chlorophyll, coloured dissolved organic matter and blue-
triad factors along with pathogens and host that can cause disease in green algae. These sensors transmit data to their cloud-based aquacul-
aquatic species.42 Dissolved oxygen requirement varies with age and ture platform known as Realfish Pro. This high-precision aquaculture
fish species and can cause hypoxia due to low oxygen and hyperoxia platform has been instrumental in remotely monitoring environmental
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
BOHARA ET AL. 839

conditions and implementing timely interventions. Numerous studies three types of unmanned vehicle systems: (1) unmanned aerial
have explored the application of sensor-based cloud technologies in vehicles (UAVs), (2) autonomous underwater vehicles (AUVs) and
managing recirculating aquaculture systems (RAS) and in-land aqua- (3) unmanned surface vehicles (USVs). UAVs are also used to monitor
culture operations on a global scale.53–56 The cost of the sensors and fish farms,66 algal blooms67 and water quality.25 Equipped with sen-
buoys is one of the concerns for fish farmers. Medina et al. designed a sors, UAVs can monitor environmental conditions, feed, equipment
low-cost water quality monitoring buoy costing about $658 to make function, fish abnormality and suspicious activity. Drones can be fitted
57
in-house. An AI-integrated water-quality monitoring buoy was with RGB, multispectral and hyperspectral imaging sensors. Thermal
developed by Lu et al., which cost about $2015, including all the sen- and infrared cameras may also be utilised for specific objectives. For
sors and other hardware.58 example, researchers captured images of harmful algal blooms (HABs)
These sensors have some challenges to operate due to their pres- using a DJI Matrice 200 HD camera on a UAV, then collected water
ence in submerged conditions for extended periods and can become samples from the area to confirm the presence of HAB using Akabot
clogged, therefore needing to be cleaned more frequently. Further- II.68 The images captured using drones are subjected to image proces-
more, wireless sensors face constraints related to their finite battery sing and segmentation to fit into AI and ML models. A recent study
life, and those employed in marine aquaculture are susceptible to cor- has demonstrated the use of UAVs integrated with water quality sen-
rosion. Additionally, sensors submerged in water contend with the sors measuring the water quality indices in larger size fish farms in the
influence of microorganisms, which can distort sensor readings when United States using a hybrid aerial underwater system.1
they colonise in close proximity to the sensors. Another issue arises Feed is one of the most expensive components in aquaculture,
when sensors utilise UV radiation, acoustic beams or magnetic fields, contributing a significant amount of money to the operation. Over-
as these methods can potentially impact aquatic organisms. Sensors feeding and underfeeding can cause water quality and health issues,
integrated with remote data transfer capacity can be expensive and respectively. Optimising the feeding rate according to the growth is
need a reliable internet connection, making it a challenge for small- crucial to reduce the cost incurred. UAVs can be deployed to capture
scale farmers and rural areas. images from different ponds in a farm to understand the feeding
Additional types of sensors utilised in aquaculture include current rate.66,69 Additionally, underwater drones are popular in marine aqua-
and water flow sensors, which can measure the water levels and cur- culture management due to the low turbidity and prevalence of sea
rents in real time. Researchers are also using movement and imaging cage culture.70 Monitoring sea cages is essential to prevent fish from
sensors to monitor fish behaviour. Cameras and imaging sensors escaping and cage clogging.71 Amphibious vehicles equipped with dis-
observe fish growth, health and behaviour. These sensors can provide solved oxygen sensors have been used to reduce the number of sen-
real-time photos and videos of fish behaviour, allowing farmers to sors required in each pond on a large farm.72 If necessary, the drone's
identify potential issues and take corrective action. They can also flight plan can be programmed to cover multiple ponds on a farm
monitor fish feeding and ensure sufficient nutrition for proper growth using set coordinates and maps.1
59
and development. However, in the developing phase, these sensors Da-Jiang Innovations (DJI) is one of the leading multi-rotor drone
are only limited to some RAS systems and cage cultures in oceans. manufacturers worldwide, designing drones for all uses, including agri-
cultural monitoring and surveillance (https://www.dji.com/). These
drones range from $759 to $ 2500; however, based on camera type,
2.2 | Drones it can cost more. DJI Agras series, designed for agricultural operations
such as farm surveillance and spraying, can cost up to $30,000. Fur-
Imaging using unmanned aerial vehicles (UAVs) is a relatively new thermore, American companies like Ag Eagle (https://ageagle.com/)
agricultural concept for farm monitoring.60,61 Previously used for mili- specialise in producing fixed-wing drones and various sensors, includ-
62
tary operations and surveillance, UAVs have gained popularity in ing RGB, thermal and multispectral sensors. On the other hand,
different sectors, including agriculture for surveillance.63 Farm moni- American Robotics (https://www.american-robotics.com/) focuses on
toring is crucial in in-land and off-shore aquaculture management the manufacturing of automated multi-rotor drones.73 Underwater
practices. Larger size aquaculture farm monitoring can be labour- drones are popular in the European aquaculture industry, helping cage
intensive as well as costly. Additionally, cage culture in the open culture farmers monitor cages and fish health efficiently.74,75
ocean is generally accessed via boats or ships, making it hard to moni- Blueye (https://www.blueyerobotics.com/) and SeaSmart (https://www.
tor the cages and well-being of the aquatic species reared. Remotely seasmart.no/) are among the leading companies providing cutting-edge
operated vehicles (ROVs) have been extensively used in aquaculture technology solutions to support the needs of ocean aquaculture farmers.
farm monitoring.21,64,65 With the advent of technology in aquaculture, However, navigating and communicating with AUVs underwater
new tools such as high-resolution cameras on underwater drones or can be challenging due to the geography.76 Additionally, these devices
ROVs can capture images of fish to study their behaviour, growth and can deteriorate quickly due to the saline environment, weather and
health indices. These technologies integrate artificial intelligence water currents.77 In contrast, UAVs are smaller, lightweight and easy
(AI) and machine learning (ML) to detect diseases and parasites. Image to navigate and communicate; however, they have limited flight time
processing technologies interpret external symptoms, enabling real- and are prone to wind and other environmental conditions.78 UAVs
21,64,65
time pathogen detection and saving time. Aquaculture employs are regulated by the Federal Aviation Authority (FAA) in the
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
840 BOHARA ET AL.

