Involvement of host recognition by oncomiracidia and post-larval survivability in the host specificity of Heterobothrium okamotoi (Monogenea: Diclidophoridae)

Heterobothrium okamotoi, a monogenean gill parasite, shows high host specificity for tiger puffer, Takifugu rubripes. In the present study, in vivo and in vitro experimental infections were conducted using various fish species, including T. rubripes, to understand the mechanisms of specificity. In in vivo experiments, T. rubripes, grass puffer, Takifugu niphobles, olive flounder, Paralichthys olivaceus, and red sea bream, Pagrus major, were exposed to oncomiracidia of H. okamotoi labelled with a fluorescent dye, 5- (and -6) carboxyfluorescein diacetate succinimidyl ester, and the numbers of parasites on the gills and skin were recorded at intervals. Oncomiracidia were attached to gills and skin of all the experimental fish species immediately after exposure, and the infection intensity on T. rubripes was higher than that on T. niphobles and much higher than those on the other two species. After 2 days, the attached parasites remained on the gills of T. rubripes, but disappeared from the other hosts. During in vitro experiments, gill filaments excised from seven different fish species (four fish species used in the in vivo experiments and panther puffer, Takifugu pardalis, southern flounder, Paralichthys lethostigma and spotted halibut, Verasper variegates) were exposed to oncomiracidia and the attachment to each fish species and subsequent larval behaviour was observed. The percentage of post-larvae that attached to T. rubripes was slightly higher than those which attached to congeneric fish species and much higher than those of non-tetraodontid fish species. In vivo and in vitro experiments demonstrated that oncomiracidia of H. okamotoi have an affinity for their natural host, T. rubripes, and congeneric fish species. The disappearance of attached post-larvae from 'alien' hosts within 2 days during in vivo experiments suggested that host recognition by oncomiracidia and subsequent post-larval survivability are involved in the host specificity of H. okamotoi.     

Gonadal Development and Fertility of Triploid Grass Puffer Takifugu niphobles Induced by Cold Shock Treatment

Tiger puffer Takifugu rubripes is one of the most valuable fish species in Japan; however, there has not been much progress in their selective breeding until recently despite their potential in aquaculture. Their long generation time and the large body size of their broodstock make breeding difficult. Recently, we made a surrogate broodstock, which produced gametes of different species in salmonids. Therefore, by using closely related recipients, which have small body sizes and short generation times, it is possible to accelerate breeding of the tiger puffer. Thus, we considered the grass puffer Takifugu niphobles, which has a short generation time and a small maturation size, as a potential recipient for gamete production of the tiger puffer. Furthermore, if sterile triploid individuals are used as recipients, the resulting surrogate broodstock would produce only donor-derived gametes. Therefore, we examined conditions for inducing triploidy by suppressing meiosis II to retain the second polar body in grass puffer. We found that cold shock treatment, which is 5°C for 30 min starting from 5 min after fertilization, is optimal to obtain high triploidization and hatching rates. Although the resulting triploid grass puffers produced small amounts of gametes in both sexes, the offspring derived from the gametes could not live for over 3 days. Furthermore, we found that triploid grass puffer showed normal plasma sex steroid levels compared with diploids. These are important characteristics of triploid grass puffer as surrogate recipients used for germ cell transplantation.     

Production of Tiger Puffer <Emphasis Type="Italic">Takifugu rubripes</Emphasis> Offspring from Triploid Grass Puffer <Emphasis Type="Italic">Takifugu niphobles</Emphasis> Parents

The tiger puffer Takifugu rubripes is one of the most popular aquacultural fish; however, there are two major obstacles to selective breeding. First, they have a long generation time of 2 or 3 years until maturation. Second, the parental tiger puffer has a body size (2–5 kg) much larger than average market size (0.6–1.0 kg). The grass puffer Takifugu niphobles is closely related to the tiger puffer and matures in half the time. Furthermore, grass puffer can be reared in small areas since their maturation weight is about 1/150 that of mature tiger puffer. Therefore, to overcome the obstacles of maturation size and generation time of tiger puffer, we generated surrogate grass puffer that can produce tiger puffer gametes through germ cell transplantation. Approximately 5000 tiger puffer testicular cells were transplanted into the peritoneal cavity of triploid grass puffer larvae at 1 day post hatching. When the recipient fish matured, both males and females produced donor-derived gametes. Through their insemination, we successfully produced donor-derived tiger puffer offspring presenting the same body surface dot pattern, number of dorsal fin rays, and DNA fingerprint as those of the donor tiger puffer, suggesting that the recipient grass puffer produced functional eggs and sperm derived from the donor tiger puffer. Although fine tunings are still needed to improve efficiencies, surrogate grass puffer are expected to accelerate the breeding process of tiger puffer because of their short generation time and small body size.     

