Free-swimming Zoospores Research Articles

Characterizing the “fungal shunt”: Parasitic fungi on diatoms affect carbon flow and bacterial communities in aquatic microbial food webs

Microbial interactions in aquatic environments profoundly affect global biogeochemical cycles, but the role of microparasites has been largely overlooked. Using a model pathosystem, we studied hitherto cryptic interactions between microparasitic fungi (chytrid Rhizophydiales), their diatom host Asterionella, and cell-associated and free-living bacteria. We analyzed the effect of fungal infections on microbial abundances, bacterial taxonomy, cell-to-cell carbon transfer, and cell-specific nitrate-based growth using microscopy (e.g., fluorescence in situ hybridization), 16S rRNA gene amplicon sequencing, and secondary ion mass spectrometry. Bacterial abundances were 2 to 4 times higher on individual fungal-infected diatoms compared to healthy diatoms, particularly involving Burkholderiales. Furthermore, taxonomic compositions of both diatom-associated and free-living bacteria were significantly different between noninfected and fungal-infected cocultures. The fungal microparasite, including diatom-associated sporangia and free-swimming zoospores, derived ∼100% of their carbon content from the diatom. By comparison, transfer efficiencies of photosynthetic carbon were lower to diatom-associated bacteria (67 to 98%), with a high cell-to-cell variability, and even lower to free-living bacteria (32%). Likewise, nitrate-based growth for the diatom and fungi was synchronized and faster than for diatom-associated and free-living bacteria. In a natural lacustrine system, where infection prevalence reached 54%, we calculated that 20% of the total diatom-derived photosynthetic carbon was shunted to the parasitic fungi, which can be grazed by zooplankton, thereby accelerating carbon transfer to higher trophic levels and bypassing the microbial loop. The herein termed "fungal shunt" can thus significantly modify the fate of photosynthetic carbon and the nature of phytoplankton-bacteria interactions, with implications for diverse pelagic food webs and global biogeochemical cycles.

Molecular phylogenetic and zoospore ultrastructural analyses of Chytridium olla establish the limits of a monophyletic Chytridiales

Chytridium olla A. Braun, the first described chytrid and an obligate algal parasite, is the type for the genus and thus the foundation of family Chytridiaceae, order Chytridiales, class Chytridiomycetes and phylum Chytridiomycota. Chytridium olla was isolated in coculture with its host, Oedogonium capilliforme. DNA was extracted from the coculture, and 18S, 28S and ITS1-5.8S-ITS2 rDNA were amplified with universal fungal primers. Free swimming zoospores and zoospores in mature sporangia were examined with electron microscopy. Molecular analyses placed C. olla in a clade in Chytridiales with isolates of Chytridium lagenaria and Phlyctochytrium planicorne. Ultrastructural analysis revealed C. olla to have a Group II-type zoospore, previously described for Chytridium lagenaria and Phlyctochytrium planicorne. On the basis of zoospore ultrastructure, family Chytridiaceae is emended to include the type of Chytridium and other species with a Group II-type zoospore, and the new family Chytriomycetaceae is delineated to include members of Chytridiales with a Group I-type zoospore.

Phylogenetic Position of the Copepod-Infesting Parasite Syndinium turbo (Dinoflagellata, Syndinea)

Sequences were determined for the nuclear-encoded small subunit (SSU) rRNA and 5.8S rRNA genes as well as the internal transcribed spacers ITS1 and ITS2 of the parasitic dinoflagellate genus Syndinium from two different marine copepod hosts. Syndinium developed a multicellular plasmodium inside its host and at maturity free-swimming zoospores were released. Syndinium plasmodia in the copepod Paracalanus parvus produced zoospores of three different morphological types. However, full SSU rDNA sequences for the three morphotypes were 100% identical and also their ITS1-ITS2 sequences were identical except for four base pairs. It was concluded that the three morphotypes belong to a single species that was identified as Syndinium turbo, the type species of the dinoflagellate subdivision Syndinea. The SSU rDNA sequence of another Syndinium species infecting Corycaeus sp. was similar to Syndinium turbo except for three base pairs and the ITS1-ITS2 sequences of the two species differed at 34-35 positions. Phylogenetic analyses placed Syndinium as a sister taxon to the blue crab parasite Hematodinium sp. and both parasites were affiliated with the so-called marine alveolate Group II. This corroborates the hypothesis that marine alveolate Group II is Syndinea.

