Ubiquitin-dependent Argonauteprotein MEL1 degradation is essential for rice sporogenesis and phasiRNA target regulation.

MEIOSIS ARRESTED AT LEPTOTENE1 (MEL1), a rice (Oryza sativa) Argonaute (AGO) protein, has been reported to function specifically at premeiotic and meiotic stages of germ cell development and is associated with a novel class of germ cell-specific small noncoding RNAs called phased small RNAs (phasiRNAs). MEL1 accumulation is temporally and spatially regulated and is eliminated after meiosis. However, the metabolism and turnover (i.e. the homeostasis) of MEL1 during germ cell development remains unknown. Here, we show that MEL1 is ubiquitinated and subsequently degraded via the proteasome pathway in vivo during late sporogenesis. Abnormal accumulation of MEL1 after meiosis leads to a semi-sterile phenotype. We identified a monocot-specific E3 ligase, XBOS36, a CULLIN RING-box protein, that is responsible for the degradation of MEL1. Ubiquitination at four K residues at the N terminus of MEL1 by XBOS36 induces its degradation. Importantly, inhibition of MEL1 degradation either by XBOS36 knockdown or by MEL1 overexpression prevents the formation of pollen at the microspore stage. Further mechanistic analysis showed that disrupting MEL1 homeostasis in germ cells leads to off-target cleavage of phasiRNA target genes. Our findings thus provide insight into the communication between a monocot-specific E3 ligase and an AGO protein during plant reproductive development.

G protein α subunit suppresses sporangium formation through a serine/threonine protein kinase in Phytophthora sojae.

Eukaryotic heterotrimeric guanine nucleotide-binding proteins consist of α, ß, and γ subunits, which act as molecular switches to regulate a number of fundamental cellular processes. In the oomycete pathogen Phytophthora sojae, the sole G protein α subunit (Gα; encoded by PsGPA1) has been found to be involved in zoospore mobility and virulence, but how it functions remains unclear. In this study, we show that the Gα subunit PsGPA1 directly interacts with PsYPK1, a serine/threonine protein kinase that consists of an N-terminal region with unknown function and a C-terminal region with a conserved catalytic kinase domain. We generated knockout and knockout-complemented strains of PsYPK1 and found that deletion of PsYPK1 resulted in a pronounced reduction in the production of sporangia and oospores, in mycelial growth on nutrient poor medium, and in virulence. PsYPK1 exhibits a cytoplasmic-nuclear localization pattern that is essential for sporangium formation and virulence of P. sojae. Interestingly, PsGPA1 overexpression was found to prevent nuclear localization of PsYPK1 by exclusively binding to the N-terminal region of PsYPK1, therefore accounting for its negative role in sporangium formation. Our data demonstrate that PsGPA1 negatively regulates sporangium formation by repressing the nuclear localization of its downstream kinase PsYPK1.

Germination of Spores of the Orders Bacillales and Clostridiales.

Dormant Bacillales and Clostridiales spores begin to grow when small molecules (germinants) trigger germination, potentially leading to food spoilage or disease. Germination-specific proteins sense germinants, transport small molecules, and hydrolyze specific bonds in cortex peptidoglycan and specific proteins. Major events in germination include (a) germinant sensing; (b) commitment to germinate; (c) release of spores' depot of dipicolinic acid (DPA); (d) hydrolysis of spores' peptidoglycan cortex; and (e) spore core swelling and water uptake, cell wall peptidoglycan remodeling, and restoration of core protein and inner spore membrane lipid mobility. Germination is similar between Bacillales and Clostridiales, but some species differ in how germinants are sensed and how cortex hydrolysis and DPA release are triggered. Despite detailed knowledge of the proteins and signal transduction pathways involved in germination, precisely what some germination proteins do and how they do it remain unclear.

The zoospores of the thraustochytrid Aurantiochytrium limacinum: Transcriptional reprogramming and lipid metabolism associated to their specific functions.

