Diseases of the giant river prawn Macrobrachium rosenbergii : A review for a growing industry

The giant river prawn, Macrobrachium rosenbergii , is a major focus of aquaculture in tropical and sub-tropical regions around the globe. Over the last 30 years, culture of M . rosenbergii has increased exponentially as demand has risen both for domestic consumption and for international export trade. As with many aquaculture species increases in production have been accompanied by the emergence of diseases affecting yield, profit and trading potential. Disease-causing agents include pathogens infecting other crustaceans, such as Decapod Iridescent Virus (DIV1), as well as pathogens only known from M . rosenbergii such as White Tail Disease caused by Macrobra-chium rosenbergii nodavirus (MrNV) and extra small virus (XSV). Here, we review the pathogenic agents associated with the culture of M . rosenbergii since commercial culture began in earnest during the 1970s. Particular emphasis is given to pathogens first identified in other aquaculture host species, but which have subsequently been shown to infect and cause disease in M . rosenbergii . As polyculture of M . rosenbergii with other aquaculture species is common practice, including culture with other decapods, crabs and fish, increased pathogen transfer among these farmed species may occur as M . rosenbergii aquaculture increases in the future.


| INTRODUCTION
Aquaculture is the fastest-growing farmed food sector, with the proportion of cultured to caught seafood increasing year-on-year. 1,2obal aquaculture production for fish, crustaceans and molluscs surpassed 87 million tonnes in 2020, just under half of all world production. 2 Crustacean production by aquaculture far surpasses that by capture, with more than 11,000,000 tonnes in 2020 from aquaculture compared to 6,000,000 tonnes by capture. 2 Despite these considerable production figures, it is estimated that up to 40% of tropical shrimp aquaculture production is lost annually, 3 equating to losses of over 3 billion USD, and this is primarily due to infection with viral agents. 4Losses on this scale threaten global food security, with most of the consumption of cultured crustaceans being outside of producing countries, making management and mitigation of disease of key importance to the future success of crustacean aquaculture. 4Stentiford et al. (2012) outlines a holistic strategy involving both producer and consumer nations to improve husbandry and farm management, better understand pathogens and their spread (either by movement of animals or alternative hosts) and learn lessons from previous disease outbreaks.This review focuses on these points to outline the pathogenic agents in Macrobrachium rosenbergii culture, identify other hosts that these agents are able to infect, and discuss how culture practices can mitigate disease transfer.
The freshwater prawn genus Macrobrachium comprises over 200 species, distributed throughout the world's tropical and subtropical regions. 5The giant river prawn, M. rosenbergii, is indigenous to the Indo-Pacific area, 6 but has been transferred from its native locations to almost every continent for farming purposes following the development of methods to mass-produce postlarvae. 5M. rosenbergii is primarily found in inland freshwater areas, including but not limited to rivers, swamps and canals, particularly where turbidity makes the water cloudy or opaque.Most species of Macrobrachium, including M. rosenbergii, require brackish water in the larval stages of development before moving to freshwater as postlarvae, and thus are frequently found in habitats that are connected to the sea. 5 M. rosenbergii is cultured worldwide in tropical and sub-tropical climate regions. 7Its culture provides significant income, as well as both direct and indirect employment and a food source in areas of poverty. 7The first countries to report production of M. rosenbergii to FAO of the UN (Food and Agriculture Organisation of the United Nations) were Thailand and Vietnam in 1975, followed over the next two decades by Myanmar, Taiwan, India, Bangladesh and China. 2 Prior to 1980, global production was below 3000 metric tonnes per year 2 ; since then, the species has become a desirable target for commercial aquaculture due to its large size, tolerance to some diseases that cause mass mortality in shrimp aquaculture, and considerably higher market price than the same weight of marine penaeid shrimp. 7,8In 2020, worldwide production of M. rosenbergii reached 294,081 tonnes ($2.4bn USD), with over 50% of global production in China. 2 The expansion of M. rosenbergii culture has resulted in numerous technological advancements and investments to facilitate the sector.
These include the production of monosex cultures of M. rosenbergii by RNA interference (RNAi) or injection of suspended hypertrophied androgenic gland cells in order to reduce aggression (reducing mortality by injury, decreasing losses), increase yield and to make harvest size more uniform 9,10 ; the construction of intensive hatcheries 11 and, the production of specific-pathogen-free (SPF) stocks. 12However, despite these significant advances, increased incidence of disease has accompanied expansion of the industry, with the emergence of novel pathogens that current SPF stock production programmes do not consider.
4][15][16] One of the reasons for deploying polyculture of M. rosenbergii with other aquaculture species is to mitigate losses from disease in one species by maintaining yield and profit from the others. 17Another potential advantage derives from the hypothesis that the effect/impact of pathogens of one cultured species can be modified/quenched by the presence of (an)other(s). 18sh-prawn polyculture systems are thought to reduce the frequency of algal blooms commonly observed in finfish monoculture. 194][15][16] Prawn-shrimp polyculture systems are less common but have been reported in China and Bangladesh with the former culturing M. rosenbergii with the shrimp Penaeus (Litopenaeus) vannamei, and the latter farming M. rosenbergii with P. monodon. 13,20 China, prawn-prawn polyculture with M. rosenbergii and Macrobrachium nipponense, the oriental freshwater prawn, also occurs. 13China is the world's largest producer of the Chinese mitten crab, Eriocheir sinensis 2 ; in 2012, 90% of all M. nipponense polyculture was with E. sinensis, with over 300,000 ha of land dedicated to this type of culture. 13Reports suggest that E. sinensis-M.rosenbergii polyculture as well as M. rosenbergii-Anodonta woodiana (Chinese pond mussel) polyculture also occurs in China. 21,22tection of known and emerging pathogens has become more accessible through advances in molecular biology techniques that have allowed the rapid characterisation of novel pathogens without relying on traditional pathogen culturing techniques. 23Numerous disease-causing agents are known to infect M. rosenbergii, including several bacterial, viral, fungal and other eukaryotic pathogens, causing losses to the aquaculture sector either by mortality, or by the production of a smaller or substandard animal with decreased market value. 24en new aquaculture practices emerge concurrently with a concerted effort to improve the productivity of freshwater prawn farming, similar to marine shrimp farm operations, the risk of novel prawn diseases also increases (Figure 1).Since the last review of disease in M. rosenbergii in 2012, 24 several agents have been newly identified as pathogenic in giant river prawn culture, mainly aided by enhancement in molecular detection.In this review, we describe pathogenic agents of disease that have impacted M. rosenbergii since the origins of commercial culture of this species; further, we discuss the potential host range of these pathogens and their possible impact on the health of M. rosenbergii and those species with which it is often co-cultured.