United States (https://www.faa.gov/), and to operate an unmanned healthy and diseased salmon by classifying images sourced from the
aerial system, FAA guidelines must be followed by the pilot. FAA has internet. When applied to different captured images, the trained
recently imposed regulations such as using Remote ID in every drone model exhibited an accuracy range of 91%–94%.80 Another group of
79
to identify the drone's location. These regulations are constantly researchers integrated a cloud-based AI model with an autonomous
changing, and it can be challenging for the farmers to keep track of drone to monitor off-shore cages for feeding, fish length, facial recog-
them. Image capturing in aquaculture is a bottleneck due to the lack nition and monitoring. The drone's sensors and cameras collected data
of appropriate ROVs capable of capturing high-quality fish images from the cages, which were transmitted to the cloud-based AI system
underwater. Despite a few disadvantages, drones have been a popular during flight via Wi-Fi, then to operator.66 The combined work of sen-
method for farm monitoring due to the reduced operational cost and sors, ML and AI can be utilised to move towards the decision-making
labour.77 process easily using intelligent devices such as aerators and feeders.
Dissolved oxygen sensors integrated with the controller have been
utilised to maintain the oxygen level in whiteleg shrimp culture using
2.3 | Artificial intelligence (AI) and machine adaptive predictive control.81 ML has also been employed to separate
learning (ML) dead embryos from live ones to prevent fungal infection in rainbow
trout. This ML approach resulted in an average accuracy of 99.45%,
Despite the advantages of drones and sensors in facilitating rapid and suggesting its possible use in commercial aquaculture.82
real-time data acquisition, effectively utilising the gathered data In addition to these technologies, several modern technologies, in
remains a challenge. AI and ML use the learning process by feeding combination with imaging, cloud AI and experts in the field, are being
data and tasks to train the model to execute the decision process. In used for problem diagnosis. Fish-expert is one of the earlier developed
aquaculture, different ML models can be implemented to analyse the diagnosis software that uses more than 400 images of diseased fish as
images without going through manual analysis. Information sourced databases and can diagnose about 126 freshwater diseases.83 In
from individuals, drones and sensors can be seamlessly channelled recent years, Rossi et al. developed FishAPP, a mobile-based software,
into ML and AI platforms through various data transmission mecha- by training the model with fish images captured from a fish market.84
nisms, including wired connections, wireless technologies, mobile This application utilises the smartphone's camera to predict the fish
internet and data storage solutions. The ML models necessitate prior species, achieving nearly 100% accuracy. Web-based expert systems
training, after which processes like feature extraction, segmentation developed85 are now transitioning to AI-based web systems.86 The
and final output generation can be executed (Figure 2). A team of former system uses the images uploaded by the user for diagnosis by
researchers successfully trained a model to distinguish between a human expert. In contrast, the latter uses data from previous

FIGURE 2 A visual representation illustrating the integration of drones and sensors with AI and ML in aquaculture.
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
BOHARA ET AL. 841

