AIM To study the cell types,localization,distribution density and morphology of APUDcells in the intestinal mucosa of stomachlessteleost fishes.METHOD By using the peroxidase-antiperoxidase complex(PAP)immunocytochemi...AIM To study the cell types,localization,distribution density and morphology of APUDcells in the intestinal mucosa of stomachlessteleost fishes.METHOD By using the peroxidase-antiperoxidase complex(PAP)immunocytochemical staining technique theidentification,localization and morphology ofimmunoreactive(IR)endocrine cells seattered inthe intestinal mucosa of grass carp(Cyenopharyngodon idellus),black carp(Mylopharyngodon piceus)and common carp(Cyprinus carpio)were investigated with 20kinds of antisera prepared against mammalianpeptide hormones of APUD cells,and likewise byusing avidin-biotin-peroxidase complex(ABC)method those of silver carp(Hypophthalmichthys molitrix),bighead(Aristichthys nobilis),silver crucian carp(Carassius gibelio)and bluntnose black bream(Megalobrama amblyocephala)were alsostudied with 5 different antisera.Thereplacement of the first antiserum by phosphatebuffered saline(PBS)was employed as a control.IR endocrine cells were counted with asquare-mesh ocular micrometer from 10 fieldsselected randomly in every section of each partof the intestine specimen.The average numberof IR endocrine cells per mm2 was counted toquantify their distribution density.RESULT Gastrin(GAS)-,Gastric inhibitorypeptide(GIP)-,glucagon(GLU)-,glucagon-likeimmunoreactants(GLI)-,bovine pancreaticpolypeptide(BPP)-,leucine-enkephalin(ENK)-and substance P(SP)-IR endocrine cells werefound in the gut of grass carp,black carp andcommon carp,and somatostatin(SOM)-IRendocrine cells were only seen in common carp.GAS-,GIP-and GLU-IR endocrine cells werefound in the intestinal mucosa of silver carp,bighead,silver crucian carp and bluntnose blackbream.Most of IR endocrine cells had the higherdistribution density in the foregut and midgut,and were longer in shape.They had a long apicalcytoplasmic process extended to the gut lumenand a basal process extended to adjacent cellsor basement membrane and touched with it.Sometimes,the basal cytoplasmic processformed an enlarged synapse-like structure in thecontiguous part with basement membrane.Thisphenomenon provided new morphologicalevidence for neuroendocrine and paracrinesecretory function of these enteroendocrinecells.CONCLUTION At least 8 kinds of IR endocrinecells were found in the gut of stomachlessteleost species for the first time in China.TheseIR endocrine cells scattering in the gut mucosabelong to the APUD system.Among them,thehormones secreted by SP-,ENK-,SOM-and GLU-IR endocrine cells belong to the peptides of dualdistribution in the brain and gut.This providednew evidence for the concept of brain-gutpeptide.According to the cell types,distribution density,morphologicalcharacteristics and variety in shape of APUDcells in the gut of stomachless teleost fishes,itis deemed that the digestive tract of fishes isalso an endocrine organ of great importance andcomplexity.展开更多
AIM To identify the type localization andmorphology of APUD endocrine cells in thegastroenteropancreatic(GEP)system ofstomach-containing teleosts,and study APUDendocrine system in the stomach,intestine andpancreas of ...AIM To identify the type localization andmorphology of APUD endocrine cells in thegastroenteropancreatic(GEP)system ofstomach-containing teleosts,and study APUDendocrine system in the stomach,intestine andpancreas of fish species.METHODS Two kinds of immunocytochemical(ICC)techniques of the streptavidin biotin-peroxidase complex(SABC)and streptavidin-peroxidase(S-P)method were used.Theidentification,localization and morphology ofAPUD endocrine cells scattered in the mucosa ofdigestive tract,intermuscular nerve plexus andglandular body of northern snakehead(Channaargus),ricefield eel(Monopterus albus),yellow catfish(Pelteobagrus fulvidraco),mandarinfish(Siniperca chuatsi),largemouthbass(Micropterus salmoides),orientalsheatfish(Silurus asotus),freshwater pomfret(Colossoma brachypomum)and nile tilapia(Tilapia nilotica)were investigated with 8 kindsof antisera.RESULTS The positive reaction of 5-hydroxytryptamine(5-HT)immunoreactiveendocrine(IRE)cells was found in the digestive tract and glandular body of 8 fish species indifferent degree.Only a few gastrin(GAS)-IREcells were seen in C.argus,M.albus and P.fulvidraco.Glucagon(GLU)-IRE cells were notfound in the digestive tract and glandular bodybut existed in pancreatic island of most fishspecies.The positive reaction of growthhormone(GH)-IRE cells was found only inpancreatic island of S.Chuatsi and S.Asotus,no positive reaction in the other 6 fish species.Somatostatin(SOM)-,calcitonin(CAL)-,neurofilament(NF)-and insulin(INS)-IRE cellsin the stomach,intestine and pancreas of 8 kindsof fish were different in distribution and types.The distribution of all 8 APUD cells was the mostin gastrointestinal epithelium mucosa and then indigestive glands.The positive reaction of SOM-and 5-HT-IRE cells was found in intermuscularnerve plexus of intestine of P.fulvidraco andS.chuatsi.Only GH-IRE cells were denselyscattered in the pancreatic islands of S.chuatsiand S.asotus,and odd distribution in thepancreas of S.asotus,SOM-IRE cells weredistributed in the pancreatic islands of S.asotus,C.Brachypomum and T.nilotica.There were INS-IRE cells in the pancreaticislands of S.chuatsi and S.asolus.Eightkinds of APUD cells had longer cell body andcytoplasmic process when they were located inthe gastrointestinal epithelium,and had shortercell body and cytoplasmic process in the gastricgland,and irregular shape in the esophagus andpancreatic island.CONCLUSION Eight kinds of IRE cells were identified in the GEP system of stomach-containing teleosts. These endocrine cells were scattered in gastrointestinal mucosa, intermuscular nerve plexus, gland body, pancreatic gland and islands under APUD system. CAL- and GH-IRE cells in the pancreatic islands of fishes showed functional diversity for these two hormones. Their morphological feature provides evidence of endocrine-paracrine and endocrine-exocrine acting mode. This research can morphologically prove that the GEP endocrine system of fish ( the lowest vertebrate) is almost the same as of mammal and human.展开更多
Macrophages exist in most tissues and play a variety of functions in vertebrates.Teleost fish species are found in most aquatic environments throughout the world and are quite diverse for a group of vertebrate animals...Macrophages exist in most tissues and play a variety of functions in vertebrates.Teleost fish species are found in most aquatic environments throughout the world and are quite diverse for a group of vertebrate animals.Due to whole genome duplication and en vironme ntal adaptati on,teleost monocytes/macrophages possess a variety of different functions and modulations compared with those of mammals.A deeper understanding of teleost monocytes/macrophages in the immune system will not only help develop teleost-specific methods of disease prevention but will also help improve our understanding of the various immune mechanisms in mammals.In this review,we summarize the differences in polarizati on and phagocytosis of teleost and mammalian macrophages to improve our understanding of the various immune mechanisms in vertebrates.展开更多
