Ultrastructural morphogenesis of dimorphic arcuate infection (gun) cells of Haptoglossa erumpens an obligate parasite of Bunonema nematodes

Haptoglossa is a genus of biflagellate organisms that has been placed in the oomycetes and is characterised by producing unique infective gun cells, which usually infect by physically rupturing the nematode cuticle. Haptoglossa erumpens is a parasite of Bunonema nematodes that produces arcuate infection cells and aplanospores that are discharged following the swelling and rupture of the thallus wall and distended host cuticle. Recent isolations of H. erumpens have revealed that the germinating aplanospores develop into two similar-sized but morphologically distinct infection cells. The uni-nucleate, convexly arcuate, gun cells were observed to fire in response to host nematodes, producing a cylindrical sporidium inside the host body. These gun cells had an apical missile chamber containing a needle with a unique arrangement of investing cones. Unlike previously described gun cells, the tube tail did not wind around the nucleus but continued into the basal vacuole where it terminated. The second type of infection cell was a concavely arcuate, bi-nucleate, cell that had an unusually large and elongate annulus component in the missile chamber. These modified bi-nucleate gun cells were never observed to fire in response to contact with Bunomena nematodes. The patterns of morphological and structural variations in these infection structures in this genus are reviewed in the light of these findings.     

Injection tube differentiation in gun cells of a haptoglossa species which infects nematodes

The gun cells which develop from germinating cysts in Haptoglossa produce a specialized infection apparatus, the injection tube. Upon eversion this tube fires a missile-like projectile which penetrates the host cuticle and then forms an infective sporidium within the body cavity of the nematode host. The temporal assembly of this complex cell organelle has been determined by serial-section reconstructions of maturing gun cells in a previously undescribed Haptoglossa species. The differentiation of the partially walled inverted injection tube is an unusual example of internal tube growth, in which membrane and wall assembly are temporally separated. There is no evidence that the shape of this inverted tube, which coils around the nucleus until it doubles back on itself, is dictated by the disposition of cytoplasmic microtubules. However, actin-like material was associated with the delimiting membrane of the differentiating tube, particularly in the regions of extension. From these studies it seems likely that the "head and buttress" structures previously depicted as the barbed tip of the "harpoon-like" penetration missile are part of a separate, structurally complex system which we suggest locks the "missile" into position in the invaginated injection tube. From this detailed account of cell architecture, models for the likely mechanism of infection cell firing are discussed, and unresolved questions relating to the cell biology and biochemistry of these complex organelles are highlighted. Copyright 1998 Academic Press.     

Shooting of sporidium by "gun" cells in <Emphasis Type="Italic">Haptoglossa heterospora</Emphasis> and <Emphasis Type="Italic">H. zoospora</Emphasis> and secondary zoospore formation in <Emphasis Type="Italic">H. zoospora</Emphasis>

Haptoglossa spp. (Lagenidiales, Oomycetes) have been known to parasitize microscopic animals by means of a "gun" cell that shoots an infection cell, named the sporidium, into the body of the animal. A thallus grown from the sporidium changes into a zoosporangium at maturation to produce a number of zoospores that encyst after a swarming period, and the resulting cysts germinate to produce gun cells. In Haptoglossa zoospora, endoparasitic in nematodes, the cysts of primary zoospores that swam for about 5 min did not develop gun cells but produced secondary zoospores that swam for about 3 h. After encystment of the secondary zoospores, each secondary cyst germinated to produce a gun cell. In the present study, the secondary zoospores of the genus Haptoglossa could be recorded with a videocassette recorder for the first time. The videocassette recording also revealed the infection of a nematodes by H. zoospora and H. heterospora to be composed of two steps of injection of a sporidium by the gun cell, in which the gun cell came in contact with the cuticle of a nematode and produced a spherical adhesorium on the tip of the cell in 0.07–0.1 s in both species. The adhesorium was ∼2 μm in H. zoospora and ∼4 μm in H. heterospora. When the adhesorium infiated to full size, it shot the sporidium into the nematode's body in 0.5–0.65 s and in 0.2–0.5 (or rarely 1.0) s in H. zoospora and H. heterospora, respectively. After shooting, the empty gun cell with an empty cyst case was separated from the cuticle immediately in both species. Received: October 3, 2001 / Accepted: December 13, 2001     

