Sphaerospora epinepheli n. sp. (Myxosporea: Sphaerosporidae) observed in grouper (Epinephelus malabaricus)

Sphaerospora epinepheli n. sp. is described from grouper, Epinephelus malabaricus, in cage-cultured and wild fish collected from both coastal lines of southern Thailand. Subspherical to spherical spores and mono- or disporous pseudoplasmodia were observed in the lumen of kidney tubules. Pseudoplasmodia were round to elongate, size range 15.6-22.9 microns (length) x 8.4-21.6 microns (width). Spores were 7.8-10.0 microns (length) x 12.3-14.5 microns (thickness), and 7.0-9.5 microns (width) with two spherical polar capsules of equal size measuring 2.9-4.4 microns in diameter and containing polar filaments with six or seven windings. Two uninucleate sporoplasms showed iodine vacuoles. Blood stages, similar to C-blood protozoans observed from freshwater fish in Europe, were found from peripheral blood smears of grouper. Ultrastructural studies of blood stages showed a similar structure to unidentified mobile protozoans from the blood of carp. Electron dense bodies were observed in the cytoplasm of the primary cell blood stages. Infected proximal-tubular epithelial cells showed highly vacuolated cytoplasm and pycnotic nuclei.     

Light and Electron Microscope Observations On Presporogonic and Sporogonic Stages of Sphaerospora Epinepheli (Myxosporea) In Grouper (Epinephelus Malabaricus)

ABSTRACT. Presporogonic (blood) stages of Sphaerospora epinepheli Supamattaya, Fischer-Scherl. Hoffmann, Boonyaratpalin, 1990 were observed in the circulating blood, sinus of kidney, glomerurar capillaries and liver arteries of grouper Epinephelus malabaricus. the earliest detectable stage was a primary cell with one secondary cell. After cell divisions, nine to 16 secondary cells were found in one primary cell. Ultrastructural examination revealed electron-dense bodies (118-145 nm) in the cytoplasm of primary cells. Sporogonic stages and spores were located in Bowman's space and in kidney tubule lumens. Electron micrographs revealed a similar pattern of spore development as described from other Sphaerospora spp. Kidneys infected with S. epinepheli showed highly vacuolated tubular epithelial cells and severely affected renal corpuscles.     

Sphaerospora ictaluri N. Sp. (Myxosporea: Sphaerosphoridae) Observed in the Kidney of Channel Catfish, Ictalurus punctatus Rafinesque

Sphaerospores were found in the kidneys of alevin channel catfish (Ictalurus punctatus) from a farm in Central California. MulticelluUr developmental stages, similar to C-blood protozoans described for Sphaerospora spp. from cyprinid fishes, were observed in circulating blood and numerous tissues. Upon a 2nd examination of the same population offish 10 days later, sporogonic stages were seen developing into mature Sphaerospores in the lumina of the kidney tubules. Sporogeoesis was asynchronous with simple unicellular stages adjacent to more complex forms with developing polar capsules and valves. Only one elliptical spore (5.6 μm in width, 6.5 μm in thickness by 5.8 μm in length) developed within the surrounding pscudoplasmodium. Thin valves surrounded two sporoplasm cells and two subspherical polar capsules (1.7 × 1.9 μm) which contained a polar filament with four to five turns. The blood stages of the Sphaerospora sp. described here are similar to the trophozoites seen in channel catfish with proliferativc gill disease (PGD). Early stages of PGD also observed in the same population of channel catfish containing developmental and sporogonic stages of this newly recognized Sphaerospora sp. may suggest a causal relationship between this new myxosporean and the gill disease.     

Light and Electron Microscopic Description of Sphaerospora dicentrarchi N. Sp. (Myxosporea: Sphaerosporidae) from Wild and Cultured Sea Bass, Dicentrarchus labrax L.

ABSTRACT. A new myxosporean, Sphaerospora dicentrarchi n. sp., was found in numerous organs of wild and cultured sea bass ( Dicentrarchus labrax L.). It is distinguished from all previously reported Sphaerospora spp. by the shape and small size of the spores, location in the host and its geographical distribution. Prevalence of infection was 100% and 83.5% in wild and cultured fish, respectively. Outstanding ultrastructural features are the presence of a binucleate sporoplasm, valvogenic spheroidal structures and capsulogenic lipid inclusions. Other data concerning ultrastructure and sporogenesis are presented.     