| VIRUSES
As the culture of freshwater prawns (Genus: Macrobrachium) and penaeid shrimp (Family: Penaeidae) has increased, the incidence of viral infections has also increased.It has also become apparent that viral host specificity is not bound to fresh-or saline water, with several viruses of penaeid shrimps able to infect freshwater prawn species and vice versa.A summary of viruses known to infect M. rosenbergii is provided in Table 1.
Hepatopancreatic parvovirus (HPV) was the first reported virus of M. rosenbergii, 25 and has since been reclassified in the genus Aquambidensoviridae. 26 The 25-30 nm icosahedral virus, discovered in giant river prawns in Malaysia, was not associated with mortalities and primarily infected the hepatopancreatic tissue of postlarvae, with basophilic intranuclear inclusions present in tubule epithelial cells.It was first thought that this parvo-like virus was the same as another parvovirus infecting the Korean Penaeus (Fenneropenaeus) chinensis, a species of marine shrimp.However, differences in virus size and infected host cell histopathology justified its designation as separate species. 27nce its discovery, there have been few reports of HPV in M, rosenbergii, most likely due to the lack of mortality associated with infection; however, HPV is associated with mortalities in penaeid shrimp. 28e only reports of HPV infection of M. rosenbergii since the distinction between the two parvoviruses have been in postlarvae from Thailand and Malaysia in 2007 and 2009, respectively. 29,30A more recent study attempted to detect HPV in M. rosenbergii collected from wild populations in India, but could not amplify the virus by PCR. 31 Another parvovirus, Penaeus stylirostris penstyldensovirus 1 (PstDV1), 26 commonly named infectious hypodermal haematopoetic necrosis virus (IHHNV), a 20-22 nm icosahedral-shaped virus, has been shown to cause disease in penaeid shrimp. 32Gross clinical signs of infections vary with species, from runt deformity syndrome in P. vannamei and P. monodon to a whitish colour and opaque abdominal musculature resulting in mortality of Penaeus stylirostris postlarvae and adults. 32In 2004, a pond in Taiwan reported mortality of M. rosenbergii postlarvae and juveniles. 33The mortality was linked to the slow growth of animals, later determined to be infected with IHHNV, which manifested as muscular atrophy, a reddish colouration, and deformities to the cuticle and opaque musculature.Histopathological examination identified Cowdry type A and B eosinophilic intranuclear inclusions in hepatopancreatic epithelial cells, a different presentation to the basophilic inclusion seen in infection with HPV.
By far the biggest threat to the viability of M. rosenbergii aquaculture worldwide has been the emergence of a novel nodavirus, Macrobrachium rosenbergii nodavirus (MrNV) (Family: Nodaviridae) and an associated satellite virus, extra small virus (XSV), infecting M. rosenbergii and causing white tail disease (WTD). 34The impact this nodavirus has had on the culture of M. rosenbergii has resulted in a large number of research publications, from pathogenicity of the virus to host response.The first report of MrNV was in hatchery-reared M. rosenbergii postlarvae in the French West Indies in 1994. 35However, MrNV is likely synonymous with Macrobrachium muscle virus (MMV), a virus of the same size that caused similar clinical signs of disease and post-larval mortalities since 1992 in Taiwan. 36The clinical signs of WTD are whitish colouration of the abdominal and tail muscle, with discolouration spreading from the tail towards the head as the infection progresses. 35Histopathological signs of WTD include hyaline necrosis of muscle fibres, with moderate oedema, necrosis, haemocyte infiltration and fibrosis in affected muscles; with the presence of pale to dark basophilic intracytoplasmic inclusions in muscle cells and hepatopancreatic connective tissue cells. 35MrNV is a 26-27 nm nonenveloped, icosahedral virus with a genome composed of two   White triangle under the carapace at the bottom of the rostrum, yellow gills, loss of swimming ability, migration to deep water, mortality. 79sinophilic inclusions and karyopyknosis in haemopoietic tissue. 79Gill-associated virus Roniviridae 52 Unknown. 53nknown.
b Macrobrachium rosenbergii nodavirus (MrNV) 35 Nodaviridae 37 recent years, M. rosenbergii culture in China has been facing problems associated with growth retardation of prawns known as iron prawn syndrome (IPS)-prawns have body weights and lengths much smaller than normal. 43Several studies have been conducted to determine a cause for this phenomenon, including environmental factors and known pathogens [44][45][46] ; however, no one factor has been identified as the primary cause.The syndromic nature of this disease is discussed further in the Section 6.When IPS prawns are cultured with unaffected prawns, the latter begin to exhibit reduced growth, suggesting that the cause may be of pathogen aetiology. 47One potential cause of IPS is a novel hepe-like virus identified in M. rosenbergii with slow growth from a farm in China. 48Crustacea hepe-like virus 1 (CHEV1), identified via metatranscriptomics, has a 7750 nt positive sense ssRNA genome with three hypothetical open reading frames.
Although CHEV1 could not be identified as the primary cause of ironprawn by Dong et al. (2020), it highlights the importance of emerging pathogens and syndromic disease conditions in aquaculture.
Further to the discovery of CHEV1, another virus, infectious precocity virus (IPV) or Crustaflavivirus infeprecoquis, has been linked to IPS in M. rosenbergii. 49The 12,630 nt single-stranded RNA virus is proposed to be of the family Flaviviridae, and was assembled from metatranscriptomic sequencing of IPS prawns.A disease challenge using a viral extract from IPS prawns was able to reproduce the IPS phenotype, with eosinophilic viral inclusions were seen in multiple tissue types (Table 1).The same study 49  Problems with feeding and moulting, reddish exuviae, decreased response to stimuli, mortality. 42le to dark basophilic cytoplasmic inclusions in the cuticle epithelium. 42nodon-type baculovirus (MBV) 60 AKA Penaeus monodon Nudivirus (PmNV) 58 Baculoviridae 60 234-316 nm 82 Unknown 62 Large nuclei in hepatopancreatic tubule epithelial cells with diffuse, central, eosinophilic inclusions and marginated nucleoli and chromatin. 62ite spot syndrome virus b (WSSV) 67 Nimaviridae 67 250-380 nm 83 White spots on carapace, lethargy, increased cannibalism, mortality.[70][71][72][73] Hypertrophied Feulgen-positive nuclei in target tissues, basophilic and Cowdry A type inclusion bodies, chromatin margination and karyorrhexis.(MrGV)-assembled from metatranscriptomic data, proposed to belong to Nidovirales, was present in hatcheries with mass mortality events. 50ylogenetically, the closest known relative of MrGV is Gillassociated virus (GAV), which comprises genotypes that cause mass mortalities in penaeid shrimp culture. Sincee publication of MrGV, the virus has also been identified in metatranscriptomic data from postlarvae in China, but in the absence of disease.51 The yellow head viruses (YHV) are positive sense ssRNA viruses of the order Roniviridae. 