Aurantiochytrium limacinum (Thraustochytriaceae, class Labyrinthulomycetes) is a marine Stramenopile and a pioneering mangrove decomposer. Its life cycle involves a non-motile stage and zoospore production. We observed that the composition of the medium, the presence of amino acids in particular, affects the release of zoospores. Two opposite conditions were defined, one with a cell population mainly composed of zoospores and another one with almost only non-motile cells. In silico allelic frequency analysis and flow cytometry suggest that zoospores and non-motile cells share the same ploidy level and are diploid. Through an RNA-seq approach, the transcriptional reprogramming accompanying the formation of zoospores was investigated, with a particular focus on their lipid metabolism. Based on a differential expression analysis, zoospores are characterized by high motility, very active signal transduction, an arrest of the cell division, a low amino acid metabolism and low glycolysis. Focusing on lipid metabolism, genes involved in lipase activities and peroxisomal ß-oxidation are upregulated. qRT-PCR of selected lipid genes and lipid analyses during the life span of zoospores confirmed these observations. These results highlight the importance of the lipid dynamics in zoospores and show the metabolic processes required to use these energy-dense molecules as fuel for zoospore survival during their quest of new territories.

Topical Application of Adult Cecal Contents to Eggs Transplants Spore-Forming Microbiota but Not Other Members of the Microbiota to Chicks.

The intestinal microbiota plays an essential role in the metabolism and immune competence of chickens from the first day after hatching. In modern production systems, chicks are isolated from adult chickens, instead hatching in a clean environment. As a result, chicks are colonized by environmental bacteria, including potential pathogens. There is a need to investigate methods by which chicks can be exposed to a more appropriate microbial community at hatching. Such methods must be easy to apply in a hatchery and produce consistent results. The development of the intestinal microbiota of chicks hatched from eggs sprayed with dilute adult cecal content during incubation was observed at 0, 3, 7, and 14 days posthatching (dph) across two experiments. High-throughput Illumina sequencing was performed for the V4 hypervariable region of the 16S rRNA gene. A topical treatment of dilute adult cecal content was sufficient to transplant spore-forming bacteria such as Lachnospiraceae and Ruminococcaceae However, this treatment was not able to transplant other taxa that are considered to be core elements of the chicken cecal microbiota, such as Bacteroidaceae, Lactobacillaceae, Bifidobacteriaceae, and Burkholderiaceae The topical treatment significantly altered the microbiota of chicks immediately posthatching and accelerated the normal development of the microbiota with earlier colonization by Ruminococcaceae in the cecum and "Candidatus Arthromitus" in the ileum. The effect of the treatment on the cecal microbiota was maximal at 3 dph but diminished over time.IMPORTANCE Over the last 60 years poultry production has intensified in response to increased demand for meat. In modern systems, chicks hatch without contacting chickens and their gut bacteria. Consequently, they are colonized by environmental bacteria that may cause disease. The normal bacteria that live in the gut, or intestinal microbiota, play an important role in the development of the immune system. Therefore, it is essential to find easy ways to expose chicks to the more appropriate bacteria at hatching. This experiment investigated whether spraying eggs with adult cecal contents was sufficient to transfer an adult microbiota to chicks. Our findings show that spore-forming bacteria were transplanted, but other members of the microbiota were not. In this respect, the spray application was partially successful, but the timing of the spray needs to be modified to ensure that more bacteria are transferred.

Effects of sophorolipids on fungal and oomycete pathogens in relation to pH solubility.

AIMS: The objective of this study was to determine the effects of sophorolipids on several fungal and oomycete plant pathogens and the relationship between sophorolipids at different pH and antimicrobial activities. METHODS AND RESULTS: Sophorolipids had different solubility at different pH with a dramatic increase in solubility when pH was 6 or higher. Inhibition of mycelial growth of Phytophthora infestans by sophorolipids was affected by pH values, showing that when the pH value was higher, the inhibition rate was lower. Sophorolipids inhibited spore germination and mycelial growth of several fungal and oomycete pathogens in vitro including Fusarium sp., F. oxysporum, F. concentricum, Pythium ultimum, Pyricularia oryzae, Rhizoctorzia solani, Alternaria kikuchiana, Gaeumannomyces graminis var. tritici and P. infestans and caused morphological changes in hyphae by microscope observation. Sophorolipids reduced ß-1,3-glucanase activity in mycelia of P. infestans. In greenhouse studies, foliar application of sophorolipids at 3 mg ml-1 reduced severity of late blight of potato caused by P. infestans significantly. CONCLUSION: Sophorolipids influenced spore germination and hyphal tip growth of several plant pathogens and pH solubility of sophorolipids had an effect on their efficacy. Application of sophorolipids reduced late blight disease on potato under greenhouse conditions. SIGNIFICANCE AND IMPACT OF THE STUDY: The findings indicated that sophorolipids have the potential to be developed as a convenient and easy-to-use formulation for managing plant diseases.