Factors influencing the sporulation and cyst formation of Aphanomyces invadans, etiological agent of ulcerative mycosis in Atlantic menhaden, Brevoortia tyrannus

Oomycete infections caused by Aphanomyces invadans occur in freshwater and estuarine fishes around the world. Along the east coast of the USA, skin ulcers caused by A. invadans are prevalent in Atlantic menhaden, Brevoortia tyrannus. From laboratory observations low salinities appear crucial to transmission of the pathogen. To better understand aspects of transmission, we characterized sporulation and cyst formation of secondary zoospores of two isolates of A. invadans at different salinities and temperatures. Sporulation occurred only at low salinities. At room temperature (ca. 20–22 C), using “pond water” augmented with artificial sea salts, the endemic strain WIC and the Thailand strain PA7 of A. invadans produced free-swimming secondary zoospores at salinities of 0, 1 and 2 psu (practical salinity unit= ‰), but not at 4 psu or higher. Secondary zoospores of another species, ATCC-62427 (Aphanomyces sp.), were observed at 1, 2, 4 and 8 psu but not at 0 and 12 psu. Secondary zoospores of all three isolates, especially WIC, were abundant and motile 1–2 d post-sporulation. Sporulation was temperature dependent and occurred over a relatively narrow range. No sporulation occurred at 4, 30 or 35 C for either WIC or PA7. For both strains zoospore production within 1–3 d after the initiation of sporulation was more prolific at 25 C than at 20 and 15 C. At 15 C production of zoospores was sustained over 11 d for WIC and 5 d for PA7. At room temperature single WIC secondary zoospores remained motile 12–18 h. Salinities exceeding 4 psu or vigorous shaking caused immediate cyst formation of WIC secondary zoospores. Exposure to menhaden tissue, but not tissues of other fishes to secondary zoospores (WIC), caused rapid (2 h) cyst formation. Cysts were capable of excysting when transferred to 1 psu water within 2–3 h of cyst formation. Cysts that had remained encysted in 6.5 psu for 24 h did not excyst when transferred to 1 psu water. Salinity and temperature requirements for sporulation indicate that juvenile menhaden must acquire infections during rain or in low salinity oligohaline waters.

Factors influencing the sporulation and cyst formation of Aphanomyces invadans, etiological agent of ulcerative mycosis in Atlantic menhaden, Brevoortia tyrannus

Oomycete infections caused by Aphanomyces invadans occur in freshwater and estuarine fishes around the world. Along the east coast of the USA, skin ulcers caused by A. invadans are prevalent in Atlantic menhaden, Brevoortia tyrannus. From laboratory observations low salinities appear crucial to transmission of the pathogen. To better understand aspects of transmission, we characterized sporulation and cyst formation of secondary zoospores of two isolates of A. invadans at different salinities and temperatures. Sporulation occurred only at low salinities. At room temperature (ca. 20-22 C), using "pond water" augmented with artificial sea salts, the endemic strain WIC and the Thailand strain PA7 of A. invadans produced free-swimming secondary zoospores at salinities of 0, 1 and 2 psu (practical salinity unit = per thousand), but not at 4 psu or higher. Secondary zoospores of another species, ATCC-62427 (Aphanomyces sp.), were observed at 1, 2, 4 and 8 psu but not at 0 and 12 psu. Secondary zoospores of all three isolates, especially WIC, were abundant and motile 1-2 d postsporulation. Sporulation was temperature dependent and occurred over a relatively narrow range. No sporulation occurred at 4, 30 or 35 C for either WIC or PA7. For both strains zoospore production within 1-3 d after the initiation of sporulation was more prolific at 25 C than at 20 and 15 C. At 15 C production of zoospores was sustained over 11 d for WIC and 5 d for PA7. At room temperature single WIC secondary zoospores remained motile 12-18 h. Salinities exceeding 4 psu or vigorous shaking caused immediate cyst formation of WIC secondary zoospores. Exposure to menhaden tissue, but not tissues of other fishes to secondary zoospores (WIC), caused rapid (2 h) cyst formation. Cysts were capable of excysting when transferred to 1 psu water within 2-3 h of cyst formation. Cysts that had remained encysted in 6.5 psu for 24 h did not excyst when transferred to 1 psu water. Salinity and temperature requirements for sporulation indicate that juvenile menhaden must acquire infections during rain or in low salinity oligohaline waters.