52 Tote, YHV comprises eight genotypes.53 The most notable genotype of YHV is yellow head virus genotype 1 (YHV-1), the only genotype notifiable to the World Organisation for Animal Health (WOAH, previously OIE), which forms enveloped, rodshaped virions and is the causative agent of yellow head disease (YHD) that has resulted in mass mortalities of cultured penaeid shrimp.54 YHV genotype 2 (YHV-2), more commonly known as Gillassociated virus (GAV), is also associated with mortalities caused by gill-associated virus disease.Genotypes three to seven (YHV-3 to YHV-7) commonly occur in healthy P. monodon and are rarely or never associated with disease or mortality.55 However, in recent years, yellow head virus genotype eight (YHV-8) has been detected in shrimp exhibiting acute hepatopancreatic necrosis disease (AHPND) in Chinese F. chinensis, but the pathology of YHV-8 in the absence of other pathogens is not known.56 Longyant et al. ( 2005) screened for YHV-1 in M. rosenbergii collected from or nearby a YHV-affected P. monodon farm in Thailand as well as experimentally infected M. rosenbergii.57 YHV-1 was not detected in either group of M. rosenbergii, suggesting that it may not be a susceptible species.However, Yang et al. (2016)   were able to detect YHV in 21% of cultured M. rosenbergii (n = 19) using a novel LAMP assay designed to target a conserved region of the YHV complex in YHV-1, YHV-2 and YHV-8 genotypes.53 The degrees to which M. rosenbergii is susceptible to different YHV genotypes, and can be a reservoir of YHV, are yet to be determined.
monodon in 1977, 60,61 PmNV has since been shown to have a wide host range of penaeid shrimp and infections have been reported around the globe. 59PmNV has been observed in all life stages of penaeid shrimp, but is most lethal to late larval, post-larval and juvenile shrimp. 59 Covert mortality nodavirus (CMNV), a 32 nm icosahedral nodavirus first identified in P. vannamei, causes viral covert mortality disease (VCMD) in penaeid shrimp. 63,649 A study in 1998 collected giant river prawns from culture farms and experimentallyinfected adults, postlarval and larval M. rosenbergii with WSSV.Again, all WSSV-positive animals from the aquaculture farms were positive only by nested PCR and no mortalities were associated with the infections. 70Experimental infection of M. rosenbergii led to clinical signs of disease, including white spots on the carapace; however, these spots appeared to have a smaller diameter, and a different shape and colour to those of P. monodon infected with the same strain of WSSV. 70Peng et al. (1998) noted that experimentally infected larvae and postlarvae appear to be more susceptible to WSSV infection than adults; this was also reported in a study in 2002 along with increased cannibalism in WSSV-infected groups. 71Interestingly, adult M. rosenbergii experimentally infected with WSSV can clear the virus within a few days of infection 72 ; infected individuals exhibit lethargy and anorexia within the first few days of infection but recover within 24 h. 72,73This ability to clear the virus is also seen from a molecular perspective: VP28 WSSV enveloping protein mRNA can only be detected up to 4 days post-infection, whereas WSSV DNA can be detected 40 days post- Reported life stage(s) affected Clinical signs of infection
Bacillus sp. 89uveniles (GO, L), adults (GO) "Black spot disease"-dark necrotic lesions on exoskeleton, mortality. 115sions heavily covered in bacterial cells, with many embedded in an amorphous mucoid material on the lesion surface. 115trobacter C. freundii 108 Juveniles (GO, L) "Water bubble disease (WDB)-Formation of a "water bubble" with a diameter of c. 7 mm under the carapace, loss of appetite, inactivity, weight loss. 108t described.
V. cholerae 100,106 Larvae (L), postlarvae (L), juveniles (GO, L), adults (GO) Mortality in larvae and postlarvae. 106Red discolouration, anorexia, swimming alone in juveniles/ adults. 100 adults, rupture of basal lamina of hepatopancreatic tubules, with severe necrosis and dilation of tubules, loss of structure, atrophy and vacuolisation.Disorganisation of intestinal villus and epithelial cells with severe necrosis and separation from the basal membrane of epithelial cells. 100 harveyi 119,120 Larvae (H), juveniles (L) "Luminescent larvae syndrome"-glowing appearance of dead and moribund larvae.119 In juveniles-swelling and deformation of the hepatopancreas with appearance of white spots. Mortity. 120ot described in larvae.Chromatin condensation in hepatopancreatic cells of juveniles. 120 mimicus Larvae (L), postlarvae (L) Mortality.106 Not described.
V. parahaemolyticus (non-AHPND strains) 99,102 All (H, GO) Red discolouration with black spots on carapace, loss of appendages and telson, brittle shells and mortality in juveniles/adults. 99Mortality in larvae. 102 infection, suggesting that the virus can persist in M. rosenbergii, but does not replicate at high levels. 72WSSV has been shown to have reduced pathogenicity in other crustacean species, including the shore crab, Carcinus maenas. 74capod iridescent virus 1 (DIV1) (Family: Iridoviridae) is a large dsDNA virus. 75Members of Iridoviridae have been shown to infect amphibia, fish and invertebrates. 76DIV1 is an icosahedral enveloped virus with a diameter of approximately 160 nm, 75 but dependent on infected tissue type, DIV1 can be enveloped or non-enveloped, determined by whether they were budded from the cell membrane or released by cell lysis. 75,76To date, DIV1 comprises two strains: shrimp iridescent virus (SHIV) 20141215 and Cherax quadricarinatus iridovirus (CQIV) CN01, 77 with the former first identified in P. vannamei, and the latter in the red claw crayfish C. quadricarinatus. 75,78However, both strains have been shown to be able to infect P. vannamei. 78tural infections of M. rosenbergii with DIV1 have been reported in China, with cumulative mortalities of ≥80%.Affected prawns migrate to deep water and exhibit a white triangle under the carapace at the bottom of the rostrum. 75,79Mass mortalities due to infection with DIV1 have been reported from a pond in China where M. rosenbergii were co-cultured with Procambarus clarkii, a freshwater crayfish, on a farm that had seen high levels of mortalities from DIV1 in an adjacent P. vannamei pond a month prior to mortalities in M. rosenbergii. 79qPCR analysis of viral load showed high copy numbers of DIV1 in M. rosenbergii and P. clarkii, as well as in dried dead P. vannamei from the side of the pond that had experienced mortalities.Wild crustaceans also in the M. rosenbergii pond, including Macrobrachium superbum, M. nipponense and a Cladocera sp., also had detectable levels of DIV1, with M. nipponense having comparable viral loads to that of M. rosenbergii and P. vannamei.DIV1 has also been detected in F. chinensis and M. japonicus, suggesting a wide host range. 80DIV1 is a disease listed by WOAH. 81