diagnostic cases in the system, and AI generates a response based on employed Oxford nanopore sequencing technology to enhance Tilapia
earlier images. Both systems have their advantages and disadvan- Lake virus (TiLV) detection with similarity between Oxford nanopore
tages. However, combining both systems with 3D images and large sequencing and other sequencing methods, such as Illumina and
databases may make them more accurate and precise. The crucial fac- Sanger, was found to be around 99%–100%.103 In addition to viral
tor for accurate disease identification using the captured image is the disease diagnostics, nanopore sequencing has been implemented in
availability of a trained model. However, a constraint is the lack of dif- other pathogenic bacteria and parasites.104,105 Furthermore, nanopore
ferent images specific to the disease for training the model. The symp- sequencing is also utilised for microbiome analysis to gain insights into
toms of different diseases can be similar, making the identification the health status of fish.106 While limited, the use of nanopore
harder. Additionally, image databases for each aquatic species can be sequencing for fish disease detection is growing due to the lack of
challenging to maintain and sometimes cause false positives due to sequenced pathogen variants in aquaculture. Ongoing studies aim to
system errors. The major hurdle in integrating AI and ML using the compare pathogen genomes using this technology.104,107
datasets obtained by drones and sensors in commercial aquaculture is The advantages of this technology include the low cost (as low as
lack of individuals trained in such area. $8/sample), long reads, portability and no requirement for a sophisti-
cated lab.108,109 Although the accuracy of nanopore sequencing cur-
rently limits its commercial use in fish disease diagnosis, it has the
3 | CONFIRMATORY DIAGNOSTIC potential to be a game changer for novel pathogen identification by
METHOD sequencing. Nanopore sequencing is still in discussion for animal dis-
ease diagnosis; however, the Food and Drug Administration (FDA) in
3.1 | Nanopore sequencing United States authorised emergency use of SARS-CoV-2 test kits
based on nanopore sequencing during the pandemic.110 By enabling
Next-generation sequencing (NGS) technology is a valuable diagnostic rapid identification of fish diseases using this tool in remote loca-
tool used to sequence pathogens to understand their evolution87 and tions, outbreaks can be quickly contained, preventing further spread
identification better. It has been successfully applied in disease diag- and economic losses. Nanopore sequencing could greatly enhance
nosis in humans,88 plants89 and animals.90 However, this technology managing and controlling fish infections, especially in isolated areas
presents several challenges, such as its reliance on sophisticated once approved by the regulatory bodies for animal health including
equipment, higher costs and longer turnaround times.91 Oxford World Organisation for Animal Health (WOAH) and United States
Nanopore Technologies introduced a new third-generation sequenc- Department of Agriculture-Animal and Plant Health Inspection
ing technology in 2014 that overcomes the challenges of NGS by pro- (USDA-APHIS).
viding long and accurate sequencing reads in real time using a small,
portable device weighing just 90 g.92 This technology uses a flow cell
sequencing chip to add a sequencing library. The DNA/RNA passes 3.2 | Biosensors
through the nanopore triggering variable current, which is then
decoded into base pairs, resulting in reliable, on-site results. In recent Biosensors use biological elements such as enzymes and antibodies to
years, nanopore sequencing has been utilised in diverse fields, includ- detect target substances, causing changes in parameters like current,
93
ing disease diagnosis. Nanopore sequencing was used as a real-time resistance or heat. These biocatalysts can produce or consume chemi-
surveillance tool for Ebola outbreak94 and Zika virus in Brazil.95 This cal substances when interacting with the target substance, making
technology offers a portable solution for sequencing DNA or RNA, them useful for identification and measurement.111 A biosensor can
requiring minimal equipment, and is practically beneficial in remote rapidly detect small signal changes to detect the organism (Figure 3).
95,96
areas. Biosensors have been researched for many years. However, their
Aquaculture is one of the fastest growing agricultural industries,8 commercial application is limited to specific pregnancy tests and glucose
yet it is threatened by different diseases and parasite outbreaks.97,98 biosensors due to regulatory issues, sensitivity, commercial viability and
Nanopore sequencing is a promising tool for aquaculture to sequence coordination between biological researchers and engineering. There are
99–101
genomes of various fish species and pathogens affecting aqua- different biosensors based on receptors, including enzymes, antibodies,
culture, including novel pathogens. Nanopore sequencing was suc- aptamers, whole-cells and nanoparticle biosensors,112 which increase
cessfully utilised to sequence the novel subtype-3 lineage of salmon the efficiency of the biological reaction. Biosensors are used in the
alphavirus, identifying a previously unrecognised strain within the food industry113 to detect contamination and pollutants in the
102
alphavirus family. It can detect pathogens easily without requiring environment,114 and for disease diagnostics.115 In aquaculture, biosen-
the samples to be taken to a laboratory. Rapid and accurate identifica- sors are used to detect cyanobacteria, cyanobacterial toxins in water
tion of viral pathogens can help with prompt planning and manage- and disease diagnostic.116,117
ment to prevent future outbreaks. Nanopore technology has been Biosensors have various applications in detecting stress and path-
shown to accurately identify Salmon alphavirus (SAV) and infectious ogens in fish. For example, a biosensor system was developed to
salmon anaemia virus (ISAV), which cause significant economic losses detect cortisol in fish plasma samples as a stress indicator in
for farmers in aquaculture.37,102 Another group of researchers has also Oreochromis niloticus.118 The biosensor showed similar results to the
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
842 BOHARA ET AL.

F I G U R E 3 General working principle of biosensors used in aquaculture. Analytes diffuse in the biosensor surface containing the bio
component and react specifically. This change in the properties of the surface led to a change in optical/electronic signal and captured
subsequently.