石斑鱼垂体腺苷酸环化酶激活多肽和生长激素释放激素基因的cDNA克隆及表达分析

垂体腺苷酸环化酶激活多肽 (PACAP)和生长激素释放激素 (GHRH)均属于血管活性肠肽家族成员 ,且两者前体基因在脊椎动物的鸟类、两栖类、鱼类中由同一基因编码 ,而哺乳动物是由两个不同基因编码。已有几例关于鱼类编码PACAP和GHRH基因克隆的报道 ,而关于重要海水养殖鱼类石斑鱼的PACAP和GHRH基因未见报道。克隆了PACAP GHRH前体cDNA序列 ,该前体有两种剪接方式 ,包括一个长序列和一个短序列 ,其中长序列编码PACAP和GHRH ,短序列缺失 10 5个碱基 ,仅编码PACAP而缺失编码GHRH的外显子区 ,同样情况在虹鳟和沟鲶中也有报道。通过半定量RT PCR方法对石斑鱼PACAP GHRH前体mRNA在胚胎发育和发育早期以及各部位的表达情况进行了分析。胚胎发育分析结果表明 ,从神经胚期开始 ,PACAP GHRH前体mRNA大量表达 ,提示该蛋白质在神经发育或神经营养方面具有重要作用。PACAP GHRH前体基因在中枢系统的表达量远高于外周组织。在鱼类的眼和鳃发现PACAP GHRH前体分布。     

斜带石斑鱼淋巴器官个体发育的组织学

本文应用连续组织切片技术和组织学观察,对出膜后1～60天的斜带石斑鱼(Epinephelus coioides)各期仔鱼、稚鱼和幼鱼的淋巴器官组织进行了研究,描述了淋巴器官的个体发育过程和组织学结构特征。研究表明：实验水温为22．0～27．8℃时,孵化后第10天出现头肾原基。头肾原基由未分化的造血干细胞组成。随着鱼体的生长,头肾原基的造血干细胞很快分化成不同类型的细胞;头肾主要由网状内皮系统支持下的淋巴造血组织构成。第11天出现脾脏原基。脾脏原基由造血细胞组成,淋巴化速度相对较慢。脾脏在整个发育过程中,红细胞和类红细胞占优势,没有红髓和白髓之分。第13天出现胸腺原基。胸腺发育速度较快,是明显的淋巴器官。胸腺主要由胸腺细胞(淋巴细胞)和上皮细胞组成,外区和内区没有明显的界限,但很容易区分。胸腺外被单层的上皮细胞层与咽腔相隔,保持浅表的位置,并且在整个发育过程中,胸腺与头肾是独立分开的。免疫器官原基出现顺序是头肾、脾脏和胸腺;而免疫器官淋巴化的顺序是胸腺,头肾和脾脏。和其它硬骨鱼类一样,斜带石斑鱼在早期发育阶段,淋巴器官的发育较迟,出现相对滞后的现象[动物学报49(6)：819～828,2003]。     

Sphaerospora ovophila N. Sp. and Myxobolus algonquinensis N. Sp. (Myxozoa, Myxosporea), Ovarian Parasites of Fish from Algonquin Park, Ontario, Canada

ABSTRACT. Sphaerospora ovophila n. sp. and Myxobolus algonquinensis n. sp., found in the ovary of the pumpkinseed ( Lepomis gibbosus ) and the golden shiner ( Noremigonus crysoleucas ) respectively from Algonquin Park, Ontario, are described using light and electron microscopy. Ovoid cysts of S. ovophila measured up to 500 pm in length. Monosporic pseudoplasmodia were ovoid or ellipsoid in shape and measured up to 12.5 pm in length. Spores were 7.2–8.4 pm long × 6.0–7.0 pm wide (in sutural plane) × 7.4–8.2 pm thick (perpendicular to sutural plane), with two subspherical polar capsules of equal size measuring 2.7–3.2 × 2.6–3.1 pm and each of which contained a polar filament with 6–7 coils. The spore had a straight sutural ridge which protruded slightly at the anterior end and contained two uninucleate sporoplasms. The spore valve had ornate folds on the posterior end. Cysts of M. algonquinensis ranged from ovoid to elongated ellipsoid in shape and measured up to 800 pm in length. Mature spores measured 13.G15.7 pm long × 10.1–12.1 pm wide × 5.0–6.9 pm thick. with two pyriform polar capsules of equal size measuring 5.1–5.5 pm × 2.5–2.9 pm, each of which contained a polar filament with 4–6 coils. The spores of M. algonquinensis had smooth valves, a straight sutural ridge and a distinct small intercapsular appendix.     