| BACTERIA
Since the start of M. rosenbergii culture, opportunistic bacterial infections have been associated with mortalities and clinical signs of disease in all prawn life stages.Vibrio, Aeromonas and Pseudomonas spp.
have been associated with mortalities in the hatchery stage of culture through to adults in grow-out ponds. 5,84,85A summary of bacteria known to infect M. rosenbergii is provided in Table 2.
In juvenile and adult prawns, bacterial infections commonly manifest as 'black spot' lesions on the animal's exoskeleton; melanised, necrotic lesions, primarily on the appendages. 86As in other crustaceans, the causative agents Vibrio, Aeromonas, Pseudomonas and Bacillus have been associated with mortalities in all life stages of M. rosenbergii. 87,88Even when animals can be harvested before mortalities occur, market value is still reduced due to the substandard appearance of the animals. 86Aeromonas hydrophila has been associated with black spot disease in M. rosenbergii. 89Experimental Genus Species Reported life stage(s) affected Clinical signs of infection

Histopathological signs of infection
Infiltration of cells in intertubular spaces. 99Histopathology of larvae not described. 102 parahaemolyticus a (AHPND strains) 114,116 Larvae (L), adults (L) Mortality in larvae and adults.114,116 Not described in larvae.In adults, haematopoietic tissue showed karyorrhexis, pyknotic nuclei and accumulation of cells with eosinophilic structures. 116 V. nificus 98,121 Larvae (H), postlarvae (L), juveniles (GO) Dark brown focal lesions and necrosis on appendages of juveniles/adults. 98nfected larvae are weak, have poor appetite and slowed growth prior to mortality. 121 adults, necrosis of hepatopancreatic tubules and presence of bacterial haemocytic nodules with melanised and marked haemolytic enteritis.Accumulation of haemocytes in the haemocoelic space of gills and diffuse necrosis of gill lamellae.Erosion through the epicuticle of the exoskeleton extending to into the exocuticle. 98Histopathology of larvae not described.[92][93] Several other bacterial species have been associated with opaque and white musculature in the cephalothorax and abdominal segments of adult and juvenile M. rosenbergii.5][96] L. garvieae and L. lactis were first reported a decade apart in association with mortalities in ponds in Taiwan, with 30%-40% and 25%-60% mortality occurring in farms infected with each species, respectively. 94,95fection with Vibrio spp. was identified as a risk to M. rosenbergii farming when its culture was in its infancy.Huang et al. (1981) experimentally infected M. rosenbergii with a strain of V. anguillarum and observed mortalities of up to 100% with a high dose. 97Since then, numerous Vibrio species infections have been reported including V.

Note
vulnificus causing dark brown local lesions and necrosis of appendages, V. parahaemolyticus and V. cholerae, which both cause red discolouration of infected prawns, [98][99][100] and V. alginolyticus causing mortalities in adult prawns. 101rvae appear to be particularly susceptible to Vibrio spp. in hatchery settings.Ma et al. (2020) showed that four species of Vibrio, with multiple strains of each species, were prevalent in moribund hatcherycultured zoea-stage larvae. 85V. parahaemolyticus, V. neocaledonicus, V.
vulnificus and V. alginolyticus could cause larval mortalities in a dosedependent manner individually.However, mortalities in hatcheries were likely to be caused by co-infections of multiple Vibrio species and other opportunistic bacteria. 85Previous studies have also identified other species of Vibrio that cause larval mortalities including V. harveyi and V. campbellii. 102,103Other than Vibrio spp., the same study identified Enterobacter and Bacillus associated with moribund prawns.E. cloacae was previously identified as the causative agent of mortalities at the zoea stage of larval development in China, where mortalities exceeded 50% in hatcheries with affected larvae exhibiting reduced growth and rapid death when captured by netting. 104Despite multiple studies investigating the role of bacteria in larval disease and mortality, very few studies have investigated the bacterial communities involved in postlarval mortality. 105,106cently, Enterobacter cloacae has been associated with slow growth syndrome, also known as iron prawn syndrome (introduced in Section 2 -Viruses).Reports of E. cloacae in aquatic animals are rare; however, E. cloacae was detected in 100% of slow growth prawns collected from farms in China between 2017 and 2019.Challenging prawns experimentally with an isolate of E. cloacae reproduced the slower-growth phenotype, with slow-growth prawns significantly smaller in size than non-challenged prawns. 107study carried out in 2000 challenged juvenile M. rosenbergii with Citrobacter freundii but found no clinical signs of disease or mortalities associated with infections. 91Despite this, a 2022 study found C. freundii to be associated with juvenile mortalities of up to 30% in grow-out ponds in China, with laboratory challenges with C. freundii replicating clinical signs of disease seen in culture settings. 108Typically, prawns infected with C. freundii develop a 7mm diameter bubble under the carapace, show loss of appetite, weight loss and inactivity.Clinical signs of disease are quickly followed by mortality and spread of the disease to other prawns.The discrepancies in the results of these two studies could be explained by different strains of the same species of bacteria showing differing pathogenicity to the host, or differences in host genetics of the challenged prawns resulting in resistance to infection.
Many bacterial infections in M. rosenbergii appear to be opportunistic and caused by bacteria that are ubiquitous in the aquatic environment and/or known as (opportunistic) pathogens of other aquatic animals.However, other bacteria appear to be more specifically hostassociated.Spiroplasma eriocheiris, described from an infection of the Chinese mitten crab Eriocheir sinensis, has been identified as a pathogen of M. rosenbergii across Asia. 109,110S. eriocheiris, a mollicute with a distinctive helical morphology, is the causative agent of tremor disease in E. sinensis, which manifests as paroxysmal tremors prior to mortality. 111A Spiroplasma infection was first identified in M. rosenbergii in 2010 in China. 109This strain of Spiroplasma, provisionally named MR-1008, caused disease and mortalities in juveniles and adults in infected ponds.Experimental infections with the MR-1008 strain of Spiroplasma resulted in >80% mortality. 112,113A publication in 2013 investigating the proteomic response in M. rosenbergii to Spiroplasma infection confirmed that strain MR-1008 was S. eriocheiris. 113ere were no reports of S. eriocheiris infection outside of China until 2015, when naturally infected M. rosenbergii were reported from a pond in Thailand with unusually high mortality. 110Infections of S. eriocheiris have also been identified in wild river populations of M. rosenbergii in Bangladesh in 2018 and 2019 by molecular and histopathological techniques (Our own data, not published).