Enzyme-linked immunosorbent assay (ELISA), indicating its potential 3.3 | Clustered regularly interspaced short
as an alternative to ELISA. In addition to that, different types of sen- palindromic repeats-based (CRISPR) diagnostic
sors are being developed and used to determine the stress level in method
fish.119,120 Biosensors combined with nanoparticles can also be
used as a rapid test kit for the presence of pathogens. Gold nano- Nucleic acid-based diagnosis is a global disease diagnostic method
particle biosensors have detected fish necrosis virus RNA with a relying on PCR and sequencing as standards. These methods have limi-
sensitivity of 270 picograms (pg),64 and graphene oxide-based sen- tations like the need for sophisticated equipment, non-specific amplifi-
sors have been used to detect Vibrio alginolytics up to 31 colony- cation and costly reagents.124 Rapid diagnostics with deployable
117
forming units (CFU/ml). DNAzyme (D-AS-2) combined with technology are needed. CRISPR is an emerging technology for rapid dis-
graphene can detect bacterial disease caused by Aeromonas salmo- ease detection and various applications.125 This system identifies and
nicida, producing fluorescence within 2 min.116 Some viral dis- eliminates foreign sequences using associated endonucleases based on
eases, such as viral hemorrhagic septicemia (VHSV), cause similarity.126 Various CRISPR-endonucleases are identified from bacte-
significant losses in fish farming. Traditional diagnosis using cell ria and archaea,125 targeting foreign DNA or RNA bases in the CRISPR
36,121
culture and RT-PCR can take up to a month. However, gold RNA sequence. The RNA can be modified as needed to eliminate the
biosensors have been developed to target specific G-protein of target.127 CRISPR is extensively used in agriculture128–130 and human
VHSV, providing a more rapid and sensitive (0.0016 μM detection disease diagnostics.131,132 In aquaculture, CRISPR-Cas systems like
limit) detection method. Biosensors are also used to detect food- cas12a (Cpf1) and Cas13 are extensively used for diagnostic purposes
borne pathogens in fish,122 but there are still limitations to their due to their unique characteristics (Table 1).
reliability and applicability in aquaculture. Biosensors can not only Cas13-based diagnostics, known as Specific High Sensitivity Enzy-
detect pathogenic bacteria but this technology can also be used to matic Reporter unLOCKing (SHERLOCK), initially detected the White
monitor stress as well as spawning prediction of aquatic animals. spot syndrome virus (WSSV) with sensitivity up to 2 femtograms of
Additionally, biosensors have the potential to be used in fish DNA.133 SHERLOCK has been used for various aquaculture pathogens
health, fish physiology and behavioural monitoring research. How- since then.134,135 Although SHERLOCK targets both DNA and RNA, its
ever, operating cost for biosensors can be higher if used with label- popularity in aquaculture is limited due to its qualitative nature.136 None-
ling compounds like nanoparticles. Several low-cost techniques are theless, SHERLOCK shows potential as a field-deployable test kit for dis-
being used such as the use of polystyrene and 3D moulding tech- ease detection in remote areas with limited access to laboratory facilities.
niques to reduce the manufacturing cost. A rapid test was devel- The DNA endonuclease-targeted CRISPR trans reporter
oped for detecting Escherichia coli using 3D printed biosensor (DETECTR) system incorporates pre-amplification stages like recombi-
costing about $2.5 USD per sample. Biosensors have the potential nase polymerase amplification (RPA) or rolling circle amplification
to empower farmers by allowing them to swiftly detect pathogens (RAA) to amplify the target sequences before detection (Figure 4),
through uncomplicated test strips. This accessibility can extend to ensuring accurate and sensitive on-field diagnosis at a constant tem-
remote and developing areas across the globe. The FDA has perature (37–42
C). RPA for target sequence pre-amplification offers
approved numerous biosensors for the detection of various forms a time-saving alternative to regular PCR or qPCR.137 Integrating RPA
123
of cancer in humans. However, the process of obtaining into the diagnostic assay for white spot syndrome virus (WSSV) signif-
approvals for biosensors in disease diagnosis, beyond cancer, is icantly increased t detection by almost five million-fold.138 Some
still a complex and lengthy journey. Still in development phase, researchers explored pre-amplification-free CRISPR-Cas systems for
with the enhancement of sensitivity and specificity biosensors disease diagnosis to minimise pre-amplification equipment,135,139 lim-
have potential in animal health monitoring. ited to high target amounts.
 BOHARA ET AL.

TABLE 1 Characteristics of different CRISPR-Cas systems reported for aquatic pathogen diagnostic.

Pre-
SN Name Enzyme amplification Assay time Readout Sample prep Application LOD References
1 CRISPR Cas12a RPA <1 h Fluorescence Crude extraction White spot syndrome virus 200 copies 138
(WSSV)
2 CRISPR-based SHERLOCK Cas13a RPA 1h Fluorescence and Lateral- Crude extraction White spot syndrome virus 530,000 copies 133
flow strips (WSSV)
3 CRISPR-Cas12a fluorescence Cas12a RPA 1h Fluorescence Crude extraction Enterocytozoon 50 copies of DNA 141
assay hepatopenaei (EHP)
4 RAA-CRISPR/Cas12a Cas12a RAA 40 m Fluorescence NS Vibrio vulnificus 2 copies of DNA 137
5 CRISPR/Cas13a Cas13a NS 20 m Fluorescence NS Red-spotted grouper 100 fM 134
nervous necrosis virus
(RGNNV)
6 RPA-CRISPR/Cas12a assay Cas12a RPA 2h Fluorescence and Lateral NS Acute hepatopancreatic 20 copies/μL for fluroscence, 142
flow necrosis disease (AHPND) 200 copies/μL (0.3 fmol/L)
for lateral flow
7 RPA-CRISPR/Cas12a Cas12a RPA 1h Fluorescence and Lateral Crude extraction Scale drop disease virus 40 copies per reaction 143
flow (SDDV)
8 Dualplex dRAA-CRISPR/ Cas12a dRAA 45 m Fluorescence Crude extraction Aeromonas hydrophila 2 copies of DNA 144
Cas12a
9 RT-RPA-CRISPR-Cas12a Cas12a RT-RPA 1h Smartphone fluorescence, Tilapia lake virus (TiLV) 200 copies of RNA 38
lateral-flow strip
10 Signal-off Cas14a1-based Cas14a1 PCR 1h Fluorescence NS Staphylococcus aureus 10,000 CFU/mL 145
platform (SOCP)
11 CRISPR-Cas13a Cas13a Amplification 15 m Fluorescence NS Taura syndrome virus 1,000,000 copies (1.66 pM) 135
free
12 RPA-CRISPR/Cas12a Cas12a RPA 45 m Fluorescence Crude extraction Acute hepatopancreatic 100 copies/reaction 140
necrosis disease (AHPND)
13 LAMP-CRISPR-Cas12b Cas12b LAMP 30 m Fluorescence Crude extraction Taura syndrome virus (TSV) 100 copies for WSSV and 146
(SHERLOCKv2) and White spot syndrome 200 copies for TSV
virus (WSSV)
16 RR-Cas Cas 12a RT-RPA 1.5 h Fluorescence/lateral flow Crude extraction Yellow head virus genotype 100 copies plasmid and 147
1 100 fg total RNA