点带石斑鱼的亲鱼培育、产卵受精和胚胎发育

在水泥池人工养殖条件下,对人工诱导性逆转的“功能性雄性”、天然雄性及天然雌性点带石斑鱼(Epinephelus malabaricus)亲鱼进行强化培育,自然产卵受精。2002年繁殖季节,共获3cm以上的点带石斑鱼苗145万尾。亲鱼培育、产卵受精和胚胎发育的程序：(1)经过强化培育的亲鱼,自然产卵受精,受精率平均81％,最高97％;(2)成熟的“人工变性雄鱼”的精子头部呈球型,精子头径约3．7—6．1gm,尾长12．4～44．5μm;在水温26℃、盐度3％、pH7．9条件下,精子游动时间最长超过58min,精液中90％精子在31min停止游动,与天然雄亲鱼的精子无异,可作为生产雄亲鱼的来源;(3)在东山岛,产卵期由4月初持续到11月中旬,产卵高潮集中在4月底至5月底、9月底至10月初。在24h内,自然产卵通常从16：30持续到次日1：30;(4)产卵适宜水温为23．5—28．6℃,最适产卵水温为25．5—26．5℃;(5)水温26℃时,受精卵在盐度2．82％～2．96％,的海水中呈悬浮状态,盐度2．82％以下时下沉,盐度2．96％以上时上浮;(6)在水温24．9—27．6℃、盐度3％—3．3％、pH7．6—8．2的情况下,胚胎发育时间为21h。     

重组斜带石斑鱼生长激素及其抗血清在放射免疫测定中的应用

将斜带石斑鱼(Epinephelus coioides)生长激素成熟多肽cDNA序列克隆到质粒pRSET,与6x组氨酸等原核编码序列融合获得重组质粒pRGH6,转入大肠杆菌BL21(DE3),获得高效表达,表达量占细菌总蛋白的43％。免疫印迹证明表达产物为含斜带石斑鱼生长激素的融合蛋白,Ni^2 亲合层析柱纯化融合蛋白,以此为抗原免疫家兔制备特异性的抗血清。以纯化的重组生长激素和特异性的抗血清建立斜带石斑鱼生长激素的放射免疫测定法,该方法的灵敏度、特异性和重复性均达到测定血液生长激素的水平。研究了多巴胺的受体激动剂阿扑吗啡对静态孵育斜带石斑鱼脑垂体碎片释放生长激素的影响,结果表明,阿扑吗啡能以剂量依存方式促进斜带石斑鱼垂体释放生长激素。     

Establishment,Characterization, and Virus Susceptibility of a New Marine Cell Line from Red Spotted Grouper (<Emphasis Type="Italic">Epinephelus akaara</Emphasis>)

A marine fish cell line from the snout of red spotted grouper Epinephelus akaara, a protogynous hermaphrodite, was established, characterized, and subcultured with more than 60 passages. The grouper snout cell line (GSC) cells multiplied well in Dulbecco’s modified Eagle’s medium (DMEM) medium supplemented with 10% fetal bovine serum. The optimal growth temperature was 25°C, and morphologically the cells were fibroblastic. Chromosome analysis revealed that the GSC cell line has a normal diploid karyotype with . A virus titration study indicated that the cells were susceptible to turbot Scophthalmus Maximus rhabdovirus (SMRV) (10^8.5 TCID₅₀ ml⁻¹), while the viral titer of frog Rana grylio virus 9807 (RGV₉₈₀₇) reached 10^3.5 TCID₅₀ ml⁻¹. The infection was confirmed by cytopathic effect (CPE), immunofluorescence, and electron microscopy experiments, which detected the viral particles in the cytoplasm of virus-infected cells, respectively. Further, significant fluorescent signals were observed when the GSC cells were transfected with pEGFP vector DNA, indicating their potential utility for transgenic and genetic manipulation studies.     