A quantitative method to assess the role of indoor air decontamination to simultaneously reduce contamination of environmental surfaces: testing with vegetative and spore-forming bacteria.

Indoor air can spread pathogens, which can be removed/inactivated by a variety of means in healthcare and other settings. We quantitatively assessed if air decontamination could also simultaneously reduce environmental surface contamination in the same setting. Two types of vegetative bacteria (Staphylococcus aureus and Acinetobacter baumannii), and a bacterial spore-former (Geobacillus stearothermophilus) were tested as representative airborne bacteria. They were separately aerosolized with a Collison nebulizer into a 24-m3 aerobiology chamber and air samples collected with a programmable slit-to-agar sampler. Settling airborne particles were collected on culture plates placed at, and collected from, five different locations on the floor of the chamber with a custom-built remote plate-placement and -retriever system. Experimentally contaminated air in the chamber was decontaminated for 45 min with a device based on HEPA filtration and UV light. The plates were incubated and CFU counted. The device reduced the viability levels of all tested bacteria in the air by >3 log10 (>99·9%) in 45 min. Based on two separate tests, the average reductions in surface contamination for S. aureus, A. baumannii and G. stearothermophilus were respectively, 97, 87 and 97%. We thus showed that air decontamination could substantially and simultaneously reduce the levels of surface contamination in the same setting irrespective of the type of pathogen present. SIGNIFICANCE AND IMPACT OF THE STUDY: The innovative and generic test protocol described can quantitatively assess the reduction in environmental surface contamination from microbial decontamination of indoor air in the same setting. This added advantage from air decontamination has implications for infection prevention and control in healthcare and other settings without the need for additional expense or effort. Continuous operation of an air decontamination device, such as the one tested here, can lead to ongoing reductions in pathogens in air and on environmental surfaces.

New host records of three Kudoa spp. (K. yasunagai, K. thalassomi, and K. igami) with notable variation in the number of shell valves and polar capsules in spores.

To date, 26 Kudoa spp. (Myxozoa: Myxosporea: Multivalvulida) have been recorded in edible marine fishes in Japan. In the future, it is likely that even more marine fish multivalvulid myxosporeans will be characterized morphologically and genetically, which will aid the precise understanding of their biodiversity and biology. We examined 60 individuals of six fish species collected from the Philippine Sea off Kochi or from the border between the Philippine Sea and East China Sea around Miyako Island, Okinawa, i.e., the southern part of Japan. Newly collected parasite species included Kudoa yasunagai from the brain of Japanese meagre (Argyrosomus japonicus) and Japanese parrotfish (Calotomus japonicus), Kudoa miyakoensis n. sp. and Kudoa thalassomi from the brain and trunk muscle, respectively, of bluespine unicornfish (Naso unicornis), and Kudoa igami from the trunk muscle of Carolines parrotfish (Calotomus carolinus), African coris (Coris gaimard), and Pastel ringwrasse (Hologymnosus doliatus). With the exception of Japanese parrotfish for K. yasunagai, all these fish are new host records for each kudoid species. Notable variation in the number of shell valves (SV) and polar capsules (PC) was observed for all four kudoid species. In particular, spores with seven or eight SV/PC were prominent in K. igami isolates, despite the original Japanese parrotfish-derived description characterizing it as having spores with six, or less commonly five, SV/PC. However, molecular genetic characterization based on the ribosomal RNA gene (rDNA) and mitochondrial DNA (cytochrome c oxidase subunit 1 and ribosomal RNA small and large subunits) found no significant differences in the nucleotide sequences of isolates with different phenotypical features as far as examined in the present study. A newly erected species, K. miyakoensis n. sp., was determined to be phylogenetically closest to brain-parasitizing species, such as K. chaetodoni, K. lemniscati, and K. yasunagai based on rDNA nucleotide sequences, but differed from them morphologically.

The Effect of pH on Spore Germination, Growth, and Infection of Strawberry Roots by Fusarium oxysporum f. sp. fragariae, Cause of Fusarium wilt of Strawberry.