Shooting of sporidium by “gun” cells in Haptoglossa heterospora and H. zoospora and secondary zoospore formation in H. zoospora

Haptoglossa spp. (Lagenidiales, Oomycetes) have been known to parasitize microscopic animals by means of a “gun” cell that shoots an infection cell, named the sporidium, into the body of the animal. A thallus grown from the sporidium changes into a zoosporangium at maturation to produce a number of zoospores that encyst after a swarming period, and the resulting cysts germinate to produce gun cells. In Haptoglossa zoospora, endoparasitic in nematodes, the cysts of primary zoospores that swam for about 5min did not develop gun cells but produced secondary zoospores that swam for about 3h. After encystment of the secondary zoospores, each secondary cyst germinated to produce a gun cell. In the present study, the secondary zoospores of the genus Haptoglossa could be recorded with a videocassette recorder for the first time. The videocassette recording also revealed the infection of a nematodes by H. zoospora and H. heterospora to be composed of two steps of injection of a sporidium by the gun cell, in which the gun cell came in contact with the cuticle of a nematode and produced a spherical adhesorium on the tip of the cell in 0.07–0.1 s in both species. The adhesorium was ~2 μm in H. zoospora and ~4 μm in H. heterospora. When the adhesorium inflated to full size, it shot the sporidium into the nematode's body in 0.5–0.65 s and in 0.2–0.5 (or rarely 1.0) s in H. zoospora and H. heterospora, respectively. After shooting, the empty gun cell with an empty cyst case was separated from the cuticle immediately in both species.     

An electron-microscopical analysis of capture and initial stages of penetration of nematodes byArthrobotrys oligospora

A detailed analysis was made of the capture and subsequent penetration of nematodes by the nematophagous fungusArthrobotrys oligospora using different electron-microscopical techniques. Capture of nematodes by this fungus occurred on complex hyphal structures (traps) and was effectuated by an adhesive coating, present on these trap cells. The adhesive layer was largely fibrillar in nature and was absent on cells of normal hyphae. Following capture, penetration hyphae were formed at those sites where the trap cell wall was anchored to the nematode cuticle by the adhesive. New walls of these hyphae were formed underneath the original trap cell walls, which were partly hydrolysed to allow growth and development of the penetration tubes through the adhesive coating towards the cuticle. Our observations indicated that the cuticle of the nematode was subsequently penetrated by the penetration tubes by mechanical means. After penetration a large infection bulb was formed from which trophic hyphae arose. Cytochemical experiments indicated that the sites of penetration of the cuticle were intensely stained for acid phosphatase activity. At later stages of infection activity of this enzyme was present throughout the nematode contents; the enzyme was most probably secreted by complex membranous structures associated with the cytoplasmic membrane of the infection bulb and the trophic hyphae.     

Paternal cytoplasmic transmission in Chinese pine (Pinus tabulaeformis)