In this current paper, the exposure time effects on four endocrine disruptors and teleost fishes were evaluated using the reduced life expectancy (RLE) model based on the effect concentration (EC<sub>50</sub&...In this current paper, the exposure time effects on four endocrine disruptors and teleost fishes were evaluated using the reduced life expectancy (RLE) model based on the effect concentration (EC<sub>50</sub>) of available literature published. The result on the regression analysis over different exposure times has demonstrated that the EC<sub>50</sub> of hepatic biomarkers falls with increasing exposure times in a predictable manner. The slopes of the regression equations reflect the strength of the toxic effects on the various teleost fish. The EC<sub>50</sub> reduction over time can be interpreted based on the bioconcentration process, which can be used to understand transfer routes of the compounds from water to fish body. RLE model also provides useful information in assessing the toxic effects on fish life expectancy as a result of the occurrence of compounds.展开更多
Time-dependent tissue distribution of mercury(Hg) was studied in a freshwater perch,Anabas testudineus which revealed that the liver and kidneys are the major sites of Hg reten tion. The role of reduced glutathione (...Time-dependent tissue distribution of mercury(Hg) was studied in a freshwater perch,Anabas testudineus which revealed that the liver and kidneys are the major sites of Hg reten tion. The role of reduced glutathione (GSH) in the clearance of Hg was also investigated to e valuate the ameliorative effect of this nucleophile. For this purpose, the perch was given GSH 15 min before or after they received 203Hg by injection. The fish were then sacrificed at 24 h and 48 h later. The results clearly indicate that exogenous GSH can significantly reduce Hg retention in both the liver and kidneys, demonstrating a direct role of this nucleophile in the amelioration of Hg-induced toxicity in the early phase of intoxication展开更多
The toxic effect of fertilizer Diammonium phosphate resulted in alterations of 5'-Nucleotidase activity of tissues liver, kidney and muscles offish C. batrachus at varying intervals and exposures. Alterations in 5...The toxic effect of fertilizer Diammonium phosphate resulted in alterations of 5'-Nucleotidase activity of tissues liver, kidney and muscles offish C. batrachus at varying intervals and exposures. Alterations in 5'-Nuclcotidase activity of body organs gave an idea of the toxicity caused by the fertilizer. Thus the enzyme 5'-Nucleotidase can be used to monitor the pollution in aquatic ecosystem.展开更多
NLRP3 inflammasome activation is pivotal for cytokine secretion and pyroptosis in response to diverse stimuli,playing a crucial role in innate immunity.While extensively studied in mammals,the regulatory mechanisms go...NLRP3 inflammasome activation is pivotal for cytokine secretion and pyroptosis in response to diverse stimuli,playing a crucial role in innate immunity.While extensively studied in mammals,the regulatory mechanisms governing NLRP3 activation in non-mammalian vertebrates remain largely unexplored.Teleosts,as basal vertebrates,represent an ideal model for exploring the evolutionary trajectory of inflammasome regulation.In this study,ABE assays,confocal microscopy,and biochemical analyses were applied to systematically characterize the mechanisms underlying NLRP3 inflammasome in teleosts,using large yellow croakers(Larimichthys crocea,Lc)and zebrafish(Danio rerio,Dr)as representative models.Our findings revealed a previously unrecognized palmitoylation-dependent regulatory mechanism essential for teleost NLRP3 activation.Specifically,zDHHC18-mediated palmitoylation at a teleost-specific cysteine residue(C946 in LcNLRP3,C1037 in DrNLRP3)was required for the translocation of NLRP3 to the dispersed trans-Golgi network,facilitating its subsequent recruitment to the microtubule-organizing center.This membrane trafficking was crucial for inflammasome assembly and downstream inflammatory responses.These findings provide new insights into the distinct regulatory mechanisms of NLRP3 activation in teleosts,highlighting an evolutionary divergence that contributes to innate immunity adaptation in early vertebrates.展开更多
The defense system of teleost fish organized on innate and adaptive immunity protects them against a wide variety of pathogenic microorganisms in the aquatic environment.Phagocytosis is one of the most effective defen...The defense system of teleost fish organized on innate and adaptive immunity protects them against a wide variety of pathogenic microorganisms in the aquatic environment.Phagocytosis is one of the most effective defense strategies against microbial challenge mainly performed by classical‘professional’phagocytes(including monocytes,macrophages and granulocytes).They contain,kill and process the internalized pathogens for antigen presentation by providing antigenic ligands to initiate activation and clonal expansion of T and B cells,which bridge the innate and adaptive immunity.The discovery of phagocytic B cells in teleost fish has broken the paradigm that primary vertebrate B cells are lack of phagocytosis of particulates,as well as led to the investigation of phagocytic activity of mammalian B-1 B cells.The active phagocytic,microbicidal capabilities and antigen presentation in teleost phagocytic B cell have demonstrated to be similar as professional phagocytes,providing a potential impact on development of new vaccination strategies to prevent and control infectious diseases.In this review,we aim to address current progress on the antimicrobial role of phagocytic B cells in teleost fish by comparing it with other professional phagocytes and mammalian B-1 B cells,and provide the application prospect of phagocytic B cells in developing vaccines as well as the prevention of fish diseases.展开更多
The origin of T cells in the teleost's brain is unclear.While viewing the central nervous system(CNS)as immune privileged has been widely accepted,previous studies suggest that T cells residing in the thymus but n...The origin of T cells in the teleost's brain is unclear.While viewing the central nervous system(CNS)as immune privileged has been widely accepted,previous studies suggest that T cells residing in the thymus but not in the spleen of the teleost play an essential role in communicating with the peripheral organs.Here,we identified nine T cell subpopulations in the thymus and spleen of orange-spotted grouper(Epinephelus coioices)through single-cell RNA-sequencing analysis.After viral CNS infection with red-spotted grouper nervous necrosis virus(RGNNV),the number of slc43a2^(+)T cells synchronously increased in the spleen and brain.During the infection tests in asplenic zebrafish(tlx1^▲zebrafish model),no increase in the number of slc43a2^(+)T cells was observed in the brain.Single-cell transcriptomic analysis indicated that slc43a2^(+)T cells mature and functionally differentiate within the spleen and then migrate into the brain to trigger an immune response.This study suggests a novel route for T cell migration from the spleen to the brain during viral infection in fish.展开更多