Cytogenetic and molecular analysis of the pufferfish Tetraodon fluviatilis (Osteichthyes)

In view of their compact genome, pufferfish (Tetraodontiformes) have been proposed as model animal for the study of the vertebrate genome. Despite such interest, cytogenetic information about puffers is still scanty. To fill this gap, a cytogenetic analysis of T. fluviatilis has been performed using both classical and molecular techniques. C-banding, followed by DAPI staining, evidenced that in T. fluviatilis, like all other puffer species so far examined, heterochromatin is essentially AT-rich and it is located at centromeres, whereas staining with CMA₃, silver staining and FISH with a 28S ribosomal RNA gene DNA probe showed 2–4 nucleolar organizing regions (NORs) located in heterochromatic regions in the considered puffer species. FISH with the 5S probe put in evidence both in T. fluviatilis and in T. nigroviridis only a 5S cluster per haploid genome that is physically unlinked with the major ribosomal RNA genes including the 28S rRNA genes. Hybridization with the (TTAGGG)_n probe showed in all the puffers brightly fluorescent signals uniform both in size and intensity at the end of all the chromosomes. Finally, mariner-like elements (MLEs) have been identified in T. fluviatilis and they have located into the NOR-associated heterochromatin.     

Developmental study of the shortened spinal cord in the adult tiger puffer fish, Takifugu rubripes (Teleostei)

ABSTRACT The spinal cords of vertebrates are generally divided into the cord proper and the minute filum terminale. While the spinal cord extends the entire length of the vertebral canal in the adult tiger puffer, Takifugu rubripes, the cord proper is greatly reduced in length and almost all of the canal is occupied by the filum terminale, which is tape-like rather than thread-like. The dorsal and ventral roots of the spinal nerves extend, respectively, above and below the filum terminale; as a whole, these form a massive cauda equina. Supramedullary cells are found in the rostral half of the medulla oblongata caudal to the cerebellum. In 4-mm long tiger puffers, the spinal cord is cylindrical and supramedullary cells are found in the rostral half of the cord. In 7-mm puffers, the longitudinally arranged ventral roots appear ventrally in the middle portion of the spinal cord. In 15-mm puffers, the dorsal and ventral roots run longitudinally along the spinal cord and have noticeably increased in number. Supramedullary cells are located in the rostral 15% of the cord. In 21-mm puffers, the spinal cord in large part becomes dorsoventrally flattened. In 30-mm puffers, the spinal cord becomes much flatter, and supramedullary cells now are located mainly in the medulla oblongata. These observations indicate that formation of the shortened spinal cord proper is due to at least two developmental processes. First, the elongation of the spinal cord proper is remarkably less than that of the vertebral canal. Second, the bulk of the spinal cord proper is translocated to the cranial cavity, where it is transformed into part of the medulla oblongata.     

不同体质量和饥饿程度对红鳍东方鲀幼鱼游泳能力的影响

为了考察不同体质量和饥饿程度红鳍东方鲀(Takifugu rubripes)幼鱼的游泳能力,利用行为生态学方法测定了不同体质量(0.22—3.31 g)和饥饿天数(1d、3d、6d和9d)下红鳍东方鲀幼鱼的绝对临界游速(U_crit,cm/s)、相对临界游速(U_crit’,体长/s,BL/s)、偏好游速(U_pref,cm/s)、6个流速区域(2—36 cm/s)下停留时间百分比(P_t,%)、偏好区域平均流速(V_mean,cm/s)和总游泳距离(D_1h,m)等游泳行为指标。结果显示,体质量和饥饿显著影响红鳍东方鲀幼鱼的U_crit、U_crit’、P_t、V_mean和D_1h。随体质量增加,实验鱼的U_crit、U_pref、V_mean和D_1h均逐渐升高,而U_crit...     

High permissivity of the fish cell line SSN-1 for piscine nodaviruses.