Previous work has shown that raising the pH of acidic soil to near neutrality can reduce the incidence of Fusarium wilt. The basis for this effect has not been established. The present study assessed effects of pH on spore germination, growth, and infection of strawberry roots by Fusarium oxysporum f. sp. fragariae, the cause of Fusarium wilt of strawberry. There was not a significant effect of pH (5 versus 7) on the rate of spore germination at either 20 or 25°C for any of the three tested isolates (one representative of each clonal lineage of F. oxysporum f. sp. fragariae found in California). Likewise, pH did not have a significant effect on fungal growth at 20°C. At 25°C, two isolates grew faster at pH 7 than at pH 5. Growth of the third isolate was unaffected by the difference in pH. For the strawberry cultivar Albion, the frequency of root infection was significantly higher for plants grown in acidified soil (near pH 5) than for plants grown in soil near neutrality. The higher frequency of root infection in acidified soil was associated with a lower level of microbial activity, as measured by hydrolysis of fluorescein diacetate.

Ultrasensitive Response of Developing Myxococcus xanthus to the Addition of Nutrient Medium Correlates with the Level of MrpC.

Upon depletion of nutrients, Myxococcus xanthus forms mounds on a solid surface. The differentiation of rod-shaped cells into stress-resistant spores within mounds creates mature fruiting bodies. The developmental process can be perturbed by the addition of nutrient medium before the critical period of commitment to spore formation. The response was investigated by adding a 2-fold dilution series of nutrient medium to starving cells. An ultrasensitive response was observed, as indicated by a steep increase in the spore number after the addition of 12.5% versus 25% nutrient medium. The level of MrpC, which is a key transcription factor in the gene regulatory network, correlated with the spore number after nutrient medium addition. The MrpC level decreased markedly by 3 h after adding nutrient medium but recovered more after the addition of 12.5% than after 25% nutrient medium addition. The difference in MrpC levels was greatest midway during the period of commitment to sporulation, and mound formation was restored after 12.5% nutrient medium addition but not after adding 25% nutrient medium. Although the number of spores formed after 12.5% nutrient medium addition was almost normal, the transcript levels of "late" genes in the regulatory network failed to rise normally during the commitment period. However, at later times, expression from a reporter gene fused to a late promoter was higher after adding 12.5% than after adding 25% nutrient medium, consistent with the spore numbers. The results suggest that a threshold level of MrpC must be achieved in order for mounds to persist and for cells within to differentiate into spores.IMPORTANCE Many signaling and gene regulatory networks convert graded stimuli into all-or-none switch-like responses. Such ultrasensitivity can produce bistability in cell populations, leading to different cell fates and enhancing survival. We discovered an ultrasensitive response of M. xanthus to nutrient medium addition during development. A small change in nutrient medium concentration caused a profound change in the developmental process. The level of the transcription factor MrpC correlated with multicellular mound formation and differentiation into spores. A threshold level of MrpC is proposed to be necessary to initiate mound formation and create a positive feedback loop that may explain the ultrasensitive response. Understanding how this biological switch operates will provide a paradigm for the broadly important topic of cellular behavior in microbial communities.

Ecophysiology and lipid dynamics of a eukaryotic mangrove decomposer.

Aurantiochytrium limacinum is an osmo-heterotrophic Stramenopile and a pioneering mangrove decomposer which is taxonomically assigned to the family of Thraustochytriaceae (class: Labyrinthulomycetes). The life cycle of A. limacinum involves different cell types including mono- and multi-nucleated cells as well as flagellated zoospores which colonize new fallen leaves. The ecological relevance of thraustochytrids is underestimated and eclipsed by their biotechnological importance, due to their ability to accumulate large amount of lipids, mainly triacylglycerols (TAGs). In this study, we aimed to understand the ecophysiological parameters that trigger zoospore production and the interplay between the life cycle of A. limacinum and its lipid metabolism. When grown in a rich medium, cells accumulated large amounts of TAGs at the end of their growth period, but no zoospores were produced. In poor media such as artificial sea water, zoospores were produced in massive quantities. In the absence of organic carbon, the zoospores remained swimming for at least 6 days, consuming their TAGs in the process. Addition of glucose rapidly triggered the maturation of the zoospores. On the basis of these data, we propose a life cycle for A. limacinum integrating the potential perturbations/changes in the environment surrounding a mangrove leaf that could lead to the production of zoospores and colonization of new areas.