Gonadal soma-derived factor(Gsdf)is a member of the transforming growth factor beta(TGF-β)superfamily and a teleost-specific gene.Early studies suggested that Gsdf promotes spermatogonial proliferation;however recent...Gonadal soma-derived factor(Gsdf)is a member of the transforming growth factor beta(TGF-β)superfamily and a teleost-specific gene.Early studies suggested that Gsdf promotes spermatogonial proliferation;however recent reports have shed light on its role in sex differentiation based on expression profiles and gain-and loss-of-function studies.This paper discusses the involvement of Gsdf in gonadal sex differentiation and sex changes in teleosts.展开更多
In mammals,chemokines are a superfamily of cytokines that regulate cell migration or exert direct antimicrobial activity.The presence and organization of four invariant cysteine residues within the mature protein sequ...In mammals,chemokines are a superfamily of cytokines that regulate cell migration or exert direct antimicrobial activity.The presence and organization of four invariant cysteine residues within the mature protein sequence can classify chemokines into four major subfamilies:CXC,CC,CX_(3)C,and XC.In teleost,a novel subfamily of chemokines named CX has been identified.Since the first report of a CC chemokine(CK1)in rainbow trout in 1998,the number of identified chemokine genes in different fish species has expanded dramatically.In this paper,we reviewed the chemokine genes that have been identified so far in teleost fish and their tissue expressions and responses post-stimulation.We also discussed the functions of fish chemokines based on currently available information and compared them to human chemokines.Additionally,we predicted the structures of trout chemokines for the first time,and we found that the structures of trout chemokines share significant similarities.Finally,the adjuvant effects of fish chemokines in aquaculture were also reviewed.Overall,this review will give us a better understanding of the chemokines of teleost fish.展开更多
Viral infection induces the initiation of antiviral effectors and cytokines which are critical mediators of innate antiviral responses.The critical molecular determinants are responsible for triggering an appropriate ...Viral infection induces the initiation of antiviral effectors and cytokines which are critical mediators of innate antiviral responses.The critical molecular determinants are responsible for triggering an appropriate immune response.Long noncoding RNAs(lncRNAs)have emerged as new gene modulators involved in various biological processes,while how lncRNAs operate in lower vertebrates are still unknown.Here,we discover a long noncoding RNA,termed antiviral-associated long noncoding RNA(AANCR),as a novel regulator for innate antiviral responses in teleost fish.The results indicate that fish MITA plays an essential role in host antiviral responses and inhibition of Siniperca chuatsi rhabdovirus(SCRV)production.miR-210 reduces MITA expression and suppress MITA-mediated antiviral responses,which may help viruses evade host antiviral responses.Further,AANCR functions as a competing endogenous RNA(ceRNA)for miR-210 to control protein abundance of MITA,thereby inhibiting SCRV replication and promoting antiviral responses.Our data not only shed new light on understanding the function role of lncRNA in biological processes in teleost fish,but confirmed the hypothesis that ceRNA networks exist widely in vertebrates.展开更多
Endocrine disruptors(EDs)are synthetic or natural chemical molecules occurring in environment that have the potential to impart adverse effects on homeostasis of endocrine axis leading to neurological,developmental,im...Endocrine disruptors(EDs)are synthetic or natural chemical molecules occurring in environment that have the potential to impart adverse effects on homeostasis of endocrine axis leading to neurological,developmental,immunological and reproductive disarray at organismal level.A wide range of structurally diverse EDs such as,sex-steroid hormone mimics,pesticides and fertilizers,prevail in the environment originating from waste of industries,pharmaceutics,sewage treatment plants and agriculture.In addition,some metals,such as Cu,Hg and Zn,have endocrine disrupting potency in their metallic as well as synthesized nano-particulate forms.There is an increasing concern in research for the plausible threat posed by EDs that can disrupt the endocrine system in aquatic fauna as these compounds are frequently discharged or run-off into water stream.Fishes are well known bio-indicators to understand toxicity of EDs as they are vulnerable to endocrine disruption.Furthermore,EDs have the potential to affect fish-feeding higher vertebrates including mammals and subsequently human,as they make their way up on the food web pyramid due to biomagnification.In light of this,several observations suggesting adverse effects of EDs and the mechanism contributing to endocrine disruption in fish are discussed extensively in this review.This article highlights the necessity to choose a credible model for assessing the toxic effects exerted by EDs.Furthermore,the toxic effects of EDs will be comprehensively reviewed with reference to sexual plasticity,neuroendocrine mechanisms,thyroid and immune modulation,gonadal development and maturation as well as changes in transcriptome/genome profile using fish models to imply ED-induced aquatic pollution in a larger perspective.For decades now,studies on EDs have challenged traditional concepts in toxicology to develop new molecular markers to improve methodologies and to assess the ecological risks associated with field conditions.In this regard,it is imperative to highlight the development of modern diagnostic tools including biosensors to monitor the inadvertent usage of EDs and the resultant environmental risks.Lastly,current limitations in knowledge along with future research perspectives in the field are also highlighted in this article.展开更多
Among vertebrates,teleosts display a wide array of sex determination and differentiation mechanisms ranging from chromosomal sex determination on one end of the spectrum to environmental sex determination on the other...Among vertebrates,teleosts display a wide array of sex determination and differentiation mechanisms ranging from chromosomal sex determination on one end of the spectrum to environmental sex determination on the other end.However,the interplay of both these mechanisms is also not uncommon.Several gonochoristic fishes exhibit gonadal plasticity often resulting in sex reversal.The major manipulation of sex differentiation in teleost is affected by sex steroids.In this context,the increasing contamination of aquatic ecosystems by estrogen-like compounds,commonly known as xenoestrogens,is of major concern.This often leads to deleterious effects on the reproductive success of fish and thereby adversely impacts aquatic biodiversity.In the present review,we have focused on impact of xenoestrogen at different levels of the reproductive system influencing not only gonadal differentiation in teleosts but also their reproductive functions.The review would also explore the mitigation strategies and regulations in place for aquatic xenoestrogen management.展开更多