Summary. In this paper, the stages of normal sexual reproduction between pollen tube penetration of the archegonium and early embryo formation in Pinus tabulaeformis are described, emphasizing the transmission of parental cytoplasm, especially the DNA-containing organelles – plastids and mitochondria. The pollen tube growing in the nucellus contained an irregular tube nucleus followed by a pair of sperm cells. The tube cytoplasm contained abundant organelles, including starch-containing plastids and mitochondria. The two sperm cells differed in their volume of cytoplasm. The leading sperm, with more cytoplasm, contained abundant plastids and mitochondria, while the trailing one, with a thin layer of cytoplasm, had very few organelles. The mature egg cell contained a great number of mitochondria, whereas it lacked normal plastids. At fertilization, the pollen tube penetrated into the egg cell at the micropylar end and released all of its contents, including the two sperms. One of the sperm nuclei fused with the egg nucleus, whereas the other one was retained by the receptive vacuole. Very few plastids and mitochondria of male origin were observed around the fusing sperm and egg nuclei, while the retained sperm nucleus was surrounded by a large amount of male cytoplasm. The discharged tube cytoplasm occupied a large micropylar area in the egg cell. In the free nuclear proembryo, organelles of maternal and paternal origins intermingled in the neocytoplasm around the free nuclei. Most of the mitochondria had the same features as those of the egg cell, but some appeared to be from sperm cells and tube cytoplasm. Plastids were obviously of male origin, with an appearance similar to those of the sperm or tube cells. After cellularization of the proembryo, maternal mitochondria became more abundant than the paternal ones and the plastids enlarged and began to accumulate starch. The results reveal the cytological mechanism for paternal inheritance of plastids and biparental inheritance of mitochondria in Chinese pine. Correspondence and reprints: State Key Laboratory of Plant Physiology and Biochemistry, College of Biological Science, China Agricultural University, Beijing 100094, People’s Republic of China.     

Ultrastructural and Cytochemical Study on Mature Pollen of Pinus tabulaeformis

The pollen of Pinus tabulaeformis Cart. comprised two prothallial cells, a generative cell and a tube cell which degenerated at pollen maturation. The generative cell had its own cell wall, seperating from the intine of pollen, but with its side wall attached to the infine. Cytoplasmic channels were present on the side of the generative cell wall, which faced to the tube cell cytoplasm. The generative cell differed conspicuously from the tube cell. The main differences include: ( 1 ) The chromatin in the generative cell nucleus was condensed, but was dispersed and had numerous nueleare pores in the tube cell nucleus; (2)There was no microbody in the generative cell but many microbodies were present in the tube cell cytoplasm; (3)More inclusions were present in the tube cell than in the generative cell. Both the generative cell and the tube cells contained lipid bodies and amyloplasts in the cytoplasm, but there were more amyloplasts in the former. The tube cell also contained a few proteins which was absent in the generative cell. In addition, there were numerous mitochondria, polyribosomes, and a few endoplasmic reticulums and dictyosomes in the generative and tube cells. DAPI staining demonstrated numerous cytoplasmic DNA in both generative cell and tube cell. The mode of cytoplasmic inheritance, and the composition, structure and the nature of the pollen wall of P. tabulaefonnis are also discussed in this paper.     

Time-lapse photomicrography and electron microscopy on initiation of infection of nematodes by Dactylella ellipsospora

 Infection of nematodes by two strains of Datylella ellipsospora was observed using video photomicrography and electron microscopy. By light microscopy, each cell with adhesive knobs contained a number of particles that were distributed evenly before capture of a nematode. The cytoplasmic particles moved to and fro at random. At the moment when the knob cell came into contact with a nematode, the particles accumulated at the place where the cell wall of the knob stuck firmly to the nematode cuticle and exuded adhesive at the same time. The adhesive can be seen near the point of contact between the cell wall of the knob and the cuticle of the nematode. At that point, the knob cell produced an infection peg in most cases, and the cell showed a preference to invade the body of the nematode rather than the tail and head. During capture, accumulation of cytoplasmic particles was seen until infection-bulb formation began. In electron micrographs of ultrathin sections, most of the particles could be seen as electron-dense vesicles, 0.2–0.6 μm in diameter. After attachment of the knob cell to the nematode cuticle, the vesicles were found to fuse with plasmalemma one after another to exude adhesive seen as an amorphous electron-dense substance. Received: January 30, 2002 / Accepted: April 25, 2002     

The ultrastructure of the cuticle of adult female Mermis nigrescens (Nematoda)