Abbreviations: CFU, colony-forming unit; fg, femtogram; fM, femtomolar; NS, not specified; pM, picomolar; RPA, rapid polymerase amplification; RR or RT-RPA, reverse transcription recombinase polymerase
amplification.
843

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 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
844 BOHARA ET AL.

F I G U R E 4 Two main CRISPR-Cas systems used in aquatic animal disease diagnostics. SHERLOCK targets the RNA, while DETECTR can
target both DNA and RNA.

Wang et al. (2023) developed a one-pot CRISPR-Cas12a assay, WOAH and USDA-APHIS is essential for CRISPR tools in aquatic dis-
integrating RPA and CRISPR/Cas12a in a single tube for acute hepato- ease diagnostics, potentially revolutionising disease surveillance.
pancreatic necrosis disease (AHPND) detection with sensitivity of up CRISPR-based disease diagnosis is a relatively new tool developed by
to 100 copies per reaction. Cas12a's optimal cleavage temperature is researchers for diagnosing diseases. Further improvements are needed
37
C, compatible with RPA (37–42
C).140 Remote conditions can uti- to enhance its analytical sensitivity and specificity to obtain regulatory
lise human body temperature for pre-amplification and cleavage. approvals. The limitations include the need for trained professionals,
RPA-Cas12a detection was adapted for lateral flow using FAM- time-consuming reaction preparation and the applicability only to
ssDNA-Biotin and FAM-antibody with gold nanoparticles, more feasi- known sequences. CRISPR diagnostics aims to be field-deployable in
ble than a fluorescence-based system.38,141 aquaculture, yet specific temperature requirements and lengthy sample
CRISPR-Cas-based diagnostics are gaining attention for their high preparation hinder progress. Despite challenges, CRISPR has the poten-
sensitivity in detecting low copy numbers and user-friendliness. While tial to transform conventional diagnostic methods.
the cost per sample for CRISPR diagnostics in aquaculture has not
been studied extensively, SHERLOCK-based COVID-19 test kits cost
around $15–$30 per sample.148,149 Successful implementation of a 3.4 | Matrix-assisted laser desorption ionisation-
diagnostic technique requires sensitivity, reliability, affordability and time of flight mass spectrometry (MALDI-TOF MS)
portability. Cas12a targets double-stranded DNA, necessitating
reverse transcription for RNA-based targets, posing an additional limi- MALDI-TOF MS is a method utilising mass spectrometry to diagnose
tation. Despite SHERLOCK's ability to target both DNA and RNA, its microbial organisms. This method is rapid and economical, making it a
popularity in aquaculture is restricted due to its qualitative nature potential candidate for use in aquaculture diagnostics. In mass spec-
rather than quantitative.136 Approval from organisations like the trometry, the compounds are ionised into charged molecules, which
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
BOHARA ET AL. 845