Seventeen isolates of piscine nodavirus from larvae or juveniles of 13 marine fish species affected with viral nervous necrosis (VNN) were examined for their infectivity to a fish cell line SSN-1. Based on cytopathic effects (CPE) and virus antigen detection by fluorescent antibody technique (FAT) after incubation at 25 degrees C, the infectivity of these virus isolates was divided into 4 groups. Group 1, including 9 virus isolates from 4 species of grouper, 2 species of sea bass, barramundi, rock porgy, and Japanese flounder showed CPE characterized by rounded, granular cells with heavy cytoplasmic vacuoles within 3 d post-incubation (p.i.), and the monolayer partially or completely disintegrated over 3 to 6 d p.i. Scattered FAT-positive cells appeared at 3 h p.i. and spread through the cell sheet with an increasing fluorescence signal over 24 h p.i. Group 2, consisting of 3 virus isolates from striped jack, induced CPE with thin or rounded, granular, refractile cells without conspicuous vacuole formation, and extensive FAT-positive reaction was observed in a time course similar to that of Group 1. Cells inoculated with Group 3 (1 isolate from tiger puffer) developed no distinct CPE but viral infection was evidenced by localized FAT-positive cells. There were no FAT-positive cells in Group 4, which included 4 isolates from Japanese flounder, Pacific cod and Atlantic halibut. However, when incubation was performed at 20 degrees C, the SSN-1 cells inoculated with the Group 3 isolate showed CPE similar to that of Group 1 and extensive FAT-positive reaction. Evidence of virus proliferation at 20 degrees C was also obtained in Group 4 isolates. The virus titers in the infected fish varied from 10(11) to 10(16) tissue culture infectious dose (TCID50) g(-1) of fish. There is a good correlation between these infectivities to the SSN-1 cells and the coat protein gene genotypes of the isolates. The present results indicate that SSN-1 cells are useful for propagating and differentiating genotypic variants of piscine nodavirus.     

Impacts of diclidophorid monogenean infections on fisheries in Japan

Many monogeneans are pathogenic to economically important fish in Japan. However no other monogenean is comparable with the diclidophorids, Heterobothrium okamotoi and Neoheterobothrium hirame, on the scale of impacts they inflict on Japanese fisheries. The shared importance of the two monogenean infections lies in their pathogenicity, fecundity and tolerance to chemical treatment. Heterobothrium okamotoi infects the gills and wall of the branchial cavity of the tiger puffer, Takifugu rubripes (Tetraodontidae), which is widely cultured in western Japan. The main presenting signs of infected fish are anaemia and extensive necrosis caused by adult worms. This monogenean deposits long strings of eggs, which reach lengths of almost 3m. Egg entanglement with the mesh of culture nets increases the chance of hatched larvae encountering susceptible fish. The oncomiracidium maintains infectivity for up to 4 days after hatching. Hydrogen peroxide is the only commercially available chemical able to control the infection, but can only kill immature worms on the gills. Neoheterobothrium hirame infects the gills and wall of the buccal cavity of the Japanese flounder, Paralichthys olivaceus (Paralichthyidae). Since the first known occurrence of this monogenean in 1993, the species has been recorded from almost all areas where the host is distributed. Neoheterobothrium hirame has the potential to produce 781 eggs per day at 20 degreeC. In the western Sea of Japan, wild young-of-the-year flounder became infected in early summer, followed by a sharp increase in prevalence in late summer. By late summer, juvenile flounder have nearly disappeared from the area, strongly suggesting that N. hirame is responsible for mortality of young fish. This is in good agreement with the recent decline in the local flounder population. Neoheterobothrium hirame has also been considered the causative agent of anaemia among wild Japanese flounder since the late 1990s.     

Age specific food utilization and spatial distribution of the puffer,Takifugu niphobles, over an intertidal sand flat

Synopsis Spatial distribution and feeding habits of the puffer fish,Takifugu niphobles, were studied to understand its mode of resource utilization in an intertidal sand flat at Tomioka in the western coast of Kyushu Island, Japan. The transect survey during high tides showed the presence of young puffers near the shoreline, while adults were mainly found in the offshore area of the sand flat. The gut contents analysis showed that although the young puffers foraged more frequently on organisms dwelling in the inshore area than the adults, both puffers fed mainly on the ghost shrimpCallianassa japonica inhabiting the entire sand flat. There was no apparent difference in the size of consumedC. japonica between the young and adult puffers. However, comparison of size structures of ghost shrimps in the field and in the gut contents showed that the young puffers foraged both from inshore and offshore areas of the sand flat, while the adults foraged mainly in the offshore area. This suggests that young puffers move more frequently over the sand flat with tides than adult ones. This pattern of trophic separation among size classes may reflect environmental features of the intertidal sand flat with large tidal fluctuations.     