The ultrastructure of the cuticle of the adult female nematode Mermis nigrescens has been described. There is an epicuticle and three-layered membrane covering the cuticle. The cortex is penetrated by canals which extend from the surface of the cuticle to the matrix of the layer beneath the cortex. Beneath the cortex are two layers of giant fibres which spiral around the nematode, a thick layer containing a network of fibres and a basal layer containing a vacuolated matrix material. it is thought that the epicuticle is secreted from the canals in the cortex. The possible functions of the layers in the cuticle have been discussed and similarities with the cuticle of the Acanthocephala have been noted.     

菜青虫感染蜡蚧轮枝菌后的组织病理变化

对感病菜青虫Pieris rapae组织切片的观察研究表明, 蜡蚧轮枝菌Verticillium lecanii主要通过昆虫的体壁侵染虫体。菜青虫各龄幼虫在体壁接菌12 h后,附着在虫体表面的孢子即可萌发。2～3龄幼虫,蜡蚧轮枝菌菌丝24 h就可穿透体壁进入血腔,48 h可见脂肪体等器官发生病变; 4～5龄幼虫,36 h菌丝才可穿透体壁,48 h可见虫体内有部分菌丝体。侵入虫体内的菌丝对寄生组织没有选择性。菌丝首先在入侵的血腔中生长,然后侵入脂肪体和肌肉,随菌丝在虫体内的增殖,中肠、马氏管、丝腺等相继被侵染。受侵的组织器管均发生明显的病变,如体壁分离,脂肪体变形、溶解,肌纤维排列松散,中肠上皮细胞脱落并出现许多空泡等。     

Developmental sequence of the attack apparatus ofHaptoglossa mirabilis

Summary The most prominent ultrastructural characteristics of the cyst ofHaptoglossa mirabilis are a large centrally-placed nucleus which is partially ringed by three or four parallel cisternae of rough endoplasmic reticulum (r-ER), a centriole pair and single large Golgi complex which occupy the anterior end of the cell, and a population of provacuoles which occupies the posterior. During germination these organelles migrate into a narrow germ tube which subsequently expands to form the gun cell initial. The extracellular components of the attack apparatus (i.e. missile and injection tube) are formed entirely in the developing gun cell; indirect evidence suggests that both the Golgi complex and r-ER are involved in their synthesis. The intra-cellular component of the attack apparatus comprises the posterior, anterior and apical vacuoles. The posterior vacuole forms by fusion and expansion of the original cyst provacuoles; the formation of the anterior and apical vacuoles occurs late in gun cell differentiation and involves fusion of Golgi-derived vesicles.     

Bacterial parasite of a plant nematode: morphology and ultrastructure.

The life cycle of a bacterial endoparasite of the plant-parasitic nematode Meloidogyne incognita was examined by scanning and transmission electron microscopy. The infective stage begins with the attachment of an endospore to the surface of the nematode. A germ tube then penetrates the cuticle, and mycelil colonies form in the pseudocoelom. Sporulation is initiated when terminal cells of the mycelium enlarge to form sporangia. A septum within each sporangium divides the forespore from the basal or parasporal portion of the cell. The forespore becomes enclosed by several laminar coats. The parasporal cell remains attached to the forespore and forms the parasporal microfibers. After the newly formed spores are released into the soil, these microfibers apparently enable a mature spore to attach to the nematode. These results indicate that the endoparasite is a procaryotic organism having structural features that are more common to members of Actinomycetales and to the bacterium Pasteuria ramosa than to the sporozoans or to the family Bacillaceae, as previous investigatios have concluded.     