Molecular cloning and expression analysis of interferon-gamma-inducible-lysosomal thiol reductase gene in orange-spotted grouper, Epinephelus coioides

In mammals, interferon-gamma-inducible-lysosomal thiol reductase (GILT) has been demonstrated to play a key role in the processing and presentation of MHC class II-restricted antigen (Ag) by catalyzing disulfide bond reduction, thus unfolding native protein Ag and facilitating subsequent cleavage by proteases. In this study a cDNA containing the orange-spotted grouper GILT (OsgGILT) coding sequence has been cloned and its complete sequence determined. The full-length cDNA of OsgGILT gene is 1066 bp nucleotides (nt) encoding a protein of 260 amino acids (aa), with a putative molecular weight of 28.7 kDa. The deduced OsgGILT possesses the typical structural feature of mammalian GILT, including an active-site CXXC motif, a GILT signature sequence CQHGX(2)ECX(2)NX(4)C, and 10 conserved cysteines. The result of real-time PCR showed that OsgGILT mRNA was expressed in heart, liver, brain, gill, kidney and muscle and more highly expressed in spleen. The OsgGILT expression is obviously up-regulated in spleen and kidney after induction with LPS, these results suggest that OsgGILT may be involved in the immune response to LPS challenge in orange-spotted grouper.     

Occurrence of dusky grouper Epinephelus marginatus (Lowe, 1834) in gulfs of northern Patagonia, Argentina

The occurrence of the dusky grouper Epinephelus marginatus in northern Patagonian Gulfs San Matías and Nuevo, Argentina, is recorded. This finding represents the southernmost record of the species for the southern hemisphere.     

Ceratomyxa spp. (Protozoa: Myxosporea) infections in wild and cultured sea bass, Dicentrarchus labrax, from the Spanish Mediterranean area

Ceratomyxa labracis and Ceratomyxa diplodae , parasitic in the gall bladder of wild and cultured Mediterranean sea bass ( Dicentrarchus labrax L.) were studied. Prevalences were clearly higher in cultured fish, especially for C. labracis . Starvation seems to favour the appearance of ceratomyxosis. Infection varied seasonally, with maxima in winter and minima in summer, a pattern almost contrary to temperature. Prevalence of C. labracis decreased in older fish. No clear influence of host sex was observed, and the apparent negative association between the two Ceratomyxa spp. was not confirmed statistically. Different degrees of histopathological damage of the gall bladder were observed by light and electron microscopy, mainly consisting of vacuolation, deformation or even necrosis of epithelial cells, and thickening and inflammation of the subepithelial connective tissue. Trophozoites appeared frequently lining the epithelium, closely attached to the cell surface or even forming invaginations in it. The importance of ceratomyxosis for sea bass and the influence of culture and stress conditions are discussed.     

Zschokkella mugilis N. Sp. (Myxosporea: Bivalvulida) from Mullets (Teleostei: Mugilidae) of Mediterranean Waters: Light and Electron Microscopic Description

A new Myxosporea, Zschokkella mugilis n. sp., was found in the gall bladder of several mugilids. It is distinguished from all previously reported Zschokkella from mugilids by the absence of valve ornamentation and from other species by the size of the spores. Prevalence of infection was 62.5% for Mugil capito, 70% for M. cephalus and 64.3% for Liza saliens. Plasmodia with rhizoids attached to epithelial cells appeared to compromise the host as evidenced in light and transmission electron micrographs presented herein. General ultrastructure and disporous sporogenesis of Z. mugilis resembled other Myxosporeans, but sporogonic cells did not seem to be formed in pansporoblasts.     

Ontogenetic allometric coefficient changes: implications of diet shift and morphometric traits in Hoplias malabaricus (Bloch) (Characiforme, Erythrinidae)

This study evaluated the relationship between body size and digestive tract characteristics of the important predatory freshwater fish Hoplias malabaricus , which is widely distributed in South America. The allometric coefficients were calculated for the mass and standard length ( L _S) relationships for two different L _S groups: (1) between 20 and 100 mm (characterized as insectivores) and (2) >100 mm (characterized as piscivores). Differential growth measured from the allometric coefficient, b , between the insectivore ( b < 3) and the piscivore ( b > 3) groups was detected. Anterior intestine length and pyloric caeca zone length showed significant differences between groups. Two complementary hypotheses were developed to explain the differential growth: (1) H. malabaricus has a digestive tract adapted to a piscivorous diet, which is independent of its ontogenetic stage of development, and (2) the negative allometry observed in group 1 individuals agrees with a general behavioural strategy, allowing individuals to grow in L _S during a shorter period of time.     