Intraplant Variation in Nuclear DNA Content in Laminaria saccharina and Alaria esculenta (Phaeophyceae)

Intraplant variation in nuclear DNA content was quantified in DAPI-stained cells of Laminaria saccharina and Alaria esculenta. In both species, meristoderm, cortical and medullary cells were examined from vegetative fronds. In addition, DNA was quantified during sporogenesis, in free-swimming zoospores and gametophytes of A. esculenta. Zoospores and gametophytes had 1C (= N) levels of nuclear DNA in all cells, but vegetative cells of sporophytes varied from 2C to 16C. In A. esculenta nuclear DNA was significantly correlated with cell length, total cell volume and nuclear volume, whereas in Laminaria saccharina these correlations were not apparent. During sporogenesis of Alaria esculenta, maximal nuclear DNA levels of 8C were present in post-meiotic nuclei. During four subsequent mitotic divisions (to form 64 zoospores per sporangium at the 1C level) only one cycle of DNA synthesis occurred. These results show that mitosis can be independent of DNA synthesis in Laminariales, and that endopolyploidy is a generalized feature of multicellular plants.

Cyllamyces aberensis gen.nov. sp.nov., a new anaerobic gut fungus with branched sporangiophores isolated from cattle

A new genus of the anaerobic gut fungi (Neocallimastigales), Cyllamyces aberensis gen.nov. sp.nov., with bulbous holdfast, branched sporangiophores and limited polycentric thallus development is described. The fungus was isolated from fresh cattle faeces. Free-swimming zoospores were spherical, uninucleate, and uniflagellate. After encystment, zoospores germinated and gave rise to a single, bulbous holdfast. One or several branched sporangiophores were produced from different locations on the holdfast, each bearing several spherical sporangia. DAPI staining of thalli indicated that nuclei were present in the holdfast, sporangiophores, and sporangia. As many as 12 sporangia were observed per thallus on up to 5 sporangiophores, with zoosporogenesis beginning 16–18 h after encystment. Zoospore ultrastructure was examined by transmission electron microscopy and found to be similar to that reported for other anaerobic chytrid fungi. Organelles were evenly distributed throughout the cell, except for the posteriorly attached flagellum and associated attachment apparatus, the hydrogenosomes, which were mainly situated in the posterior parts of the cell and a posteriorly directed, beak-shaped nucleus. Limited polycentric thallus development (including branched sporangiophores), the possession of a single bulbous holdfast and the absence of rhizoids were stable features of this fungus that distinguished it from the other five genera of gut fungi. Therefore, we have used these characteristics to assign the fungus to a new genus, Cyllamyces, with the specific name C. aberensis.Key words: rumen, fungal taxonomy, Neocallimastigales, chytrid, zoospore ultrastructure.

<i>Cyllamyces aberensis</i> gen.nov. sp.nov., a new anaerobic gut fungus with branched sporangiophores isolated from cattle

A new genus of the anaerobic gut fungi (Neocallimastigales), Cyllamyces aberensis gen.nov. sp.nov., with bulbous holdfast, branched sporangiophores and limited polycentric thallus development is described. The fungus was isolated from fresh cattle faeces. Free-swimming zoospores were spherical, uninucleate, and uniflagellate. After encyst - ment, zoospores germinated and gave rise to a single, bulbous holdfast. One or several branched sporangiophores were produced from different locations on the holdfast, each bearing several spherical sporangia. DAPI staining of thalli in - dicated that nuclei were present in the holdfast, sporangiophores, and sporangia. As many as 12 sporangia were ob- served per thallus on up to 5 sporangiophores, with zoosporogenesis beginning 16-18 h after encystment. Zoospore ultrastructure was examined by transmission electron microscopy and found to be similar to that reported for other an - aerobic chytrid fungi. Organelles were evenly distributed throughout the cell, except for the posteriorly attached flagellum and associated attachment apparatus, the hydrogenosomes, which were mainly situated in the posterior parts of the cell and a posteriorly directed, beak-shaped nucleus. Limited polycentric thallus development (including branched sporangiophores), the possession of a single bulbous holdfast and the absence of rhizoids were stable features of this fungus that distinguished it from the other five genera of gut fungi. Therefore, we have used these characteris- tics to assign the fungus to a new genus, Cyllamyces, with the specific name C. aberensis.