can be measured using mass spectrometer.150 In MALDI-TOF mass including mucus, cysts and spores. Filters are used to collect these
spectrometry, the target samples are mixed with a matrix and passed genetic materials, which are then purified to determine their pres-
through high voltage to separate the ions based on their mass- ence.179 In fisheries, eDNA is widely employed to identify different
to-charge ratio, followed by detection using a time-of-flight (TOF) fish species in water bodies without harming the fish.180 The collected
detector. Peptide mass fingerprinting (PMF) is utilised to identify the DNA and RNA are subjected to PCR or qPCR tests to detect patho-
mass of the unknown protein by comparing it with the databases. gens. This non-invasive technique is helpful for disease diagnostics
151,152
MALDI-TOF MS is used for clinical bacteriology, detecting anti- and surveillance in aquaculture.181 On-farm PCR testing can be con-
153 154
biotic resistance bacteria and identifying different strains. ducted using environmental samples, providing effective surveil-
In aquaculture, this technology has been studied in different lance.182 Researchers have demonstrated the use of eDNA and eRNA
viruses,155,156 bacteria,157–162 fungus163 and parasite.164 Moussa in biosecurity measures during ornamental fish trade without the need
et al. (2021) constructed a database using different spectral projec- to sample the fish directly.183 This technology has also been utilised
tions in MALDI-TOF MS to rapidly identify 25 different species of Vibrio for eDNA metabarcoding, allowing the simultaneous detection of mul-
in molluscs.165 MALDI-TOF MS is mostly used to characterise and differ- tiple pathogen species.184 In aquaculture, indoor facilities in biomedi-
entiate species in aquaculture. However, some studies also focus on cal research laboratories face challenges with fungal diseases like
164
using these technologies in diagnosis. Cell culture is considered a gold Pseudoloma neurophilia, which require timely identification and con-
standard for viral disease diagnosis in aquaculture, followed by PCR. trol.185 The efficiency of eDNA in detecting marine pathogens that
166
MALDI-TOF MS is more suitable for bacterial diseases than viral dis- threaten wild and domestic salmon populations has been observed,
ease detection because of the low protein content and higher molecular with salmon farms identified as potential reservoirs for these patho-
167
weight of viral protein. In addition, the virus cultured in cells also con- gens.176,186 eDNA has also shown potential as a surveillance method
tains a high amount of proteins from the cells themselves, making the for aquatic viruses such as Herpesvirus and Ranaviruses.187 Addition-
168
peptide mass fingerprinting complex. Yet, there are some studies in ally, eRNA has been reported as a non-invasive technique for identify-
human viral diseases diagnostic such as human herpesvirus,169 enteric ing specific pathogens like the Tilapia Lake virus and protozoan
170 171
viruses and human papillomavirus using MALDI-TOF MS. A study parasites.188,189
conducted to compare the cost and performance of MALDI-TOF MS eDNA and eRNA-based diagnosis offer several advantages,
showed the minimal price ($0.5 per sample) with 84%–96% accuracy of including non-lethal sampling, no handling stress, cost-effectiveness,
result in identification of yeast.172 time efficiency and the ability to detect multiple pathogens and their
The advantage of MALDI-TOF MS technology is its ability to ana- hosts using a single sample. EDNA sample analysis costs approxi-
lyse samples within minutes of culturing with acceptable accuracy. Addi- mately $54 when processing 24 samples in a single run. However,
tionally, the cost per sample ranges from $1 to $3.14, excluding the when running 100 samples in a single run, the cost per sample
instrument cost.173 Furthermore, this method can be integrated into cur- decreases significantly to approximately $39.190 In 2022, WOAH
rent microbiology culture conditions and workflows. However, one major released a discussion paper on using eDNA methods in aquatic animal
limitation of this technology is the lack of precise identification of viral diseases, stating the benefits and limitations of this method in aqua-
and fungal species. Additionally, accurate identification relies on estab- culture.191 The limitation of these methods is the degradation of
lished databases for pathogen identification, which are incomplete for nucleic acids, with DNA being more stable and easier to detect than
aquatic pathogens. This makes it more challenging to adopt this technol- RNA.192 Additionally, detecting pathogens directly can be challenging
ogy for aquatic pathogen detection. Moreover, this method cannot be due to their microscopic size and different life stages in water. More-
directly applied to tissue samples, limiting its use to laboratory-based over, clinical signs of disease may only become evident in advanced
diagnosis only, making it inaccessible to farmers in rural areas with limited stages of infection, limiting treatment options. Nevertheless, eDNA
access to laboratories. However, in 2022, bioMérieux, Inc. (https://www. methodologies can potentially enhance animal health monitoring sys-
biomerieux-usa.com/) developed a laboratory instrument based on tems in aquaculture by addressing these challenges.193
®
MALDI-TOF MS, the VITEK MS PRIME, which received FDA approval The field of aquatic pathogen diagnosis has undergone continu-
as a microbial organism identification tool.174 The VITEK® MS PRIME ous evolution, transitioning from visual assessments to molecular
instrument can identify various bacterial pathogens, including those pre- techniques. Nevertheless, in aquaculture, disease diagnosis continues
sent in aquatic environments. However, the initial cost of the VITEK sys- to adhere to the three distinct levels (Levels I, II and III) of diagnosis as
tem falls within the range of $50,000–$120,000, making it a relatively established by the FAO.194 At Level I, diagnosis is primarily based on
expensive instrument, which does not include supplies. preliminary observations of the animals or their environment. In con-
trast, Level II and Level III employ advanced laboratory techniques for
definitive confirmation, as outlined in the FAO's technical manual.
3.5 | Environmental DNA/RNA (eDNA/eRNA) Each of the confirmatory diagnostic methods mentioned above has
advantages and drawbacks, with considerations including specificity,
eDNA and eRNA are innovative techniques for detecting DNA or sensitivity, time requirements and cost-effectiveness (Table 2).
RNA from environmental samples such as air, water and soil.175–178 Nanopore sequencing has the advantage of identifying novel
This method captures nucleic acids released externally by organisms, pathogens, but it lacks accuracy as compared to CRISPR-based
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
846 BOHARA ET AL.

T A B L E 2 Comparison of emerging diagnostic techniques with the current aquatic disease diagnostic methods for sensitivity, specificity,
labour, cost and time.