It is well known that proteins existing in the biomembrane usually appear to be dispersed and disordered. However, there are a few exceptions to proteins appearing to be ordered in the membrane. For example, mammalian...It is well known that proteins existing in the biomembrane usually appear to be dispersed and disordered. However, there are a few exceptions to proteins appearing to be ordered in the membrane. For example, mammalian spermatozoa are highly polarized cells. There is an acrosome in the front of the sperm head. Therefore, the head of sperm can be often divided into diverse structural domains. The membrane is differentiated展开更多
The aim of the present paper was to check for the presence of cerebrovascular dystroglycan in vertebrates,because dystroglycan,which is localized in the vascular astroglial end-feet,has a pivotal function in glio-vasc...The aim of the present paper was to check for the presence of cerebrovascular dystroglycan in vertebrates,because dystroglycan,which is localized in the vascular astroglial end-feet,has a pivotal function in glio-vascular connections.In mammalian brains,the immunoreactivity ofβ-dystroglycan subunit delineates the vessels.The results of the present study demonstrate similar patterns in other vertebrates,except for anurans and the teleost groups Ostariophysi and Euteleostei.In this study,we investigated 1 or 2 representative species of the main groups of Chondrichthyes,teleost and non-teleost ray-finned fishes,urodeles,anurans,and reptiles.We also investigated 5 mammalian and 3 bird species.Animals were obtained from breeders or fishermen.The presence ofβ-dystroglycan was investigated immunohistochemically in free-floating sections.Pre-embedding electron microscopical immunohistochemistry on Heterodontus japonicus shark brains demonstrated that in Elasmobranchii,β-dystroglycan is also localized in the perivascular glial end-feet despite the different construction of their blood-brain barrier.The results indicated that the cerebrovascularβ-dystroglycan immunoreactivity disappeared separately in anurans,and in teleosts,in the latter group before its division to Ostariophysi and Euteleostei.Immunohistochemistry in muscles and western blots from brain homogenates,however,detected the presence ofβ-dystroglycan,even in anurans and all teleosts.A possible explanation is that in the glial end-feet,β-dystroglycan is masked in these animals,or disappeared during adaptation to the freshwater habitat.展开更多
The stress axis in teleost fish attempts to maintain internal homeostasis in the face of allostatic loading.However,stress axis induction has been associated with a higher predation rate in fish.To date,the physiologi...The stress axis in teleost fish attempts to maintain internal homeostasis in the face of allostatic loading.However,stress axis induction has been associated with a higher predation rate in fish.To date,the physiological and behavioral factors associated with this outcome are poorly understood.The purpose of the present study was to investigate the impact of experimental cortisol elevation on anti-predator behavior and physiological responses to predator presence.We hypothesized that semi-chronic cortisol elevation would increase susceptibility to predation by increasing stress-induced risk-taking behaviors.To test this hypothesis,schoolmaster snapper were given cocoa butter implants without cortisol(sham)or with cortisol(50 mg/kg body weight)and tethered to cover.Fish were exposed to either a lemon shark or control conditions for 15-min.Space use and activity were recorded throughout and fish were terminally sampled for blood.Cortisol implantation,relative to shams,resulted in higher blood glucose and plasma cortisol concentrations with a lower plasma lactate concentration.Shark exposure,relative to controls,elicited higher blood glucose and lactate concentrations but had no effect on plasma cortisol concentration.No interactions were detected between shark exposure and cortisol treatment for any physiological trait.Behavioral metrics,including shelter use and activity,were unaffected by either cortisol implantation or shark exposure.Physiological responses to cortisol implantation likely resulted from enhanced gluconeogenic activity,whereas alterations under predator exposure may have been the product of catecholamine mobilization.Further work should address context-specific influences of stress in mediating behavioral responses to predation.展开更多
Tudor domain-containing(TDRD)proteins,the germline enriched protein family,play essential roles in the process of gametogenesis and genome stability through their interaction with the PIWI-interacting RNA(piRNA)pathwa...Tudor domain-containing(TDRD)proteins,the germline enriched protein family,play essential roles in the process of gametogenesis and genome stability through their interaction with the PIWI-interacting RNA(piRNA)pathway.Several studies have suggested the rapid evolution of the piRNA pathway in teleost lineages with striking reproductive diversity.However,there is still limited information about the function and evolution of Tdrd genes in teleost species.In this study,through genome wide screening,13 Tdrd family genes were identified in economically important aquaculture fish,including spotted sea bass(Lateolabrax maculatus),Asian sea bass(Lates calcarifer),and tongue sole(Cynoglossus semilaevis).With copy number,structure,phylogeny,and synteny analysis,duplication of Tdrd6 and Tdrd7,as well as loss of Stk31 and Tdrd10,were characterized in teleost lineages.Codon based molecular evolution analysis indicated faster evolution of teleost Tdrd genes than that in mammals,potentially associated with the accelerated evolution of the piRNA pathway in teleost lineages.The evolutionary diversity of Tdrd genes was also detected between different teleost lineages.RNA-seq analysis showed that most teleost Tdrd genes were dominantly expressed in gonads,particularly highly expressed in testis,such as Tdrd6,Tdrd7a,Tdrd9,Ecat8,and Tdrd15.The varied expression and evolutionary pattern between the duplicated Tdrd6 and Tdrd7 in teleosts may indicate their functional diversification.All these results suggest a conserved function of teleost Tdrd family in gametogenesis and the piRNA pathway,which could lay a foundation for the evolution of Tdrd genes and be helpful for further deciphering of Tdrd functions in teleosts.展开更多
Horizontal gene transfer(HGT)is a common occurrence across all domains of life.However,most HGT events were reported between single-celled organisms or parasites and hosts(Van Etten and Bhattacharya 2020).A type II an...Horizontal gene transfer(HGT)is a common occurrence across all domains of life.However,most HGT events were reported between single-celled organisms or parasites and hosts(Van Etten and Bhattacharya 2020).A type II antifreeze protein(AFP)gene was the first and sole evidence of HGT direct vertebrate-to-vertebrate DNA transmission.AFP is only found in 3 widely separated branches of teleost fishes(herring,sea raven,and smelts),sharing amino acid similarity up to 80%(Graham et al.2008).展开更多
基金the National Natural Science Foundation of China.No.39070666.