Isospora Normanlevinei N. Sp. and Isospora Coluzzii N. Sp. (Apicomplexa: Eimeriidae) In Emberiza Cirlus (Cirl Bunting) (Passeriformes-Emberizidae)

ABSTRACT. The morphological characteristics of two new species of Isospora observed in Emberiza cirlus (Cirl Bunting) from Italy are reported. the oocysts of Isospora normanlevinei n. sp. are spherical or sub-spherical, with a smooth double-layered wall, and measure 24.2 times 23.7 (21.0-26.5 times 21.5-25.5) μm; each oocyst contains 2 to 10 polar granules. No micropyle or residuum was observed. the piriform sporocysts measure 19.4 times 11.2 (17.0-21.0 times 10.0-12.5) μm and contain a dispersed residuum. the Stieda body is flat; the substiedal body, with scattered clear and dark granules, may be either symmetrical or asymmetrical. the oocysts of I. coluzzii n. sp. are asymmetrical and rounded shape and measure 28.6 times 24.2 (25.0-31.5 times 21.5-26.0) μm. the oocyst has a double-layered wall and 2 to 3 polar granules. Neither micropyle nor residuum is present. the sub-ellipsoidal sporocyst, measuring 18.2 times 10.0(16.5-20.0 times 9.0-11.0) μm, has a dispersed sporocyst residuum. the Stieda complex is symmetrical.     

Effects of temperature on prey consumption and growth in mass of juvenile trahira Hoplias aff. malabaricus (Bloch, 1794)

The influence of temperature on prey consumption and growth in mass of juvenile trahira Hoplias aff. malabaricus were investigated. Consumption of small-sized lambari Astyanax altiparanae (mean standard length, L _S, 5·43 cm) varied from zero to 65 over a period of 30 days. Temperatures ranged from 14 to 34° C and the size of trahiras ranged from 17·5 to 24·7 cm L _S. Prey consumption differed significantly among temperatures. Trahiras at 18° C consumed significantly less than those at 30° C. A linear multiple regression model including temperature, prey consumption and L _S explained 89·4% of the variability in growth in mass. Some caution is suggested when inferring the impact of H. aff. malabaricus piscivory on assemblage structures in systems that, despite their location in tropical regions, are subjected to seasonal thermal variations.     

Molecular and Morphometric Characteristics of Ceratomyxa hamour n. sp. (Myxosporea: Bivalvulida) Infecting the Gallbladder of the Orange‐spotted Grouper Epinephelus coioides from the Arabian Gulf,Saudi Arabia

Ceratomyxa hamour n. sp. was found to infect the gallbladder of the orange‐spotted grouper, Epinephelus coioides located off the Saudi Arabian coast of the Arabian Gulf. The infection was reported as a free‐floating spore in the bile, and pseudoplasmodia were not observed. Mature spores were crescent‐shaped and measured on average 7 μm in length and 16 μm in thickness. The polar capsule, meanwhile, had length to width measurements of 4 μm and 3 μm on average. A periodical survey was conducted throughout a sampling period between December 2012 and December 2013, with the results showing that the parasite was present throughout the year with a mean prevalence of 32.6%. The objective of this study was to characterize this new species based on its morphological and molecular differences from previously described species. Molecular analysis based on the partial sequence of the SSU rDNA gene, showed the highest similarity (97.8%) to Ceratomyxa buri, reported in the cultured yellow tail Seriola quinqueradiata in Japan. Indeed, C. buri and the new species described here formed an individual cluster with a high degree of bootstrap support. This is the first reported species of genus Ceratomyxa from the Arabian Gulf fishes off Saudi Arabia.     

Ascogregarina saraviae N. Sp. (Apicomplexa: Lecudinidae) in Lutzomyia lichyi (Diptera: Psychodidae)

Ascogregarina saraviae n. sp. (Apicomplexa: Lecudinidae) is described from wild-caught Lutzomyia lichyi (Diptera: Psychodidae) females. Gametocysts adhered to the hemocoel side of the genital accessory gland walls and oocysts were injected into their lumina. Sporulated oocysts were ellipsoidal, 12.4 × 5.8 (11.6–13.1 × 5.6–5.9) micrometers, contained eight sporozoites and a refractile residuum. The elongate form of A. saraviae n. sp. oocysts, and their more delicate walls, clearly distinguish them from oocysts of A. chagasi (Adler & Mayrink, 1961).