Diagnostic methods Sensitivity Specificity Cost Speed Labour

Nanopore Sequencing High High Low High Medium
Biosensors High High High initial cost Fast Low
CRISPR High High Low Fast Low
MALDI-TOF-MS High High High initial cost Medium Low
eDNA/eRNA High High Low Slow High
PCR/qPCR High High Medium Slow High

diagnostics. MALDI-TOF MS can detect a wide range of pathogens, ROVs, sensors and IoT are undergoing extensive diagnostic and sur-
including parasites, bacteria, fungi and viruses, but its accuracy veillance enhancement research. Oxford's nanopore device (https://
depends on established databases for all the pathogens under exami- nanoporetech.com/) offers real-time DNA/RNA sequencing for about
nation. eDNA/eRNA is a non-invasive method for disease diagnosis; $1000. Environmental DNA/RNA's potential for early diagnostics
however, the samples used (water) can harbour different pathogens, faces challenges due to nucleic acid degradation in aquatic environ-
making the results unreliable. CRISPR-based disease diagnostic is in ments.181 CRISPR-based diagnostics, multiplex-PCR, loop-mediated
the initial testing phase in aquaculture. Still, it has several advan- isothermal amplification (LAMP), DNA microarray and serology are
tages over other tools, such as being less time-consuming, specific explored as rapid, sensitive tools for aquatic disease identification.197
to the tested pathogens and sensitive to low pathogen load. How- Nevertheless, these tools are primarily designed to detect established
ever, this tool is only tested on specific pathogens in the fish and pathogens in aquaculture. When it comes to identifying novel patho-
shrimp industry and has not yet been tested for fungal and parasitic gens, CRISPR, MALDI-TOF-MS and biosensors may be restricted by
diseases. The regulatory approval of any diagnostic assays has its their inherent specificity towards known pathogens. In contrast, nano-
criteria as (1) definition of intended purpose, (2) optimisation, pore sequencing and environmental DNA (eDNA) analysis can identify
(3) standardisation, (4) repeatability, (5) analytical sensitivity, (6) ana- such pathogens through NGS techniques. Commercialisation poses a
lytical specificity, (7) thresholds, (8) diagnostic sensitivity, (9) diagnos- challenge for these technologies. While aquaculture researchers
tic specificity, (10) reproducibility and (11) fitness of intended develop diagnostic kits, industry partnerships are crucial for availabil-
purpose.36 Research on refining these tools is underway to achieve ity to farmers. In contrast, CRISPR-based diagnostic kits are actively
the standards set by the regulatory bodies.33,108 developed for human and animal diseases by more than 30 companies,
outpacing aquaculture.198 Lateral-flow strips utilising biosensors and
CRISPR offer easy pathogen detection in remote areas. LAMP
4 | DISCUSSION and RPA integrated with or without CRISPR146 are field-deployable
for rapid identification. Several kits using LAMP colorimetric detection
While the aquaculture industry is embracing innovation, it lags behind are developed to detect Flavobacterium columnare (now Flavobacter-
agriculture.13 In contrast, human medicine has made remarkable ium covae) in Nile tilapia,199 and Shewanella putrefaceins200 and kidney
strides, exemplified by rapid COVID-19 diagnostic and vaccine devel- necrosis virus201 in farmed tilapia and Tilapia Lake virus.202,203 Due to
opment.195 Aquaculture struggles with less efficient diagnostic tools the temperature requirement, non-specific amplification and the pos-
36
for viral diseases, often taking days to months for diagnosis. Yet sibility of primer-dimer formation, LAMP and RPA can be more suit-
insights from human and animal disease diagnostics can aid efficient able for increasing the efficiency of CRISPR-based diagnostics.
methods for aquatic diseases. Currently, PCR and cell culture are Collaboration with companies is vital for kit development and aqua-
WOAH's standard diagnostic methods due to high sensitivity and culture commercialisation. Validity must be supported by research and
36
specificity, but they are time-consuming and costly. Additionally, demonstrations. Furthermore, the commercialisation of diagnostic kits
current water quality monitoring and farm monitoring practices are necessitates approval from regulatory bodies such as the World Orga-
labour-intensive and time-consuming, making it a challenge for nisation for Animal Health (WOAH). WOAH has strict guidelines for
countries like the United States, where farm labour shortage is a big validating and certifying diagnostic assays with high standards.204
196
issue. Once these kits are approved for diagnostic testing, they are deemed
Advancements in diagnostics across various sectors have valid for use in detecting and monitoring aquatic diseases. Ensuring
prompted the development of novel methods in aquaculture. How- the accessibility of developed technologies in developing countries
ever, diagnostic methods need to be specific and sensitive, along with poses a notable challenge. Nevertheless, ongoing initiatives led by
their cost-effectiveness. Any false positive or false negative might be organisations like the Food and Agriculture Organization (FAO) and
disastrous for farmers, the region and the country. Nanopore the United States Department of Agriculture (USDA) are actively facil-
sequencing, CRISPR diagnostics, biosensors, MALDI-TOF-MS, eDNA, itating the transfer of technology from developed nations to
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
BOHARA ET AL. 847