狗枣猕猴桃果实发育的解剖学观察

对狗枣猕猴桃果实发育进行了显微和超微结构观察。其主要过程可分为：1. 早期胚胎发育：多心皮合生子房,通常具16个心室,中轴胎座。中轴上着生大量倒生胚珠,单珠被,具珠被绒毡层。胚囊蓼型。授粉后约2～4h花粉萌发。授粉后约120h花粉管到达胚珠。受精后,初生胚乳核分裂先于合子。胚乳发育为细胞型。2. 果壁：果壁可分为果皮、果肉和果心三部分。果皮较薄,由2～3层薄壁细胞组成。外表面光滑无毛,但覆有角质层,气孔下陷,分布于其中。果肉的大部分由薄壁细胞组成,分大、小两种细胞 ,小细胞含较多淀粉粒,淀粉粒的水解是果实软化的原因之一。果肉薄壁组织细胞还含有叶绿体及两种异细胞,一种异细胞内含物为结晶,普遍存在;另一种含被番红染成红色的絮团状物。果心由维管束和薄壁细胞组成,维管束不发达。3. 种子：种皮外表面呈蜂窝状,种皮较硬,由两层细胞构成。线性直立胚,胚乳发达,主要贮藏物是蛋白质。     

Muscle cell attachment in Caenorhabditis elegans

In the nematode Caenorhabditis elegans, the body wall muscles exert their force on the cuticle to generate locomotion. Interposed between the muscle cells and the cuticle are a basement membrane and a thin hypodermal cell. The latter contains bundles of filaments attached to dense plaques in the hypodermal cell membranes, which together we have called a fibrous organelle. In an effort to define the chain of molecules that anchor the muscle cells to the cuticle we have isolated five mAbs using preparations enriched in these components. Two antibodies define a 200-kD muscle antigen likely to be part of the basement membrane at the muscle/hypodermal interface. Three other antibodies probably identify elements of the fibrous organelles in the adjacent hypodermis. The mAb IFA, which reacts with mammalian intermediate filaments, also recognizes these structures. We suggest that the components recognized by these antibodies are likely to be involved in the transmission of tension from the muscle cell to the cuticle.     

An ultrastructural study of development and reproduction in the nematode parasite Myzocytiopsis vermicola

An isolate of Myzocytiopsis vermicola, a holocarpic parasite of Rhabditis nematodes, was studied with transmission electron microscopy (TEM) to follow development during infection, asexual and sexual reproduction. Nematodes became infected after attachment of apical cystospore buds to the nematode cuticle. Apical buds were packed with vesicles with dense fibrillar contents, which were absent from the thallus. Some thalli developed into sporangia while others became paired gametangial cells. Zoospore cleavage was often intrasporangial, although during the early stages of an epidemic partially differentiated zoospores usually were released via an exit tube into a fine vesicle. Packets of tripartite tubular hairs (TTH) were not observed in the cytoplasm of either developing or mature sporangia. TEM of sectioned material and whole mounts of zoospores revealed biflagellate zoospores, some without hairs and others with a proximal row of very short hairs on the anterior flagellum. Gametangial contact was via a short, walled fertilization tube and surplus antheridial and oogonial nuclei remained in their respective gametangial cells until disintegration of the periplasm. The mature oospores had a scalloped, electron opaque, epispore wall layer. These observations will be discussed in relation to the likely phylogenetic position of the Myzocytiopsidales within the oomycetes.     

Electron microscopy of infection of nematodes byDactylaria haptotyla

Infection of nematodes byDactylaria haptotyla, a nematode-trapping hyphomycete, was studied by electron microscopy. The cytoplasm of the adhesive knob in the fungus contained a number of electron-dense, membrane-bound vesicles, 0.2–0.5 µm in diam. The vesicles were rarely seen in the stalk cell or vegetative cell cytoplasm. When the adhesive knob came into contact with the nematode's cuticle, it secreted an adhesive which was seen in ultrathin sections between the knob and the cuticle as an amorphous mass. At the same time, electron-dense vesicles in the cytoplasm were reduced in number and many small vacuoles developed. Soon after capture of a nematode, the cell wall of the adhesive knob became obscure at the prospective site of penetration, where a vesicle, 0.7 µm in diam, was found in serial thin sections of the knob's cytoplasm. At the site facing the vesicle, the peripheral part of the nematode's cell exhibited a high electron density. The vesicle, which appeared to be derived from smaller electron-dense vesicles coalesced with each other, released its enzymic contents toward the captured nematodes before penetration by the fungus.     