Acute hepatopancreatic necrosis disease (AHPND) causes morbidity
and mortality in post-larval penaeid shrimp, but it was not known until recently whether freshwater prawn species could be affected.AHPND, listed by WOAH in 2017, is caused by a strain of Vibrio parahaemolyticus made virulent by the acquisition of plasmids encoding the PirA Vp /PirB Vp toxin (VP AHPND ). 1145][116] Larvae were shown to have dose-dependent mortality in a disease challenge setting, with high doses inducing 100% mortality after 36 h. 117Postlarvae appear to be resistant to infection with VP AHPND ; however, they may have the potential to be a carrier to susceptible species that are commonly cultured together or in close proximity, 115 and as in the C.
freundii example above, may be due to host genetics of the challenged prawns conferring resistance to infection.Despite postlarvae appearing to be tolerant to VP AHPND , Pudgerd et al. (2021) demonstrated that adult M. rosenbergii could be infected with VP AHPND by intramuscular injection, and showed dose-dependent mortality. 116

| FUNGI
Fungi were some of the first pathogenic agents identified in M. rosenbergii culture, discovered prior to the commercial expansion of the industry.3][124][125] Typically, yeast infections in M. rosenbergii have been shown to occur in the winter months when water temperatures are cooler. 122A summary of fungi known to infect M. rosenbergii is provided in Table 3.
A Fusarium sp. was identified in 1977 as the causal agent of "black spot disease" in juvenile and adult M. rosenbergii. 122Similar to infection with opportunistic bacteria, isolated black (melanised) lesions were apparent on the carapace and appendages of prawns, typically when the carapace had been damaged, leaving an open wound vulnerable to infection. 122Fusarium spp.have also been associated with 'black gill' in penaeid shrimp and can cause mortality over a long period of time; however, there is no evidence that this fungus causes similar clinical signs in M. rosenbergii. 122 1998, several yeast taxa were identified as causal agents of mortality in adult and juvenile M. rosenbergii in Taiwan. 126Clinical signs of the disease included behavioural changes such as slow swimming, lethargy and anorexia accompanied by a yellow-brown discolouration, white opaque eyes and swelling between the cephalothorax and the abdomen.Upon dissection, infected prawns were seen to have cloudy-white haemolymph, a light yellow hepatopancreas and white opaque muscle tissue; under light microscopy, masses of Gram-positive yeast cells were observed. 126Candida sake was the most abundant yeast species in moribund prawns and is a possible contribution to the mortalities seen in Taiwan, as a previous study showed that C. sake could cause 100% mortality in experimental infections of this host. 124,126Another species of Juveniles, adults Greyish-white body colouration, spongy appearance, lethargy, anorexia, abnormal swimming, fading of eye colour, dark spots and melanisation on cephalothorax and muscles, greyish-white filaments on claws and mortality. 123ticular erosion and atrophy with fungal cells penetrating the cuticular epithelium and intramuscular regions.Gill lamellae infiltrated by proliferating yeast cells causing distension and gross enlargement. 123ndida sake Candida mogii 126,127 Juveniles, adults Yellowish-brown discolouration, white opaque eyes, swelling between cephalothorax and abdomen, cloudy white haemolymph, light yellow hepatopancreas, slow swimming, lethargic, anorexia, mortality.126 Vacuolisation of hepatopancreatic ducts and tubules with yeast aggregates in intertubular connective tissue. Largumbers of yeast cells present in haemolymph.126 Debaryomyces hansenii 125 Juveniles, adults Yellowish-brown discolouration, swollen hepatopancreas, cloudy haemolymph, whitish opaque musculature, mortality.125 Vacuolisation of hepatopancreatic epithelial cells, infiltration of yeast cells into the hepatopancreatic sinus.
Necrosis of whiteish muscle tissue.Large numbers of yeast cells present in haemolymph. 125sarium sp. 122uveniles, adults Dark lesions on exoskeleton following injury, mortality 122 Fusarium cells present in abdominal pleura, carapace, swimmerets, uropods and walking feet and appendages.Cuticular erosion, necrosis and melanisation of musculature underlying dark lesions on the exoskeleton 122 Metschnikowia bicuspidata 128 Juveniles, adults Poor growth, anorexia, yellowish-brown discolouration of body, swelling between cephalothorax and abdomen, swollen hepatopancreas, milky haemolymph, whitish, opaque musculature with whitish-yellow spots, mortality. 128cumulation of oedematous fluid between cuticle and muscle.Numerous yeast cells infiltrating the cuticle.Fragmentation of cardiac fibres and oedema in haemocyte nodules in the heart.Hepatopancreatic tubule epithelial cells vacuolised and sinuses contain thin membrane encapsulating yeast cells with large necrotic foci observed.Abdominal, pereiopod and pleopod muscle tissue has liquefactive necrosis and oedema with a large number of yeast cells.Necrosis of gill tissues with dilation and infarction of capillaries. 128crosporidia sp. 136uveniles, adults Not described Not described Enterocytozoon hepatopenaei (EHP) 135 Postlarvae Not described-PCR positive only.

Not described
Candida, C. mogii, has also been associated with infection of M.
rosenbergii in China. 127 2001, juvenile M. rosenbergii experienced mortality events associated with similar clinical signs to those of infection with C. sake.
Metschnikowia bicuspidata was isolated from diseased prawns coinfected with Enterococcus faecium. 128Experimental challenges of M. rosenbergii with M. bicuspidata alone and M. bicuspidata with E. faecium resulted in similar clinical signs, with the co-infection challenge only slightly increasing mortality. 125A separate study in 2007 showed that a high prevalence of M. biscupidata was associated with juvenile prawn mortalities of up to 95% in some ponds. 129Other yeast species have also been associated with mortalities, including Saccharomyces cerevisiae and Candida albicans, both of which were shown to be pathogenic by experimental infection. 129M. rosenbergii infected with Debaryomyces hansenii also shows similar clinical signs to infection with M. bicuspidata. 125e chytrid Batrachochytrium dendrobatidis was identified as the causative agent of discolouration and prawn mortality in M. rosenbergii farms in southern India between 2007 and 2011. 123Affected prawns were greyish white, had a 'woolly' appearance with greyish filaments on their claws, and had problems with feeding, moulting, movement and respiration-culminating in mortality, reaching up to 90%.