Reorganization of plasma membrane lipid domains during conidial germination.

Neurospora crassa, a filamentous fungus, in the unicellular conidial stage has ideal features to study sphingolipid (SL)-enriched domains, which are implicated in fundamental cellular processes ranging from antifungal resistance to apoptosis. Several changes in lipid metabolism and in the membrane composition of N. crassa occur during spore germination. However, the biophysical impact of those changes is unknown. Thus, a biophysical study of N. crassa plasma membrane, particularly SL-enriched domains, and their dynamics along conidial germination is prompted. Two N. crassa strains, wild-type (WT) and slime, which is devoid of cell wall, were studied. Conidial growth of N. crassa WT from a dormancy state to an exponential phase was accompanied by membrane reorganization, namely an increase of membrane fluidity, occurring faster in a supplemented medium than in Vogel's minimal medium. Gel-like domains, likely enriched in SLs, were found in both N. crassa strains, but were particularly compact, rigid and abundant in the case of slime cells, even more than in budding yeast Saccharomyces cerevisiae. In N. crassa, our results suggest that the melting of SL-enriched domains occurs near growth temperature (30°C) for WT, but at higher temperatures for slime. Regarding biophysical properties strongly affected by ergosterol, the plasma membrane of slime conidia lays in between those of N. crassa WT and S. cerevisiae cells. The differences in biophysical properties found in this work, and the relationships established between membrane lipid composition and dynamics, give new insights about the plasma membrane organization and structure of N. crassa strains during conidial growth.

Cytological and Proteomic Analyses of Osmunda cinnamomea Germinating Spores Reveal Characteristics of Fern Spore Germination and Rhizoid Tip Growth.

Fern spore is a good single-cell model for studying the sophisticated molecular networks in asymmetric cell division, differentiation, and polar growth. Osmunda cinnamomea L. var. asiatica is one of the oldest fern species with typical separate-growing trophophyll and sporophyll. The chlorophyllous spores generated from sporophyll can germinate without dormancy. In this study, the spore ultrastructure, antioxidant enzyme activities, as well as protein and gene expression patterns were analyzed in the course of spore germination at five typical stages (i.e. mature spores, rehydrated spores, double-celled spores, germinated spores, and spores with protonemal cells). Proteomic analysis revealed 113 differentially expressed proteins, which were mainly involved in photosynthesis, reserve mobilization, energy supplying, protein synthesis and turnover, reactive oxygen species scavenging, signaling, and cell structure modulation. The presence of multiple proteoforms of 25 differentially expressed proteins implies that post-translational modification may play important roles in spore germination. The dynamic patterns of proteins and their encoding genes exhibited specific characteristics in the processes of cell division and rhizoid tip growth, which include heterotrophic and autotrophic metabolisms, de novo protein synthesis and active protein turnover, reactive oxygen species and hormone (brassinosteroid and ethylene) signaling, and vesicle trafficking and cytoskeleton dynamic. In addition, the function skew of proteins in fern spores highlights the unique and common mechanisms when compared with evolutionarily divergent spermatophyte pollen. These findings provide an improved understanding of the typical single-celled asymmetric division and polar growth during fern spore germination.

Down-regulation of Rad51 activity during meiosis in yeast prevents competition with Dmc1 for repair of double-strand breaks.

Interhomolog recombination plays a critical role in promoting proper meiotic chromosome segregation but a mechanistic understanding of this process is far from complete. In vegetative cells, Rad51 is a highly conserved recombinase that exhibits a preference for repairing double strand breaks (DSBs) using sister chromatids, in contrast to the conserved, meiosis-specific recombinase, Dmc1, which preferentially repairs programmed DSBs using homologs. Despite the different preferences for repair templates, both Rad51 and Dmc1 are required for interhomolog recombination during meiosis. This paradox has recently been explained by the finding that Rad51 protein, but not its strand exchange activity, promotes Dmc1 function in budding yeast. Rad51 activity is inhibited in dmc1Δ mutants, where the failure to repair meiotic DSBs triggers the meiotic recombination checkpoint, resulting in prophase arrest. The question remains whether inhibition of Rad51 activity is important during wild-type meiosis, or whether inactivation of Rad51 occurs only as a result of the absence of DMC1 or checkpoint activation. This work shows that strains in which mechanisms that down-regulate Rad51 activity are removed exhibit reduced numbers of interhomolog crossovers and noncrossovers. A hypomorphic mutant, dmc1-T159A, makes less stable presynaptic filaments but is still able to mediate strand exchange and interact with accessory factors. Combining dmc1-T159A with up-regulated Rad51 activity reduces interhomolog recombination and spore viability, while increasing intersister joint molecule formation. These results support the idea that down-regulation of Rad51 activity is important during meiosis to prevent Rad51 from competing with Dmc1 for repair of meiotic DSBs.