文摘AIM To study the cell types,localization,distribution density and morphology of APUDcells in the intestinal mucosa of stomachlessteleost fishes.METHOD By using the peroxidase-antiperoxidase complex(PAP)immunocytochemical staining technique theidentification,localization and morphology ofimmunoreactive(IR)endocrine cells seattered inthe intestinal mucosa of grass carp(Cyenopharyngodon idellus),black carp(Mylopharyngodon piceus)and common carp(Cyprinus carpio)were investigated with 20kinds of antisera prepared against mammalianpeptide hormones of APUD cells,and likewise byusing avidin-biotin-peroxidase complex(ABC)method those of silver carp(Hypophthalmichthys molitrix),bighead(Aristichthys nobilis),silver crucian carp(Carassius gibelio)and bluntnose black bream(Megalobrama amblyocephala)were alsostudied with 5 different antisera.Thereplacement of the first antiserum by phosphatebuffered saline(PBS)was employed as a control.IR endocrine cells were counted with asquare-mesh ocular micrometer from 10 fieldsselected randomly in every section of each partof the intestine specimen.The average numberof IR endocrine cells per mm2 was counted toquantify their distribution density.RESULT Gastrin(GAS)-,Gastric inhibitorypeptide(GIP)-,glucagon(GLU)-,glucagon-likeimmunoreactants(GLI)-,bovine pancreaticpolypeptide(BPP)-,leucine-enkephalin(ENK)-and substance P(SP)-IR endocrine cells werefound in the gut of grass carp,black carp andcommon carp,and somatostatin(SOM)-IRendocrine cells were only seen in common carp.GAS-,GIP-and GLU-IR endocrine cells werefound in the intestinal mucosa of silver carp,bighead,silver crucian carp and bluntnose blackbream.Most of IR endocrine cells had the higherdistribution density in the foregut and midgut,and were longer in shape.They had a long apicalcytoplasmic process extended to the gut lumenand a basal process extended to adjacent cellsor basement membrane and touched with it.Sometimes,the basal cytoplasmic processformed an enlarged synapse-like structure in thecontiguous part with basement membrane.Thisphenomenon provided new morphologicalevidence for neuroendocrine and paracrinesecretory function of these enteroendocrinecells.CONCLUTION At least 8 kinds of IR endocrinecells were found in the gut of stomachlessteleost species for the first time in China.TheseIR endocrine cells scattering in the gut mucosabelong to the APUD system.Among them,thehormones secreted by SP-,ENK-,SOM-and GLU-IR endocrine cells belong to the peptides of dualdistribution in the brain and gut.This providednew evidence for the concept of brain-gutpeptide.According to the cell types,distribution density,morphologicalcharacteristics and variety in shape of APUDcells in the gut of stomachless teleost fishes,itis deemed that the digestive tract of fishes isalso an endocrine organ of great importance andcomplexity.
基金the National Natural Science Foundation of China,No.39470554
文摘AIM To identify the type localization andmorphology of APUD endocrine cells in thegastroenteropancreatic(GEP)system ofstomach-containing teleosts,and study APUDendocrine system in the stomach,intestine andpancreas of fish species.METHODS Two kinds of immunocytochemical(ICC)techniques of the streptavidin biotin-peroxidase complex(SABC)and streptavidin-peroxidase(S-P)method were used.Theidentification,localization and morphology ofAPUD endocrine cells scattered in the mucosa ofdigestive tract,intermuscular nerve plexus andglandular body of northern snakehead(Channaargus),ricefield eel(Monopterus albus),yellow catfish(Pelteobagrus fulvidraco),mandarinfish(Siniperca chuatsi),largemouthbass(Micropterus salmoides),orientalsheatfish(Silurus asotus),freshwater pomfret(Colossoma brachypomum)and nile tilapia(Tilapia nilotica)were investigated with 8 kindsof antisera.RESULTS The positive reaction of 5-hydroxytryptamine(5-HT)immunoreactiveendocrine(IRE)cells was found in the digestive tract and glandular body of 8 fish species indifferent degree.Only a few gastrin(GAS)-IREcells were seen in C.argus,M.albus and P.fulvidraco.Glucagon(GLU)-IRE cells were notfound in the digestive tract and glandular bodybut existed in pancreatic island of most fishspecies.The positive reaction of growthhormone(GH)-IRE cells was found only inpancreatic island of S.Chuatsi and S.Asotus,no positive reaction in the other 6 fish species.Somatostatin(SOM)-,calcitonin(CAL)-,neurofilament(NF)-and insulin(INS)-IRE cellsin the stomach,intestine and pancreas of 8 kindsof fish were different in distribution and types.The distribution of all 8 APUD cells was the mostin gastrointestinal epithelium mucosa and then indigestive glands.The positive reaction of SOM-and 5-HT-IRE cells was found in intermuscularnerve plexus of intestine of P.fulvidraco andS.chuatsi.Only GH-IRE cells were denselyscattered in the pancreatic islands of S.chuatsiand S.asotus,and odd distribution in thepancreas of S.asotus,SOM-IRE cells weredistributed in the pancreatic islands of S.asotus,C.Brachypomum and T.nilotica.There were INS-IRE cells in the pancreaticislands of S.chuatsi and S.asolus.Eightkinds of APUD cells had longer cell body andcytoplasmic process when they were located inthe gastrointestinal epithelium,and had shortercell body and cytoplasmic process in the gastricgland,and irregular shape in the esophagus andpancreatic island.CONCLUSION Eight kinds of IRE cells were identified in the GEP system of stomach-containing teleosts. These endocrine cells were scattered in gastrointestinal mucosa, intermuscular nerve plexus, gland body, pancreatic gland and islands under APUD system. CAL- and GH-IRE cells in the pancreatic islands of fishes showed functional diversity for these two hormones. Their morphological feature provides evidence of endocrine-paracrine and endocrine-exocrine acting mode. This research can morphologically prove that the GEP endocrine system of fish ( the lowest vertebrate) is almost the same as of mammal and human.