| MICROSPORIDIA
In penaeid shrimp culture, the microsporidian parasite, Enterocytozoon hepatopenaei (EHP) of the Enterocytozoon Group Microsporidia (EGM) causes slow-growth in P. monodon and P. vannamei. 130,131EHP is associated with a wider range of syndromic conditions, for example, white faeces syndrome 132 and is now a major threat to the commercial shrimp farming industry, listed as an emerging disease by WOAH 133 First identified in P. monodon in ponds in Thailand, at the time, it was not considered to be causing significant problems in shrimp culture, 131 but since, although not linked to mortalities, it has spread globally, causing severe growth retardation and resulting in enormous losses for shrimp farmers. 134A study in 2018 showed that M. rosenbergii cultured in the same pond as P. vannamei were positive by PCR for EHP; however, the histology was unable to demonstrate infection of M. rosenbergii. 135Despite the inconclusiveness of this study, molecular detection of EHP from M. rosenbergii is a significant finding and should be explored in more depth to determine whether freshwater prawn species are susceptible to infection with EHP (or other EGM) to the same extent as penaeid shrimp.
Microsporidia infections have been observed in M. rosenbergii, 136 but have not been taxonomically described further than to the genus level. 137A microsporidian parasite has been identified in M. nipponense as the novel species Potaspora macrobrachium, causing progressive whitening of musculature and reduced survivability during holding and transportation of the animals. 138The description of this novel disease-causing microsporidian is an important discovery in freshwater prawn culture and should be considered as a potential threat to the culture of M. rosenbergii.

| OTHER EUKARYOTIC PATHOGENS
Eukaryotic pathogens other than fungi and microsporidia are not commonly reported in M. rosenbergii culture, likely due to the lack of, or low, mortalities associated with infection.The presence of epibionts, principally ciliates, are common in M. rosenbergii ponds predominantly associated with larger animals that moult less frequently, allowing the accumulation of symbionts on the exoskeleton. 8The presence of these epibionts can cause problems with feeding and mobility when attached to appendages, and in cases with severe fouling of the gills, mortality can occur from insufficient gaseous exchange. 8o publications have investigated the presence of eukaryotic parasites in wild populations and showed that M. rosenbergii could be infested by a range of epi-and endo-parasitic infections. 139,140Both publications describe the presence of ciliates, with Zoothamnium spp.
present at high prevalence on appendages and gills as well as Acineta spp., Epistylis spp.and Vorticella spp.The latter two were found both externally and within the gut.Jayasree et al. ( 2001) reported that apostome ciliate cysts were commonly seen attached to gill lamellae, with heavy infections frequently observed, causing a melanisation response at points of attachment. 140Gut-dwelling gregarines were also present in both M. rosenbergii populations. 139,140Other eukaryote parasites reported include bopyrid isopods and the larval digeneans Opecoelid metacercariae attached to pleopods, pereiopods and antennules and microphallid metacercariae in the musculature.Berried females may be a source of introduction of epibionts into hatchery culture systems, where they cause greater issues with feeding and swimming impairment due to the small size of M. rosenbergii larvae and postlarvae. 8spite the infrequent reports of mortality caused by eukaryotic pathogens in M. rosenbergii, one ciliate, Metanophrys sinensis, has been associated with mass mortalities of larval stage M. rosenbergii in India; prawns exhibiting lethargy, erratic movement and discolouration prior to mortality.Under microscopic examination, large numbers of ciliates were seen in the coelom. 141erwise, microeukaryote pathogens associated with Macrobrachium have received little attention.As awareness of the importance of host-associated eukaryotes (the 'eukaryome') increases, particularly with respect to host health, investigating this aspect of Macrobrachium health, particularly using the new approaches available to profile unknown pathogens, 142 will likely provide valuable information about symbiotic relationships contributing to health and disease in this host and those cultured with it.

| EMERGING AND SYNDROMIC DISEASES
The last review of diseases in M. rosenbergii 24 identified several diseases of uncertain aetiology.Many of these apparently syndromic conditions remain unresolved currently.
Many idiopathic diseases affect the early life stages of M. rosenbergii.Mid-cycle disease (MCD) affects larval stages of M. rosenbergii, typically between days 15 and 22 of production; larvae display clinical signs of lethargy, erratic swimming activity, reduced feeding and growth and a bluish discolouration of the body. 143Moult death syndrome (MDS), also known as exuviae entrapment disease (EED), is a phenomenon that most commonly occurs during the metamorphic moult of larvae to postlarvae, where larval mortality occurs during unsuccessful moults. 86Dietary supplements have been shown to reduce the occurrence of MDS. 144anchiostegal blister disease (BBD), named by farmers as balloon disease (BD), since the disease is characterised by swelling of the gill flap or branchiostegal region and deformities of the appendages. 145e disease occurs in grow-out ponds, typically within 30 days of stocking postlarvae from nurseries, with mortalities ranging from 70% to 80%.As the disease spreads rapidly between adjacent ponds, a viral aetiology was proposed; however, traditional methods of virus isolation did not detect the presence of any viruses. 145pendage deformity syndrome (ADS) was reported to cause late mortalities in grow-out ponds in the Nellore district of India in 2004. 146The disease typically affected female prawns more than males, and prevalence in affected ponds was 50%.Affected prawns had appendage deformities, broken or bent rostrums, cut antennae and wrinkled carapaces.Cohabitation challenges were unable to cause ADS in unaffected prawns, suggesting that disease cannot be transferred from prawn to prawn.Interestingly, supplementation of feed with carotenoids was able to reverse the diseased phenotype, indicating that ADS is not of pathogenic aetiology. 146owth retardation of prawns with the idiopathic condition known as iron prawn syndrome (IPS) affects the body weight and length of prawns in culture.Attempts to determine a cause of this abnormal phenotype have been significant compared to other idiopathic diseases, with studies investigating culture practices, pathogen-related disease candidates and host genetics.Studies assessing diet, 45 antibiotic use 147 and water quality 44 have not identified the leading cause of the condition.
Some studies investigated growth based on single-nucleotide polymorphisms (SNPs) and host transcriptomics. 148,149However, these mainly focused on size differences present in populations due to social dominance hierarchy, not prawns affected by IPS.Jiang et al.
(2020) investigated the genetic and epigenetic differences between prawns with a normal growth phenotype and IPS prawns and noted differences mainly in the germplasm of the two phenotypes. 43Interestingly, the same study reported enrichment for host transcripts related to immune response in infected animals, suggesting that a host response may be being initiated due to pathogen infection.Since then, a study comparing the differences in proteomics between normal and IPS prawns also reported significant variations in the expression of proteins involved in immune system functions. 150Further evidence to suggest pathogen involvement was the identification of E. cloacae infections in growth-retarded prawns 107 (Section 3 -Bacteria), the presence of a novel hepe-like virus, 48 and the presence of a novel flavivirus 49 (see Section 2 -Viruses).Screening for agents associated with IPS is missing from the three recent investigations, for example, the study that identifies IPV did not screen for Enterobacter cloacae or CHEV1, therefore it is unclear whether these pathogens can cause IPS phenotype alone, or infections of multiple pathogens are needed.