Social organization of the spotted sharpnose puffer,Canthigaster punctatissima (Tetraodontidae)

Synopsis Tagged individuals of the Eastern Pacific sharpnose puffer, Canthigaster punctatissima, were studied at Punta Santa Inez, Baja California Sur. Territories were maintained by both sexes in this sexually dimorphic fish. Male puffers appear to have harems; one large male interacts with one to four smaller females. Puffers from different harems were not seen to interact except for occasional aggressive contact. Removal of a male puffer resulted in the overnight takeover of his harem by a neighboring male. Harem-forming behavior is discussed, as well as the possible role of predation.     

Evaluation of the Possible Transmission of BSE and Scrapie to Gilthead Sea Bream (Sparus aurata)

In transmissible spongiform encephalopathies (TSEs), a group of fatal neurodegenerative disorders affecting many species, the key event in disease pathogenesis is the accumulation of an abnormal conformational isoform (PrP^Sc) of the host-encoded cellular prion protein (PrP^C). While the precise mechanism of the PrP^C to PrP^Sc conversion is not understood, it is clear that host PrP^C expression is a prerequisite for effective infectious prion propagation. Although there have been many studies on TSEs in mammalian species, little is known about TSE pathogenesis in fish. Here we show that while gilthead sea bream (Sparus aurata) orally challenged with brain homogenates prepared either from a BSE infected cow or from scrapie infected sheep developed no clinical prion disease, the brains of TSE-fed fish sampled two years after challenge did show signs of neurodegeneration and accumulation of deposits that reacted positively with antibodies raised against sea bream PrP. The control groups, fed with brains from uninfected animals, showed no such signs. Remarkably, the deposits developed much more rapidly and extensively in fish inoculated with BSE-infected material than in the ones challenged with the scrapie-infected brain homogenate, with numerous deposits being proteinase K-resistant. These plaque-like aggregates exhibited congophilia and birefringence in polarized light, consistent with an amyloid-like component. The neurodegeneration and abnormal deposition in the brains of fish challenged with prion, especially BSE, raises concerns about the potential risk to public health. As fish aquaculture is an economically important industry providing high protein nutrition for humans and other mammalian species, the prospect of farmed fish being contaminated with infectious mammalian PrP^Sc, or of a prion disease developing in farmed fish is alarming and requires further evaluation.     

Influence of puffer predation on vertical distribution of mangrove littorinids in the Gulf of Nicoya, Costa Rica

The eastern Pacific puffer fish Sphoeroides rosenblatti (family Tetraodontidae) preys on the mangrove periwinkles Littorariafasciata and L. varia up to 13 cm above the water surface. Puffers capture snails by jumping partially out of the water and removing them by mouth from Rhizophora spp. prop roots. Field predation trials suggested puffer predation is intense, averaging 40.6% h⁻¹ on L. fasciata tethered at the water surface. L.fasciata <∼8 mm in shell length and L. varia <∼12 mm remained near or below the water surface and were easily crushed by puffers; gut content analyses showed that these snails were commonly eaten by puffers. L.fasciata >∼8 mm and L. varia∼12–16 mm were also crushed by puffers, but were generally located above their reach; these snails were much less abundant in puffer guts. L.varia >∼16 mm were abundant and some were submerged at high tide; however, none were found in puffer guts presumably because puffers encounter difficulty crushing these snails. S.rosenblatti is an important predator of L. fasciata and L. varia and appears to affect their vertical distributions. Received: 21 April 1998 / Accepted: 10 August 1998     