Ceratomyxa azevedoi n. sp. (Myxozoa: Myxosporea) parasitizing the gallbladder of Lutjanus ehrenbergii in the Arabian Gulf.

A novel myxosporean species, Ceratomyxa azevedoi sp. n. is described from the gallbladder of the blackspot snapper, Lutjanus ehrenbergii (Peters), captured from the Arabian Gulf off Saudi Arabia. A total of 45 (26.8%) out of 168 fish specimens were found to be infected with Ceratomyxa azevedoi sp. n., the highest prevalence being observed in winter (42.9%, 18/42) and the lowest in autumn (11.9%, 5/42). Mature spores appeared as crescent to slightly elliptical-shaped, measuring 5-7 (6) µm in length and 12 (10-14) µm in thickness, with spherical polar capsules containing three polar filament coils. The morphometric and morphological comparison with similar species revealed the taxonomic novelty of this form, suggesting that it should be considered as new species. The phylogenetic analysis of C. azevedoi sp. n., based on partial SSU rDNA sequences, revealed close genetic relatedness to C. buri with 91.3% homogeneity and to C. hamour, with 90.1% homogeneity.

[Henneguya wolinensis (Myxosporea: Myxobolidae), a new for Russian fauna parasite from the perch Perca fluviatilis L.].

The infection of the perch Perea fluviatilis L. with myxosporean Henneguya wolinensis Romuk-Wodoracki, 1990 has been detected. This is the second finding of this parasite after its original descriptin and the first for Russia. Plasmodium of this species develops in the epidermis under scales throughout the body causing the formation of white cysts up to 1 mm. Spores are fusiform, large, their average length constitutes 25.5 µm without the caudal appendages and 62 µm with them. Slight morphological differences in spore structure comparing to original description have been revealed.

Comprehensive transcriptome-based characterization of differentially expressed genes involved in microsporogenesis of radish CMS line and its maintainer.

Microsporogenesis is an indispensable period for investigating microspore development and cytoplasmic male sterility (CMS) occurrence. Radish CMS line plays a critical role in elite F1 hybrid seed production and heterosis utilization. However, the molecular mechanisms of microspore development and CMS occurrence have not been thoroughly uncovered in radish. In this study, a comparative analysis of radish floral buds from a CMS line (NAU-WA) and its maintainer (NAU-WB) was conducted using next generation sequencing (NGS) technology. Digital gene expression (DGE) profiling revealed that 3504 genes were significantly differentially expressed between NAU-WA and NAU-WB library, among which 1910 were upregulated and 1594 were downregulated. Gene ontology (GO) analysis showed that these differentially expressed genes (DEGs) were mainly enriched in extracellular region, catalytic activity, and response to stimulus. KEGG enrichment analysis revealed that the DEGs were predominantly associated with flavonoid biosynthesis, glycolysis, and biosynthesis of secondary metabolites. Real-time quantitative PCR analysis showed that the expression profiles of 13 randomly selected DEGs were in high agreement with results from Illumina sequencing. Several candidate genes encoding ATP synthase, auxin response factor (ARF), transcription factors (TFs), chalcone synthase (CHS), and male sterility (MS) were responsible for microsporogenesis. Furthermore, a schematic diagram for functional interaction of DEGs from NAU-WA vs. NAU-WB library in radish plants was proposed. These results could provide new information on the dissection of the molecular mechanisms underlying microspore development and CMS occurrence in radish.

Ceratomyxa tunisiensis n. sp. (Myxosporea: Bivalvulida) from the Gallbladders of Two Carangid Fish Caught Off the Coast of Tunisia.