| DISCUSSION
Since a review of diseases of M. rosenbergii in 2012, 24 the number of its characterised pathogens (Figure 2 Shrimp aquaculture (P.monodon and P. vannamei) has seen more large-scale disease outbreaks than other groups of cultured aquatic animal species. 151These disease outbreaks, causing total losses exceeding 40% of global capacity, 152 have a devastating economic and socio-economic impact on shrimp farms. 153In M. rosenbergii culture, disease incidence, in particular diseases caused by viral infections, has increased with the expansion of the industry, but with no clear cause.A review of the vulnerabilities in aquatic animal production identified a number of factors contributing to increase in disease incidence, one being climate change. 154ort-term climate events such as increased incidences of storms, floods and droughts can cause instabilities in pond systems, resulting in poor water quality and a chance for opportunistic pathogens to cause disease in stressed animals.These events are likely to become more frequent and extreme as the climate crisis continues. 155Both climate change and intensification of culture has been shown to drive increased pathogen virulence, host susceptibility and incidence of disease. 156,157Inherent production and management risks also contribute to the rate of disease incidence, with the increase in production resulting in wild stocks being transported into aquaculture settings with few biosecurity measures to manage potential pathogens they may bring into the system. 154portunistic bacterial infections have the potential to cause significant losses in all stages of M. rosenbergii culture.To limit the losses due to this type of infection, use of antibiotics is common practice in culture settings, with recommendations on the type of antibiotic suitable for treating certain disease phenotypes outlined in the FAO culture handbook. 5Despite cautions surrounding the misuse of antibiotics in the handbook, these compounds have been reported to be used as a preventative measure against bacterial infections in both shrimp and other aquaculture species. 158,159Commonly used antibiotics in M. rosenbergii culture include oxolinic acid, chloramphenicol, erythromycin and oxytetracycline. 8Several studies have identified many bacteria, isolated from both prawns 160,161 and the culture environment they are grown in, 161  ing that molluscs were able to concentrate the virus by filter-feeding, delivering an infectious dose to shrimp when the molluscs were ingested. 163 China, M. rosenbergii polyculture with E. sinensis poses a significant risk of infection with S. eriocheiris, shown to infect both E. sinensis (causing tremor disease) and M. rosenbergii, with infections in both species leading to mass mortalities. 109,111Other species of crustaceans have been shown to be susceptible to infection with Spiroplasma spp., including P. vannamei 164 and P. clarkii, 165 with infections associated with mortalities.Given that M. rosenbergii is grown in polyculture systems with E. sinensis, P. vannamei and P. clarkii, 13,21,79 infection with S. eriocheiris or another Spiroplasma species is a significant risk.
A review in 2017 listed known viruses of crustaceans, with a large number infecting crab, including E. sinensis. 166Little is known about whether M. rosenbergii is also a susceptible species to these pathogens but given the broad host range of some crustacean pathogens, these risks should be considered when choosing this culture method.Interestingly, AHPND, caused by a halophilic bacterial pathogen, does not appear to pose a significant concern to M. rosenbergii under polyculture with P. vannamei at low salinities, with the prawn postlarvae and juveniles shown to exhibit resistance to VP AHPND , unlike P. vannamei, which is more susceptible to AHPND at higher salinities (e.g. 10, 15 and 20 ppt) than at 5 ppt. 115 many countries, M. rosenbergii is cultured in the same or adjacent ponds as penaeid shrimp, such as P. monodon and P. vannamei. 13,20,79Some hatcheries also produce freshwater prawns and penaeid shrimp within close proximity, increasing the likelihood of cross-contamination of species and the pathogens they are potentially carrying. 167An obvious concern relates to viruses with a wide host range such as WSSV, CMNV, IHHNV and DIV1 that are known to infect both marine and freshwater species.However, a greater concern is infection of supposedly non-susceptible species by these and other pathogens-white tail disease caused by MrNV in Indian hatchery-cultured penaeid shrimp, P. monodon and Penaeus (Fenneropenaeus) indicus, is a good case study that highlights this issue. 167Seed of the P. monodon and P. indicus was produced in very close proximity to the seed of M. rosenbergii; the postlarvae of the two penaeid shrimp species exhibited whitish abdominal muscle and lethargy, with mortality reaching 100% after the appearance of clinical signs. 167Since this disease incident, WTD has been reported in P. vannamei in China and Vietnam, 168,169 suggesting that this is not an isolated case and WTD is a threat to both freshwater and marine hosts.Experimental infection of juvenile penaeid shrimps with MrNV and XSV was unable to cause WTD, but juveniles and adults could act as virus reservoirs for larval and post-larval life stages. 170The finding that older life-stages could act as a reservoir is important in the context of berried females being used as broodstock to produce larvae in hatchery settings-many countries prefer to use berried females from rivers, canals and lakes, 5 potentially unknowingly transporting pathogens into the culture system.For almost all pathogens of M. rosenbergii, it is unknown whether berried females and their eggs can act as a reservoir, therefore caution should be taken when transporting animals with an unknown disease status into hatcheries and other culture settings.
Given the major problems currently being experienced in penaeid shrimp culture caused by the microsporidian parasite EHP, it would be naïve to assume that M. rosenbergii are not at risk from microeukaryotic pathogens-at the time of its identification, EHP was considered an incidental hazard but has since emerged as a significant disease risk to the industry. 171Despite microsporidia being ubiquitous within the environment and being able to infect vertebrates and invertebrates, 172 no microsporidian parasites have been formally described to infect M. rosenbergii, despite reports of microsporidiosis in the literature.Other eukaryotic pathogens, such as members of the Ascetosporea class of parasites, also cause problems in crustacean culture, such as the haplosporidian parasite that has been associated with high mortality and slow growth in P. vannamei. 173 addition to a gap in our understanding of eukaryotic pathogens of M. rosenbergii, there is also information missing from the literature surrounding the life-stages that pathogens affect, especially whether viruses that are lethal in postlarvae/juveniles can also infect larvae and adults.This is especially important at the stages in culture when animals are moved to new locations e.g. from hatchery to grow-out pond, where pathogens could be transferred from one life stage to another.
As the cost of sequencing continues to decrease, sequencing of all host and non-host genetic material in a sample by either metagenomics or metatranscriptomics is being applied ever more widely.As these technologies become more accessible, a greater understanding of pathogen lifecycles can be obtained by sequencing of hosts, intermediate and potential hosts of pathogens, as well as the surrounding environment.A holistic approach is beginning to be applied to pathogens causing the biggest problems in prawn aquaculture, such as MrNV and XSV.For example, natural insect carriers of viable MrNV and XSV particles have been identified in nursery ponds containing M. rosenbergii with clinical signs of WTD, 174 adding to the information currently available on transmission of the virus.A better understanding of pathogen transmission and lifecycle will enable decisions to be made, considering the risks of pathogen transfer between species and the environment, and the implementing of measures to mitigate disease.
Other molecular tools to investigate host genetics to improve M. rosenbergii culture are in their infancy-SNPs have been identified by mining transcriptomic data and have been utilised to search for markers of growth performance. 149However, there is much greater potential for this type of tool to be used to investigate genetic markers of resistance to diseases.Breeding resistance to diseases based on genetic markers could potentially replace SPF stocks that are still susceptible to infection.The most notable use of this type of technology in aquaculture has been in the identification of a single quantitative trait locus (QTL) in Atlantic salmon (Salmo salar) that confers resistance to infection with infectious pancreatic necrosis virus (IPNV), allowing the production of resistant stocks using markerassisted selection programs. 175A recent study has also shown that susceptibility of P. monodon to infection with GAV is heritable, 176 suggesting that the identification of the genetic markers involved could potentially lead to selective breeding programs to produce animals that are more tolerant, or even resistant, to diseases that cause major problems in aquaculture.
Other than the utilisation of molecular tools and breeding for disease resistance, further techniques to mitigate the risk of disease in systems have been tested and applied to giant prawn culture.Such techniques include the addition of probiotics 177 and dietary supplements 178 to the aquaculture environment, as well as changing culture systems to Recirculating Aquaculture Systems (RAS) or biofloc systems. 179These changes have seen some success, including observations that the addition of probiotics and dietary supplements can protect against disease. 177,178As a result of these positive outcomes, a more holistic approach, combining multiple methods determined to be beneficial to giant prawn culture, is starting to be adopted. 180Further adoption of a combination of methods to improve the health of M. rosenbergii is likely to reduce disease risk to prawns and animals they are cultured with.for his detailed editing of the English language of this manuscript.
water source, food or other aquaculture species Pathogen transfer from/to other aquaculture species in pond Pathogen transfer from water source, wild species and poor biosecurity between ponds !Antimicrobial resistance due to misuse of antibiotics F I G U R E 1 Culture cycle of M. rosenbergii highlighting the potential for introduction of disease into the system T A B L E 1 Viruses infecting M. rosenbergii Virus Family Structure Clinical signs of infection Histopathological signs of infection Covert mortality nodavirus (CMNV) also screened prawns from farms with and without IPS to determine whether the virus was present.All prawns collected from farms experiencing IPS were positive for IPS, whereas all farms with no gross clinical signs of IPS were negative by PCR.The role of emerging pathogens in giant river prawn aquaculture was highlighted in Bangladesh in 2011.Hatcheries in southern Bangladesh experienced mass mortalities of larval M. rosenbergii, resulting in a decline in the number of hatcheries actively producing M. rosenbergii from >60 to 12, and a drop in the number of postlarvae produced from >200 million to 27.75 million in the last decade.50Affected larvae exhibited problems with swimming, feeding and growth, with moribund animals appearing whitish in colour compared to healthy animals.A recent study determined that a novel 29,110 nt positive-sense ssRNA virus, Macrobrachium rosenbergii Golda virusT A B L E 1 (Continued)