低盐对菊黄东方鲀幼鱼生长、存活、耗氧、鳃Na /K -ATP酶以及肝抗氧化酶的影响

通过探讨低盐(盐度为0、1.7、5、10、15和20)对菊黄东方鲀(Takifugu flavidus)幼鱼生长、存活、耗氧、鳃Na+/K+-ATP酶以及肝抗氧化酶的影响,研究了菊黄东方鲀幼鱼对低盐的适应性。结果显示,菊黄东方鲀幼鱼在盐度0组实验3周后全部死亡;盐度1.7组实验6周幼鱼大量死亡,最后成活率相当低,仅17.33%;盐度5、10、15和20组的幼鱼在整个实验中没有出现死亡现象。全长特定生长率在盐度1.7~20组之间没有显著差异。体重特定生长率,1.7盐度组比其他盐度组显著低(P0.05),5~20盐度组之间没有显著差异(P0.05)。最高的全长特定生长率和最高的体重特定生长率均出现在10盐度组。前6周的饵料系数在盐度1.7~20组之间没有显著差异,但最高(1.27)和最低(1.17)的饵料系数分别出现在1.7和10盐度组,总饵料系数在5~20盐度组之间没有显著差异。幼鱼的耗氧率在5~20盐度组之间没有显著差异,但最低的耗氧率出现在10盐度组。最低的鳃Na+/K+-ATP酶(NKA)活性出现在10盐度组,5~20盐度与NKA活性的关系可以用二次函数来拟合(y=0.083 2 x2﹣2.125 2 x+20.915,r2=0.977 9),由此得到理论上最低NKA活性值出现在盐度12.77。肝超氧化物歧化酶(SOD)和谷胱甘肽过氧化物酶(GSH-PX)活性在盐度1.7~20组之间均无显著差异,而10和15盐度组的过氧化氢酶(CAT)活性比1.7和20盐度组的显著低(P0.05),1.7~20盐度与CAT活性关系可以用二次函数来拟合(y=0.257 7 x2﹣5.807 6 x+87.357,r2=0.877 1),由此得到理论上最低CAT活性值出现在盐度11.27。研究结果表明,盐度1.7是菊黄东方鲀幼鱼的生存极限低盐,盐度5以上已经能适合其存活和生长,盐度10~15是的菊黄东方鲀幼鱼的最适宜盐度范围,适当降低盐度对菊黄东方鲀幼鱼的养殖生产是有利的。建议菊黄东方鲀幼鱼养殖盐度至少在5以上,最好在10~15范围。     

Prevalence of Toxoplasma gondii in rats (Rattus norvegicus) in Grenada, West Indies

Cats are important in the natural epidemiology of Toxoplasma gondii, because they are the only hosts that can excrete environmentally resistant oocysts. Cats are infected with T. gondii via predation on infected birds and rodents. During 2005, 238 rats (Rattus norvegicus) were trapped in Grenada, West Indies, and their sera along with tissue samples from their hearts and brains were examined for T. gondii infection. Antibodies to T. gondii were assayed by the modified agglutination test (MAT, titer 1:40 or higher); only 2 (0.8%) of 238 rats were found to be infected. Brains and hearts of all rats were bioassayed in mice. Toxoplasma gondii was isolated from the brain and the heart of only 1 rat, which had a MAT titer of 1:320. All of 5 mice inoculated with the heart tissue, and the 5 mice inoculated with the brain tissue of the infected rat remained asymptomatic, even though tissue cysts were found in their brains. Genetically, the isolates of T. gondii from the heart and the brain were identical and had genotype III by using the SAG1, SAG2, SAG3, BTUB, and GRA6 gene markers. These data indicate that rats are not important in the natural history of T. gondii in Grenada.     

鳜鱼病毒病原的检出及组织病理分析

经对患暴发性传染病的鳜组织及病变组织超微切片的电镜观察,在细胞质,核及间隙中都有病毒颗粒,对该病毒进行了分离提纯,这是一种直径约为２８０ｎｍ球形病毒,将粗提病毒液接种到培养细胞中,会引起细胞病变;病毒通过人工感染健康鳜,会引起鳜发病和死亡,对患病獗进行解剖和组织病理观察,表明病鳜的５种器官都发生了病变,研究结果表明分离的鳜病毒（ＳｉｎｉｐｅｒｃａｃｈｕａｔｓｉＶｉｒｕｓ,ＳＣＶ）是引起鳜流４行的病     

Production of reactive oxygen species (ROS) by various marine fish species during the larval stage

Previous studies have indicated that the devil stinger produces reactive oxygen species (ROS) during early development from fertilized egg to larva. To determine whether ROS generation is a common feature in marine fish species, we conducted chemiluminescence analysis using ROS specific probe (L012) on larvae of six marine fish species. Marbled rockfish, black rockfish, and devil stinger showed higher levels of chemiluminescence response (CR), whereas the levels of CR of sevenband grouper, tiger puffer, and red seabream were fairly lower. These CRs were inhibited by the addition of superoxide dismutase. Hypersensitive photon-counting microscopic observation of black rockfish suggested that ROS production was concentrated in the head area. Our results suggest that the larvae of these six marine fishes produce ROS to considerably different extents depending on species, and that rockfish species, belonging to ovoviviparous fish, tend to produce much higher levels of ROS especially at the later larval stage.     