Interaction of catenaria anguillulae with romanomermis culicivorax

The development of Catenaria anguillulae, at 27 degrees C, in live and heat-immobilized Romanomermis culicivorax was studied at the ultrastructural level. C. anguillulae zoospores infected eggs, preparasites, and postparasites. The zoospores attached predominately to the head of preparasites but in postparasites attachment was primarily on the attenuated tail region. Zoospore attachment usually occurred within 4 h but at high zoospore densities attachment occurred within 30 min. Infection hyphae penetrated the cuticle and developed sporangial swellings within the nematode tissues and pseudocoelom. Sporangia produced exit tubes within 48 h. Those regions of the J3 and J4 nematodes infected by C. anguillulae became immobile whereas uninfected regions remained mobile. The J2 infected nematodes usually died within 1-2 days. Tissues adjacent to C. anguillulae hyphae or developing sporangia appeared to be relatively unaffected and ultrastructurally normal. The lack of an apparent host response in the nematode suggested either a lack of hypersensitivity or a long evolutionary host-parasite relationship. Copyright 1999 Academic Press.     

Post-Infection Development and Histopathology of Meloidogyne incognita in Resistant Cotton

The numbers of Meloidogyne incognita larvae which migrated from cotton roots declined over a 16-day period, but the difference in numbers migrating from resistant and susceptible cultivars was not significant. Larvae penetrated susceptible roots, matured, and reproduced within 14 days following inoculation, whereas nematode development in the resistant roots was greatly retarded. Three types of histological responses were observed in infected, resistant roots, and these correlated with the degree of nematode development. Some galls were examined which contained only fragments of nematodes; others contained no detectable traces of developing larvae. Formation of druses in galls, but not in healthy tissue, was noted in both cultivars 20 days after inoculation. Massive invasion of roots resulted in deep longitudinal fissures of root cortex.     

腊梅的受精作用及胚胎发生

腊梅(Chimonanthus praecox)花两性,离心皮雌蕊着生在杯状花托上,柱头线形,干性。花粉经昆虫传播,落在柱头上1 d后萌发,第8d从珠孔进入,第14d左右完成双受精,为珠孔受精。胚乳为核型胚乳;初生胚乳核经短暂休眠进行核分裂,位于合点端的游离核首先形成细胞,并从合点向珠孔端细胞化,第37d胚乳充满整个囊腔。合子经过近2周的休眠后开始分裂,随着胚的发育,大部分胚乳降解,为胚的发育提供营养。合点端的胚乳细胞则侵入合点珠心组织,为胚进一步发育提供营养。其胚胎发生为柳叶菜型。     

Histopathogenesis of Galls Induced by Meloidogyne naasi in Wheat Roots

Histopathogenesis of galls induced by Meloidogyne naasi in wheat roots was studied. Large numbers of larvae penetrated wheat root tips within 24 hr; larvae migrated both inter- and intracellularly, causing cortical hypertrophy. Giant cells were formed in the stele around the head of each nematode within 4 to 5 days. Initial pathological alterations in giant cell formation consisted of hypertrophy of protophloem and protoxylem cells, their nuclei and nucleoli. Giant ceils contained 2 to 8 agglomerated multinucleolate nuclei. Synchronous mitotic divisions were first observed 9 days after inoculation. After 21 days, giant cells became highly vacuolate. Observations 40 days after inoculation revealed a complete degeneration of cell contents in many giant cells but their thick walls remained intact. Abnormal xylem completely surrounded the degenerated or partially degenerated giant cells.