Lagenidium giganteumZoospores: Effects of Concentration, Movement, Light, and Temperature on Infection of Mosquito Larvae

Assays conducted with a California isolate ofLagenidium giganteumrevealed that the LC50against second-instarCulex quinquefasciatuslarvae in pond water was 26 zoospores/ml. The maximum distance that zoospores swam horizontally and infected second-instar larvae was 60 cm. The maximum depth from which zoospores moved upward in a column of water and infected larvae was between 30 and 40 cm. The release depth of zoospores in a column of water at which 50% larval mortality occurred was 19.6 cm. Free-swimming zoospores did not exhibit positive or negative attraction to incandescent light as evidenced by larval mortality. Survival of zoospores as indicated by mosquito larval mortality was greatest at 25°C and was similar at 30, 33, and 35°C. No infection occurred at 17°C and <20% larval mortality occurred at 19°C for any age of zoospores.

The role of anaerobic gut fungi in ruminants

Anaerobic chytridiomycete fungi are found in the gastrointestinal tracts of sheep, cattle and goats, as well as in many other domesticated ruminant and nonruminant herbivores and a wide variety of wild herbivorous mammals. They are principally found associated with the fibrous plant particles of digesta and as free swimming zoospores in the fluid phase. The presence of large fungal populations in animals consuming mature pasture or diets largely composed of hay or straw together with the production of highly active fibre degrading enzymes lead to' the belief that anaerobic fungi may have a significant role to play in the assimilation of fibrous feeds by ruminants. While many early studies focused on anaerobic fungi because of their unusual biology and metabolism, the large part of subsequent research has emphasized the biotechnological potential of their cellulases, xylanases and phenolic esterases. In recent years, the extent of the contribution of anaerobic fungi to the nutrition of ruminants has also been established through studies of fungal populations in the rumen and the dietary factors which influence them, as presented in this review. Further, we discuss the evidence supporting an important contribution of anaerobic fungal populations in the rumen to feed intake and digestion of poor quality feed by domesticated ruminants. In conclusion, the review explores some different methods for manipulating fungi in the rumen for increased feed intake and digestion.

Photosynthesis in reproductive unicells of Ulva fasciata and Enteromorpha flexuosa:implications for ecological success

MEPS Marine Ecology Progress Series Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsTheme Sections MEPS 125:229-237 (1995) - doi:10.3354/meps125229 Photosynthesis in reproductive unicells of Ulva fasciata and Enteromorpha flexuosa: implications for ecological success Beach KS, Smith CM, Michael T, Shin HW Photosynthetic performance of adult vegetative tissues is modified following the formation of free-swimming reproductive unicells by Ulvafasciata and Enteromorphaflexuosa. Comparisons of photosynthetic performance of adult haploid or diploid thalli versus gametes or zoospores (respectively) yield the following generalizations: (1) Pmax (maximum net photosynthetic rate) for motile cells (e.g. gametes of U. fasciata and zoospores of E. flexuosa) can be equivalent to Pmax of appropriate adult tissues when expressed on a chlorophyll a (chl a) or chl a+b basis; (2) respiration rates for all motile cells were 1.5- to 6-fold higher than rates for adult thalli when normalized to chl; (3) as expected with increasing respiration rates, compensation irradiance (Ic) and saturation irradiance (Ik) also increased for all motile cell types; (4) chl a:b ratios were lower in all motile cell types than in adult cells; (5) P:R (photosynthesis:respiration) ratios were 3.3- to 7.7-fold lower in zoospores of U. fasciata and E. flexuosa respectively. Photosynthetic performance of settled zoospores of U. fasciata was more similar to the vegetative parent than to the free-swimming zoospores in terms of Ik and Pmax even with altered chl a:b ratios. An unexpected finding was the increased invivo absorbance in 490 to 550 nm range for reproductive tissues of U.fasciata only. This absorbance increase suggests a potential photoadaptation. This potential photoadaptation, coupled with alteration of the chl a:b ratio, could allow reproductive unicells of U. fasciata to harvest light more readily for photosynthesis as part of the plankton when compared to zoospores of E. flexuosa. Such a difference between motile cells may enhance the competitive ability and subsequent ecological success of one over the other. High rates of Pmax have the selective advantage of subsidizing high respiratory costs associated with motility in reproductive unicells while still allowing for rapid growth upon eventual attachment to the substrate. Comparably, species such as late successional kelps do not show this pattern and may represent a different photoadaptive strategy. Photosynthesis. Zoospores . Gametes . Sporophytes. Gametophytes . Invivo absorption . Ulvafasciata . Enteromorphaflexuosa Full text in pdf format PreviousNextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in MEPS Vol. 125. Publication date: September 14, 1995 Print ISSN:0171-8630; Online ISSN:1616-1599 Copyright © 1995 Inter-Research.