A new coelozoic Myxozoan species, Ceratomyxa tunisiensis n. sp., was found infecting the gallbladders of two carangid fish, Caranx rhonchus and Trachurus trachurus (Perciforme, Carangidae), from the Gulf of Gabès, on the southern coast of Tunisia. The parasite develops in spherical mono-, diplo-, or polysporic tropozoites in the gallbladder of the hosts. Mature spores are typical of the genus Ceratomyxa. They are transversely elongated and narrowly crescent-shaped with a slightly convex anterior and concave posterior, and measure 23 ± 0. 27 (20-25) µm width × 6 ± 0.26 (5-8) µm in length. Spore shell valves are symmetrical with rounded ends. Two spherical polar capsules situated on either side of the sutural line measure 2.2 µm (2.0-3.0) in diam. Periodical sampling of C. rhonchus and T. trachurus from Marsh 2012 to February 2013 showed that infection due to C. tunisiensis occurs in 59% and 69% of the examined fish, respectively. Molecular analysis based on the small subunit (SSU) rRNA sequence shows high genetic divergence with all other ceratomyxid species. A Maximum Likelihood phylogenetic tree shows association with the species C. leatharjecketi Fiala, kova, Kodadkova, Freeman, Bartosova-Sojkova, and Atkinson, 2015 reported from the gallbladder of Aluterusmonoceros (L.) caught in the Andaman Sea, off Malaysia. Nonetheless, the SSU rRNA sequences of C. tunisiensis and C. leatharjecketi have only a 90% similarity.

MYB64 and MYB119 are required for cellularization and differentiation during female gametogenesis in Arabidopsis thaliana.

In angiosperms, the egg cell forms within the multicellular, haploid female gametophyte. Female gametophyte and egg cell development occurs through a unique process in which a haploid spore initially undergoes several rounds of synchronous nuclear divisions without cytokinesis, resulting in a single cell containing multiple nuclei. The developing gametophyte then forms cell walls (cellularization) and the resulting cells differentiate to generate the egg cell and several accessory cells. The switch between free nuclear divisions and cellularization-differentiation occurs during developmental stage FG5 in Arabidopsis, and we refer to it as the FG5 transition. The molecular regulators that initiate the FG5 transition during female gametophyte development are unknown. In this study, we show using mutant analysis that two closely related MYB transcription factors, MYB64 and MYB119, act redundantly to promote this transition. MYB64 and MYB119 are expressed during the FG5 transition, and most myb64 myb119 double mutant gametophytes fail to initiate the FG5 transition, resulting in uncellularized gametophytes with supernumerary nuclei. Analysis of cell-specific markers in myb64 myb119 gametophytes that do cellularize suggests that gametophytic polarity and differentiation are also affected. We also show using multiple-mutant analysis that MYB119 expression is regulated by the histidine kinase CKI1, the primary activator of two-component signaling (TCS) during female gametophyte development. Our data establish a molecular pathway regulating the FG5 transition and implicates CKI1-dependent TCS in the promotion of cellularization, differentiation, and gamete specification during female gametogenesis.

Callose is integral to the development of permanent tetrads in the liverwort Sphaerocarpos.

A striking feature of the liverwort Sphaerocarpos is that pairs of male and female spores remain united in permanent tetrads. To identify the nature of this phenomenon and to test the hypothesis that callose is involved, we examined spore wall development in Sphaerocarpos miche lii, with emphasis on the appearance, location and fate of callose vis-à-vis construction of the sculptoderm. All stages of sporogenesis were examined using differential interference contrast optics, and aniline blue fluorescence to locate callose. For precise localization, specimens were immunogold labeled with anti-callose antibody and observed in the transmission electron microscope. Callose plays a role in Sphaerocarpos spore wall development not described in any other plant, including other liverworts. A massive callose matrix forms outside of the sculptured sporocyte plasmalemma that predicts spore wall ornamentation. Consequently, layers of exine form across adjacent spores uniting them. Spore wall development occurs entirely within the callose and involves the production of six layers of prolamellae that give rise to single or stacked tripartite lamellae (TPL). Between spores, an anastomosing network of exine layers forms in lieu of intersporal septum development. As sporopollenin assembles on TPL, callose progressively disappears from the inside outward leaving layers of sporopollenin impregnated exine, the sculptoderm, overlying a thick fibrillar intine. This developmental mechanism provides a direct pathway from callose deposition to sculptured exine that does not involve the intermediary primexine found in pollen wall development. The resulting tetrad, encased in a single wall, provides a simple model for development of permanent dyads and tetrads in the earliest fossil plants.