基金supported by the National Natural Science Foundation of China(31772876,41776151)Natural Science Foundation of Zhejiang Province(LZ18C190001,LR18C040001)+1 种基金Scientific Innovation Team Project of Ningbo(2015C110018)K.C.Wong Magna Fund in Ningbo University
文摘Macrophages exist in most tissues and play a variety of functions in vertebrates.Teleost fish species are found in most aquatic environments throughout the world and are quite diverse for a group of vertebrate animals.Due to whole genome duplication and en vironme ntal adaptati on,teleost monocytes/macrophages possess a variety of different functions and modulations compared with those of mammals.A deeper understanding of teleost monocytes/macrophages in the immune system will not only help develop teleost-specific methods of disease prevention but will also help improve our understanding of the various immune mechanisms in mammals.In this review,we summarize the differences in polarizati on and phagocytosis of teleost and mammalian macrophages to improve our understanding of the various immune mechanisms in vertebrates.
文摘In this current paper, the exposure time effects on four endocrine disruptors and teleost fishes were evaluated using the reduced life expectancy (RLE) model based on the effect concentration (EC<sub>50</sub>) of available literature published. The result on the regression analysis over different exposure times has demonstrated that the EC<sub>50</sub> of hepatic biomarkers falls with increasing exposure times in a predictable manner. The slopes of the regression equations reflect the strength of the toxic effects on the various teleost fish. The EC<sub>50</sub> reduction over time can be interpreted based on the bioconcentration process, which can be used to understand transfer routes of the compounds from water to fish body. RLE model also provides useful information in assessing the toxic effects on fish life expectancy as a result of the occurrence of compounds.
文摘Time-dependent tissue distribution of mercury(Hg) was studied in a freshwater perch,Anabas testudineus which revealed that the liver and kidneys are the major sites of Hg reten tion. The role of reduced glutathione (GSH) in the clearance of Hg was also investigated to e valuate the ameliorative effect of this nucleophile. For this purpose, the perch was given GSH 15 min before or after they received 203Hg by injection. The fish were then sacrificed at 24 h and 48 h later. The results clearly indicate that exogenous GSH can significantly reduce Hg retention in both the liver and kidneys, demonstrating a direct role of this nucleophile in the amelioration of Hg-induced toxicity in the early phase of intoxication
文摘The toxic effect of fertilizer Diammonium phosphate resulted in alterations of 5'-Nucleotidase activity of tissues liver, kidney and muscles offish C. batrachus at varying intervals and exposures. Alterations in 5'-Nuclcotidase activity of body organs gave an idea of the toxicity caused by the fertilizer. Thus the enzyme 5'-Nucleotidase can be used to monitor the pollution in aquatic ecosystem.
基金supported by the National Natural Science Foundation of China (32473194)Natural Science Foundation of Zhejiang Province (LY23C190002)+1 种基金Natural Science Foundation of Ningbo City (202003N4011)One Health Interdisciplinary Research Project of Ningbo University (HZ202201)。
文摘NLRP3 inflammasome activation is pivotal for cytokine secretion and pyroptosis in response to diverse stimuli,playing a crucial role in innate immunity.While extensively studied in mammals,the regulatory mechanisms governing NLRP3 activation in non-mammalian vertebrates remain largely unexplored.Teleosts,as basal vertebrates,represent an ideal model for exploring the evolutionary trajectory of inflammasome regulation.In this study,ABE assays,confocal microscopy,and biochemical analyses were applied to systematically characterize the mechanisms underlying NLRP3 inflammasome in teleosts,using large yellow croakers(Larimichthys crocea,Lc)and zebrafish(Danio rerio,Dr)as representative models.Our findings revealed a previously unrecognized palmitoylation-dependent regulatory mechanism essential for teleost NLRP3 activation.Specifically,zDHHC18-mediated palmitoylation at a teleost-specific cysteine residue(C946 in LcNLRP3,C1037 in DrNLRP3)was required for the translocation of NLRP3 to the dispersed trans-Golgi network,facilitating its subsequent recruitment to the microtubule-organizing center.This membrane trafficking was crucial for inflammasome assembly and downstream inflammatory responses.These findings provide new insights into the distinct regulatory mechanisms of NLRP3 activation in teleosts,highlighting an evolutionary divergence that contributes to innate immunity adaptation in early vertebrates.
基金supported by the National Natural Science Foundation of China(32102827,31972818,31528019)China Postdoctoral Science Foundation(2019M662959)+1 种基金Guangdong Basic and Applied Basic Research Foundation(2019A1515110987)Special fund for promoting economic development(for modern fishery development)of Guangdong Province(grant number 2019A4).
文摘The defense system of teleost fish organized on innate and adaptive immunity protects them against a wide variety of pathogenic microorganisms in the aquatic environment.Phagocytosis is one of the most effective defense strategies against microbial challenge mainly performed by classical‘professional’phagocytes(including monocytes,macrophages and granulocytes).They contain,kill and process the internalized pathogens for antigen presentation by providing antigenic ligands to initiate activation and clonal expansion of T and B cells,which bridge the innate and adaptive immunity.The discovery of phagocytic B cells in teleost fish has broken the paradigm that primary vertebrate B cells are lack of phagocytosis of particulates,as well as led to the investigation of phagocytic activity of mammalian B-1 B cells.The active phagocytic,microbicidal capabilities and antigen presentation in teleost phagocytic B cell have demonstrated to be similar as professional phagocytes,providing a potential impact on development of new vaccination strategies to prevent and control infectious diseases.In this review,we aim to address current progress on the antimicrobial role of phagocytic B cells in teleost fish by comparing it with other professional phagocytes and mammalian B-1 B cells,and provide the application prospect of phagocytic B cells in developing vaccines as well as the prevention of fish diseases.
基金supported by the National Key Research and Development Program of China(2022YFD2400502)the National Natural Science Foundation of China(42176103,41825013,42230409,42276127)+2 种基金the Guangdong Provincial Natural Science Foundation(2022A1515012505)the Key-Area Research and Development Program of Guangdong Province(2021B0202040002)China Agricultural Research System(CARS-47-G16)。
文摘The origin of T cells in the teleost's brain is unclear.While viewing the central nervous system(CNS)as immune privileged has been widely accepted,previous studies suggest that T cells residing in the thymus but not in the spleen of the teleost play an essential role in communicating with the peripheral organs.Here,we identified nine T cell subpopulations in the thymus and spleen of orange-spotted grouper(Epinephelus coioices)through single-cell RNA-sequencing analysis.After viral CNS infection with red-spotted grouper nervous necrosis virus(RGNNV),the number of slc43a2^(+)T cells synchronously increased in the spleen and brain.During the infection tests in asplenic zebrafish(tlx1^▲zebrafish model),no increase in the number of slc43a2^(+)T cells was observed in the brain.Single-cell transcriptomic analysis indicated that slc43a2^(+)T cells mature and functionally differentiate within the spleen and then migrate into the brain to trigger an immune response.This study suggests a novel route for T cell migration from the spleen to the brain during viral infection in fish.