Use of zoospore concentrations and life cycle parameters in determining the population of anaerobic fungi in the rumen ecosystem

An indirect approach to quantification of the fibrolytic anaerobic fungi in the rumen is described. A mathematical model of the life cycle of anaerobic fungi, based upon observations of the life histories and growth kinetics of these organisms in vitro and in vivo, is constructed and solved in the steady-state to determine the population of particle-attached (substrate-associated) fungal thalli from the concentration of free-swimming zoospores in rumen liquid. The values obtained are broadly consistent with ruminal observations and with observations on faecal populations, which assume that a significant proportion of fungi leaving the rumen (as cysts or spores) can ultimately be accounted for in the faeces.

Chemotactic auto-aggregation in the water mould Achlya

SUMMARY: Free-swimming zoospores of several isolates of the oomycetous water mould Achlya demonstrate mutual attraction (adelphotaxis) which may result in auto-aggregation. In the apparent absence of exogenous chemotactic signals macroscopic globular aggregates or microscopic plaques of cysts are formed. Auto-aggregation was favoured by high concentrations of zoospores in suspension; the growth rate of spontaneous aggregates was estimated at 4--10 x 103spores min−1; growth of an aggregate was limited to about 7 min. By means of chemotactic assays, attractant activity for zoospores was detected in the vicinity of aggregates, and in zoospore or cyst supernatants. The attractant activity extracted from spore aggregates was heat-stable. In the absence of exogenous nutrients, the macroscopic aggregates of an homothallic isolate produced sporangia and oogonia. Aggregation was readily initiated as a result of zoospore chemotaxis towards capillary tubes containing exogenous attractant in an agar medium. Aggregating zoospores frequently assumed a slender swarm configuration moving directly towards the aggregate at the capillary-tube tip. Analysis of video recordings showed that many zoospores joining the swarm were attracted directly by the swarm rather than by the exogenous attractant. Such aggregation was a time-limited process, the duration of which was extended by the presence of amino acids in the zoospore suspension. Under natural conditions zoospore aggregation may be a dual-component chemotactic process triggered by exogenous attractants and amplified by chemical signals from the aggregating spores. The possible consequences of adelphotaxis and auto-aggregate formation, namely enhanced inoculum potential, syngamy and spore production, are discussed.

Cytochemical Detection of Polysaccharides and the Ultrastructure of the Cell Coat of Zoospores of Chytriomyces aureus and Chytriomyces hyalinus

The formation and structure of cell coats on zoospores of Chytriomyces aureus and C. hyalinus are described from an ultrastructural and cytochemical study. After zoospore initials are cleaved, but before organelles are organized into the arrangement found in free-swimming zoospores, dictyosomal vesicles deposit a cell coat on the outer surface of the plasma membrane. In free-swimming zoospores, the cell coat covers all of the zoospore except the flageUar sheath. The inner layer of the cell coat is homogeneously dense and is covered with an outer layer of helically arranged spines. Both layers of the cell coat stain with the silver methenamine technique for polysaccharides, as do glycogen and fibril-containing vesicles in the cytoplasm. Sodium bisulfite and hydrogen peroxide controls confirm the polysaccharide specificity of the reaction in glycogen deposits, vesicles, and the outer layer of the cell coat. Use of the sulfhydryl blocking agent, iodoacetate, demonstrates sulfhydryl-containing compounds in the inner layer of the cell coat.