70 a
Predicted family classification based on sequence similarity and/or virus ultrastructure.b Pathogens listed by the World Organisation for Animal Health (WOAH).
Gangnonngiw et al. (2010) identified that M. rosenbergii postlarvae from Thailand had similar lesions in the hepatopancreas as that of P. monodon in the early stages of infection with PmNV.62 Hepatopancreatic epithelial cells had enlarged nuclei with diffuse, central, eosinophilic inclusions.No mortalities were associated with infection with PmNV in M. rosenbergii; however, some individuals were noted to have co-infections of PmNV and HPV.As these infections have only been identified by histological observations, it is not clear whether the nudivirus infections seen in M. rosenbergii are caused by the same nudivirus (PmNV) that infects penaeid shrimp.As nudiviruses are quite diverse, genomic comparisons are needed to determine whether these viruses are the same or closely related.

:
Brackets in the 'reported life stage(s) affected' column indicate which settings infections were reported: H = Hatchery, N = Nursery, GO = Growout pond, W = Wild, L = Laboratory setting.a Pathogens listed by the World Organisation for Animal Health (WOAH).

T A B L E 3
Diseases of Macrobrachium rosenbergii caused by fungi and microsporidia Species Reported life stage(s) affected Clinical signs of infection Histopathological signs of infection Batrachochytrium dendrobatidis 123

F I G U R E 2
Diseases of Macrobrachium rosenbergii and the life stage(s) affected.Black circle with cross denotes that this pathogen has been shown to cause mortality.Coloured circles denote the organ(s) affected: Green = cuticle, pink = gut, yellow = gill, orange = appendages, blue = hepatopancreas, red = muscle, purple = eye/eyestalk and grey = unknown ) has increased.Many of these pathogens are viruses that have either emerged directly within M. rosenbergii culture, or are known pathogens from other aquaculture sectors that have also been shown to infect M. rosenbergii.Identifying pathogens throughout aquaculture has become easier with the decreasing cost of next-generation sequencing techniques that have allowed culture-free identification of pathogens-particularly useful where the agent is novel and favourable culture environments are not known or possible to reproduce.
Conceptualization; formal analysis; writingoriginal draft; writingreview and editing.Partho P. Debnath: Writingreview and editing.Grant D. Stentiford: Writingreview and editing.Kelly S. Bateman: Writingreview and editing.Krishna R. Salin: Writingreview and editing.David Bass: Funding acquisition; supervision; writingoriginal draft; writingreview and editing.Department of Biotechnology under the Newton Fund Global Research Partnership in Aquaculture programme BB/N00504X/1 (to David Bass).The authors would like to express their gratitude to David Currie 63 to be resistant to a number of antimi-