developing. These efforts are supported through a combination of finan- from such technologies. Nevertheless, large-scale producers with suf-
cial assistance and the sharing of technological resources aids.205,206 Fur- ficient resources and personnel could reap the long-term benefits of
thermore, the Feed the Future Innovation Lab for Fish, funded by USAID, these technologies. Furthermore, the development of low-cost rapid
and the Food and Agriculture Organization (FAO) are collaborating to diagnostic kits with higher sensitivity and specificity for identifying
support aquaculture farmers through capacity-building programmes and pathogens in aquaculture can be utilised worldwide without the need
initiatives focused on sharing technological advancements.207 for sophisticated laboratories and expensive instruments. Fostering
Efficient farm monitoring is a key for the future aquaculture partnerships to facilitate technology transfer and provide essential
industry, which is growing yearly to feed the increasing population. training will guarantee the widespread availability of these technolo-
Intelligent aquaculture farming utilising new technologies to detect gies. This will ensure accessibility to these methods in developing and
the problem promptly will be a massive factor for sustainable produc- developed countries, helping address food security issues. Identifying
tion. Farming practices, including the use of sensors to monitor water diseases on time in aquaculture is critical for better production by pre-
parameters and drones to detect abnormalities, diseases and feeding venting animal loss. However, a major constraint is the lack of rapid
behaviour, can be integrated into an AI system through the internet, identification kits for aquatic animal diseases. Efforts are underway to
creating an autonomous management system for the farm. Sensors develop these kits for use in aquaculture but are mostly limited to lab-
and drones aid monitoring in aquaculture, although challenges include oratory settings. Some researchers have developed field-deployable
availability, regulations, cost, reliability, water quality, electricity, inter- colorimetric kits for diagnosis using CRISPR, biosensors and LAMP;
net access and farmer perception.13 Regulations using drones vary however, they lack sensitivity and specificity. The use of new sensors
according to countries, such as in Canada the drones weighing more and precision aquaculture techniques is underway for easy farm man-
than 250 g must be registered while in Germany this requirement is agement and decision-making. These techniques have improved
only for a drone weighing more than 5 kg. Another requirement for aquatic health management to reduce disease incidence on farms.
drones is to fly only on the visual line of sight; however, some farms WOAH needs to research for including these technologies in aquatic
might be big, necessitating flying from different points, making it disease diagnostics to reduce the efforts and time in current methods.
208
hard. In contrast, underwater drones lack clear and comprehensive Affordable, accessible and easy-to-use surveillance and monitor-
regulatory frameworks, posing significant challenges for environmen- ing technologies are necessary to make them easy to adopt by farmers
talists concerned about their potential risks to ocean ecosystems, par- around the world. In the future, more research efforts are needed to
ticularly regarding collisions with marine organisms. AI and ML hold optimise the diagnostic protocol to make them easy to use by farmers
potential but require suitable data analysis methods, reference data- with little to no effort. In addition, research is needed for CRISPR-
bases and cost considerations in aquaculture. Nonetheless, drawbacks based field deployable kits for specificity, scalability and sensitivity by
like inadequate data analysis methods, higher initial costs and limited partnering with industry and regulatory bodies. Drones have become
reference databases hinder AI adoption. Visual AI for aquatic disease invaluable tools in aquaculture, serving various purposes beyond farm
diagnosis holds promise but requires robust training databases. Aqua- monitoring. Their versatility extends to tasks such as assessing and
culture's lower priority and support in research compared to human quantifying algal blooms, diagnosing diseases in aquatic populations,
and animal medicine pose challenges. While diagnostic, surveillance gathering sensor data and facilitating water sampling. Research on
and monitoring technologies offer advantages and disadvantages, fur- specific use cases of drones, AI and ML is needed to better under-
ther research on commercial applications with cost–benefit analysis is stand their potential applications in aquaculture. To harness the
essential. Feasibility studies for developing countries are needed. potential of AI and ML in disease identification within aquaculture, it
is necessary to gather diverse datasets encompassing various aquatic
diseases. Future research endeavours can be directed towards estab-
5 | CONCLUSION AND FUTURE REMARKS lishing a comprehensive image repository accessible to all. This reposi-
tory can serve as a valuable resource for fish health experts, enabling
Global population growth necessitates sustainable and efficient agri- them to utilise these images for training AI and ML models, thus
culture production systems, including aquaculture. The aquaculture enhancing disease detection capabilities. Sensor datasets are also
industry is evolving with the adoption of emerging technologies such valuable for AI and ML experts to forecast that region's water quality.
as recirculatory aquaculture system (RAS), off-shore aquaculture, cage Integrating different types of sensor data with AI and ML can poten-
culture, digital aquaculture, fish meal alternatives, vaccines, genetic tially reduce the environmental and stress factors in the Snieszko dis-
engineering, drones and artificial intelligence, resulting in more sus- ease triangle. Disease prevention by effective farm monitoring
tainable and profitable production. Current methods in aquatic dis- practices and identification of diseases on time are crucial aspects of
ease diagnostics need to be revised due to their ineffectiveness in sustainable aquaculture production. Research and innovation in tech-
rapidly diagnosing pathogens. Farm monitoring technologies are nological fields are vital for the growth of the aquaculture industry.
labour-intensive and costly, necessitating advanced technology adop-
tion. However, integrating advanced technologies in developing coun- AUTHOR CONTRIBU TIONS
tries is challenging due to high costs, complicated infrastructure and Kailash Bohara: Conceptualization; writing – original draft; writing –
limited internet access, making it difficult for small farmers to benefit review and editing. Pabitra Joshi: Writing – original draft; writing –
 17535131, 2024, 2, Downloaded from https://onlinelibrary.wiley.com/doi/10.1111/raq.12870 by BINGOL UNIVERSITY, Wiley Online Library on [09/05/2025]. See the Terms and Conditions (https://onlinelibrary.wiley.com/terms-and-conditions) on Wiley Online Library for rules of use; OA articles are governed by the applicable Creative Commons License
848 BOHARA ET AL.

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