基金supported by UK-Japan Research Collaboration Grants from the Ministry of the Environment,Japan,and JSPS KAKENHI grant No.18H02281 awarded to T.K.
文摘Gonadal soma-derived factor(Gsdf)is a member of the transforming growth factor beta(TGF-β)superfamily and a teleost-specific gene.Early studies suggested that Gsdf promotes spermatogonial proliferation;however recent reports have shed light on its role in sex differentiation based on expression profiles and gain-and loss-of-function studies.This paper discusses the involvement of Gsdf in gonadal sex differentiation and sex changes in teleosts.
基金supported by the Natural Science Foundation of Hubei Province(grant no.2021CFB265)Engineering Research Center of Feed Protein Resources on Agricultural By-products,Ministry of Education(grant no.DKXY2020015)Key Laboratory of Special Aquatic Formula Feed(grant no.TMKJZ1706 and TMKJZ1912).
文摘In mammals,chemokines are a superfamily of cytokines that regulate cell migration or exert direct antimicrobial activity.The presence and organization of four invariant cysteine residues within the mature protein sequence can classify chemokines into four major subfamilies:CXC,CC,CX_(3)C,and XC.In teleost,a novel subfamily of chemokines named CX has been identified.Since the first report of a CC chemokine(CK1)in rainbow trout in 1998,the number of identified chemokine genes in different fish species has expanded dramatically.In this paper,we reviewed the chemokine genes that have been identified so far in teleost fish and their tissue expressions and responses post-stimulation.We also discussed the functions of fish chemokines based on currently available information and compared them to human chemokines.Additionally,we predicted the structures of trout chemokines for the first time,and we found that the structures of trout chemokines share significant similarities.Finally,the adjuvant effects of fish chemokines in aquaculture were also reviewed.Overall,this review will give us a better understanding of the chemokines of teleost fish.
基金supported by the National Science Foundation for Outstanding Young Scholars(31822057)the National Key Research and Development Project(2018YFD0900503)。
文摘Viral infection induces the initiation of antiviral effectors and cytokines which are critical mediators of innate antiviral responses.The critical molecular determinants are responsible for triggering an appropriate immune response.Long noncoding RNAs(lncRNAs)have emerged as new gene modulators involved in various biological processes,while how lncRNAs operate in lower vertebrates are still unknown.Here,we discover a long noncoding RNA,termed antiviral-associated long noncoding RNA(AANCR),as a novel regulator for innate antiviral responses in teleost fish.The results indicate that fish MITA plays an essential role in host antiviral responses and inhibition of Siniperca chuatsi rhabdovirus(SCRV)production.miR-210 reduces MITA expression and suppress MITA-mediated antiviral responses,which may help viruses evade host antiviral responses.Further,AANCR functions as a competing endogenous RNA(ceRNA)for miR-210 to control protein abundance of MITA,thereby inhibiting SCRV replication and promoting antiviral responses.Our data not only shed new light on understanding the function role of lncRNA in biological processes in teleost fish,but confirmed the hypothesis that ceRNA networks exist widely in vertebrates.
基金SK and PS are grateful to the University of Hyderabad for Non-NET fellowships.NA is thankful to the Junior Research Fellowship support by a grant-in-aid(BT/PR15748/AAQ/3/803/2016)from the Department of Biotechnology(DBT)India awarded to BS.BS is also a recipient of TATA innovation fellowship(BT/HRD/35/01/02/2013)from DBT,India(during the years:2014-2019)which is acknowledged.
文摘Endocrine disruptors(EDs)are synthetic or natural chemical molecules occurring in environment that have the potential to impart adverse effects on homeostasis of endocrine axis leading to neurological,developmental,immunological and reproductive disarray at organismal level.A wide range of structurally diverse EDs such as,sex-steroid hormone mimics,pesticides and fertilizers,prevail in the environment originating from waste of industries,pharmaceutics,sewage treatment plants and agriculture.In addition,some metals,such as Cu,Hg and Zn,have endocrine disrupting potency in their metallic as well as synthesized nano-particulate forms.There is an increasing concern in research for the plausible threat posed by EDs that can disrupt the endocrine system in aquatic fauna as these compounds are frequently discharged or run-off into water stream.Fishes are well known bio-indicators to understand toxicity of EDs as they are vulnerable to endocrine disruption.Furthermore,EDs have the potential to affect fish-feeding higher vertebrates including mammals and subsequently human,as they make their way up on the food web pyramid due to biomagnification.In light of this,several observations suggesting adverse effects of EDs and the mechanism contributing to endocrine disruption in fish are discussed extensively in this review.This article highlights the necessity to choose a credible model for assessing the toxic effects exerted by EDs.Furthermore,the toxic effects of EDs will be comprehensively reviewed with reference to sexual plasticity,neuroendocrine mechanisms,thyroid and immune modulation,gonadal development and maturation as well as changes in transcriptome/genome profile using fish models to imply ED-induced aquatic pollution in a larger perspective.For decades now,studies on EDs have challenged traditional concepts in toxicology to develop new molecular markers to improve methodologies and to assess the ecological risks associated with field conditions.In this regard,it is imperative to highlight the development of modern diagnostic tools including biosensors to monitor the inadvertent usage of EDs and the resultant environmental risks.Lastly,current limitations in knowledge along with future research perspectives in the field are also highlighted in this article.
基金Minor Research Grant,Institution of Eminence(IoE/FRP/LS/2020/27)by University of Delhi.
文摘Among vertebrates,teleosts display a wide array of sex determination and differentiation mechanisms ranging from chromosomal sex determination on one end of the spectrum to environmental sex determination on the other end.However,the interplay of both these mechanisms is also not uncommon.Several gonochoristic fishes exhibit gonadal plasticity often resulting in sex reversal.The major manipulation of sex differentiation in teleost is affected by sex steroids.In this context,the increasing contamination of aquatic ecosystems by estrogen-like compounds,commonly known as xenoestrogens,is of major concern.This often leads to deleterious effects on the reproductive success of fish and thereby adversely impacts aquatic biodiversity.In the present review,we have focused on impact of xenoestrogen at different levels of the reproductive system influencing not only gonadal differentiation in teleosts but also their reproductive functions.The review would also explore the mitigation strategies and regulations in place for aquatic xenoestrogen management.