Cytochemical Detection of Polysaccharides and the Ultrastructure of the Cell Coat of Zoospores ofChytriomyces AureusandChytriomyces Hyalinus

The formation and structure of cell coats on zo6spores of Chytriomyces aureus and C. hyalinus are described from an ultrastructural and cytochemical study. After zoospore initials are cleaved, but before organelles are organized into the arrangement found in free-swimming zoospores, dictyosomal vesicles deposit a cell coat on the outer surface of the plasma membrane. In free-swimming zoospores, the cell coat covers all of the zoospore except the flagellar sheath. The inner layer of the cell coat is homogeneously dense and is covered with an outer layer of helically arranged spines. Both layers of the cell coat stain with the silver methenamine technique for polysaccharides, as do glycogen and fibril-containing vesicles in the cytoplasm. Sodium bisulfite and hydrogen peroxide controls confirm the polysaccharide specificity of the reaction in glycogen deposits, vesicles, and the outer layer of the cell coat. Use of the sulfhydryl blocking agent, iodoacetate, demonstrates sulfhydryl-containing compounds in the inner layer of the cell coat.

SOME GENERAL OBSERVATIONS ON ZOOSPORES OF THE CHYTRID RHIZOPHYDIUM PLANKTONICUM CANTER EMEND.

SummaryGeneral microscopic observations of zoospores of the chytrid Rhizophydium planktonicum Canter emend, contained within a small glass chamber, have shown that they have strong reactions to visible light. There appears to exist a threshold level of dim light below which zoospores will eventually enter what is termed the ‘state of quiescence’ and above which they will continue to maintain active swimming. Alternations between these two states can he manipulated with ease under the microscope. Free swimming zoospores after a period in dim light or darkness and quiescent zoospores both respond to sudden exposure to bright light.The concentration of zoospores into aggregates of individuals circling at the upper and or lower surfaces of a chamber is recorded. As viewed from above, those uppermost circle in a clockwise direction while those below go round counter‐clockwise.Adherence between zoospores as swimming individuals or in the encysted state has been seen but there is no evidence of any fusion taking place. A specially adapted channel image intensifier made possible the examination of material under infra‐red.

Ultrastructural, cytochemical, and enzymatic studies on the adhesive ?plaques? of the brown algaeLaminaria saccharina (L.) lamour. andNereocystis luetkeana (nert.) post. et rupr. 1

Newly settled zoospores of bothLaminaria saccharina andNereocystis luetkeana are surrounded by adhesive plaques. At this stage cell wall deposition has not occurred and similar structures cannot be detected in the cytoplasm. In free swimming zoospores, however, plaques are observed exclusively in small vesicles within the cytoplasm. Enzymatic extractions and cytochemical tests indicate that plaques are glycoproteid in nature. Studies on the influence of several enzymes on the attachment of zoospores show that plaques are involved in the adhesion of zoospores to the substratum.

Zoosporangia, believed to be those of Plasmodiophora brassicæ, in the Root Hairs of Non-cruciferous Plants

IN 1936, during work on the cytology of P. brassicae, zoosporangia identical with those of this well-known parasite were seen by me in root hairs of grass and dock seedlings, which had appeared in a pot of soil taken from a garden in which the club root disease of the Cruciferse was a major problem. In view of the known persistence of P. brassicae in soils without cruciferous crops, it was decided to attempt a crude cross-inoculation experiment using club root galls as a starting point. Suitable soil was partially sterilized by keeping it at 190° F. for one hour. Ten six-inch pots were filled with this soil, and club root galls with unbroken surfaces, but of an age known to contain mature spore masses, were cut up and placed in five of them. Brussels sprout seedlings germinated in these pots became infected, and an examination of the root hairs of the grass Holcus lanatus grown from seed with them in the re-infected soil revealed numerous zoosporangia and some free-swimming zoospores.