Prevalence and Intensity of Opisthorchis viverrini Metacercarial Infection in Fish from Phnom Penh,Takeo, and Kandal Provinces,Cambodia

The prevalence and intensity of Opisthorchis viverrini metacercariae (OvMc) were investigated in fish from 3 southern administrative regions along the Mekong River in Cambodia, i.e., Phnom Penh, Takeo, and Kandal Provinces from 2017 to 2020. A total of 295 freshwater fish (24 species) were transported to our laboratory with ice and examined using the artificial digestion method. In Phnom Penh, among 4 fish species positive for OvMc, 9 (23.7%) of 38 specimens examined were infected, and their intensity of infection averaged 4.3 metacercariae per infected fish. In Takeo Province, among 10 fish species positive for OvMc, 24 (38.1%) out of 63 fish examined were infected, and their intensity of infection was av. 14.4 metacercariae per infected fish. In particular, all of 3 Osteochilus schlegelii fish examined were infected, and their infection intensity was high, 34.7 metacercariae per fish. In Kandal Province, among 6 fish species positive for OvMc, 46 (90.2%) out of 51 specimens examined were infected, and their infection intensity was 24.0 metacercaraie per infected fish. All fish of Systomus orphoides (n=17), Barbonymus altus (n=14), and Rasbora aurotaenia (n=2) were infected, and their intensity of infection averaged 37.7, 21.6, and 18.5 metacercariae per fish, respectively. Metacercariae of Haplochis yokogawai, Haplorchis taichui, and Centrocestus formosanus were detected in fish from Takeo and Kandal Provinces. From these results, it has been confirmed that a variety of fish species from Phnom Penh, Takeo, and Kandal Provinces are commonly infected with OvMc, and preventive measures to avoid human O. viverrini infection should be performed in Cambodia.     

Alphavirus-induced apoptosis in mouse brains correlates with neurovirulence.

Sindbis virus induces apoptotic cell death in cultured cell lines, raising the possibility that apoptosis of infected neurons and other target cells in vivo may contribute to the resulting disease and mortality. To investigate the role of apoptosis in Sindbis virus pathogenesis, infected mouse brains were assayed by the in situ terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling technique and for DNA ladder formation. Infection with recombinant Sindbis virus strain 633 resulted in widespread apoptosis in newborn mouse brains and spinal cords, but few apoptotic cells were observed following infection of 2-week-old animals. This finding correlates with the age-dependent mortality observed in mice. The more neurovirulent virus TE, which differs from 633 by a single amino acid in the E2 glycoprotein, induced significant apoptosis in brains and spinal cords of 2-week-old animals, consistent with its ability to cause fatal disease in older animals. Double-labeling experiments demonstrated that the apoptotic cells were also infected with Sindbis virus. Thus, Sindbis virus-induced apoptosis appears to be a result of virus infection and is likely to reflect pathogenic mechanisms for other viruses.     

两种鳜病毒的共感染现象及引起感染细胞的超微变化

借助细胞培养和电镜技术,揭示了鳜球形病毒(Siniperca chuatsi spherical virus,SCSV)与鳜弹状病毒(Siniperca chuatsi rhabdovirus,SCRV)在草鱼鳍细胞(Grass carp fins,GCF)中共感染的现象。在筛选到敏感鱼类细胞系和建立了 鳜病毒体外增殖系统的基础上,取息典型病毒感染出血症的鳜组织,制备组织悬液,接种到GCF细胞中传代培养, 在攻毒后间隔不同时间收集细胞,对攻毒细胞的超薄切片进行电镜观察。揭示两种形态的鳜病毒可在同一个GCF 细胞中增殖,并描述和分析了病毒复制引起感染细胞的超微病变。本研究结果有助于阐明鱼类重要病毒病害的发 生过程及致病机理。