基金Project supported by the National Natural Science Foundation of China.
文摘It is well known that proteins existing in the biomembrane usually appear to be dispersed and disordered. However, there are a few exceptions to proteins appearing to be ordered in the membrane. For example, mammalian spermatozoa are highly polarized cells. There is an acrosome in the front of the sperm head. Therefore, the head of sperm can be often divided into diverse structural domains. The membrane is differentiated
文摘The aim of the present paper was to check for the presence of cerebrovascular dystroglycan in vertebrates,because dystroglycan,which is localized in the vascular astroglial end-feet,has a pivotal function in glio-vascular connections.In mammalian brains,the immunoreactivity ofβ-dystroglycan subunit delineates the vessels.The results of the present study demonstrate similar patterns in other vertebrates,except for anurans and the teleost groups Ostariophysi and Euteleostei.In this study,we investigated 1 or 2 representative species of the main groups of Chondrichthyes,teleost and non-teleost ray-finned fishes,urodeles,anurans,and reptiles.We also investigated 5 mammalian and 3 bird species.Animals were obtained from breeders or fishermen.The presence ofβ-dystroglycan was investigated immunohistochemically in free-floating sections.Pre-embedding electron microscopical immunohistochemistry on Heterodontus japonicus shark brains demonstrated that in Elasmobranchii,β-dystroglycan is also localized in the perivascular glial end-feet despite the different construction of their blood-brain barrier.The results indicated that the cerebrovascularβ-dystroglycan immunoreactivity disappeared separately in anurans,and in teleosts,in the latter group before its division to Ostariophysi and Euteleostei.Immunohistochemistry in muscles and western blots from brain homogenates,however,detected the presence ofβ-dystroglycan,even in anurans and all teleosts.A possible explanation is that in the glial end-feet,β-dystroglycan is masked in these animals,or disappeared during adaptation to the freshwater habitat.
基金M.J.Lawrence is supported by an NSERC PGS-D.S.JCooke is supported by NSERC and the Canada Research Chairs Program+3 种基金E.J.Eliason was supported by an NSERC PDFJ.W.Brownscombe is supported by NSERC and The Berkeley Marine Conservation Fellowship from The American Fisheries SocietyK.M.Gilmour is supported by NSERCJ.W.Mandelman is supported by the New England Aquarium.
文摘The stress axis in teleost fish attempts to maintain internal homeostasis in the face of allostatic loading.However,stress axis induction has been associated with a higher predation rate in fish.To date,the physiological and behavioral factors associated with this outcome are poorly understood.The purpose of the present study was to investigate the impact of experimental cortisol elevation on anti-predator behavior and physiological responses to predator presence.We hypothesized that semi-chronic cortisol elevation would increase susceptibility to predation by increasing stress-induced risk-taking behaviors.To test this hypothesis,schoolmaster snapper were given cocoa butter implants without cortisol(sham)or with cortisol(50 mg/kg body weight)and tethered to cover.Fish were exposed to either a lemon shark or control conditions for 15-min.Space use and activity were recorded throughout and fish were terminally sampled for blood.Cortisol implantation,relative to shams,resulted in higher blood glucose and plasma cortisol concentrations with a lower plasma lactate concentration.Shark exposure,relative to controls,elicited higher blood glucose and lactate concentrations but had no effect on plasma cortisol concentration.No interactions were detected between shark exposure and cortisol treatment for any physiological trait.Behavioral metrics,including shelter use and activity,were unaffected by either cortisol implantation or shark exposure.Physiological responses to cortisol implantation likely resulted from enhanced gluconeogenic activity,whereas alterations under predator exposure may have been the product of catecholamine mobilization.Further work should address context-specific influences of stress in mediating behavioral responses to predation.
基金funded by National Natural Science Foundation of China(31702331)China Agriculture Research System(CARS-47-G06)National Infrastructure of Fishery Germplasm Resources.
文摘Tudor domain-containing(TDRD)proteins,the germline enriched protein family,play essential roles in the process of gametogenesis and genome stability through their interaction with the PIWI-interacting RNA(piRNA)pathway.Several studies have suggested the rapid evolution of the piRNA pathway in teleost lineages with striking reproductive diversity.However,there is still limited information about the function and evolution of Tdrd genes in teleost species.In this study,through genome wide screening,13 Tdrd family genes were identified in economically important aquaculture fish,including spotted sea bass(Lateolabrax maculatus),Asian sea bass(Lates calcarifer),and tongue sole(Cynoglossus semilaevis).With copy number,structure,phylogeny,and synteny analysis,duplication of Tdrd6 and Tdrd7,as well as loss of Stk31 and Tdrd10,were characterized in teleost lineages.Codon based molecular evolution analysis indicated faster evolution of teleost Tdrd genes than that in mammals,potentially associated with the accelerated evolution of the piRNA pathway in teleost lineages.The evolutionary diversity of Tdrd genes was also detected between different teleost lineages.RNA-seq analysis showed that most teleost Tdrd genes were dominantly expressed in gonads,particularly highly expressed in testis,such as Tdrd6,Tdrd7a,Tdrd9,Ecat8,and Tdrd15.The varied expression and evolutionary pattern between the duplicated Tdrd6 and Tdrd7 in teleosts may indicate their functional diversification.All these results suggest a conserved function of teleost Tdrd family in gametogenesis and the piRNA pathway,which could lay a foundation for the evolution of Tdrd genes and be helpful for further deciphering of Tdrd functions in teleosts.
基金supported by the National Key Research and Development Program of China(2017YFA0604904)the Zhejiang Provincial Natural Science Foundation of China(LR21D060003)to Z.H.
文摘Horizontal gene transfer(HGT)is a common occurrence across all domains of life.However,most HGT events were reported between single-celled organisms or parasites and hosts(Van Etten and Bhattacharya 2020).A type II antifreeze protein(AFP)gene was the first and sole evidence of HGT direct vertebrate-to-vertebrate DNA transmission.AFP is only found in 3 widely separated branches of teleost fishes(herring,sea raven,and smelts),sharing amino acid similarity up to 80%(Graham et al.2008).
