Flagellar Length Research Articles

Level crossing statistics in a biologically motivated model of a long dynamic protrusion: passage times, random and extreme excursions

Long cell protrusions, which are effectively one-dimensional, are highly dynamic subcellular structures. The lengths of many such protrusions keep fluctuating about the mean value even in the steady state. We develop here a stochastic model motivated by length fluctuations of a type of appendage of an eukaryotic cell called flagellum (also called cilium). Exploiting the techniques developed for the calculation of level-crossing statistics of random excursions of stochastic process, we have derived analytical expressions of passage times for hitting various thresholds, sojourn times of random excursions beyond the threshold and the extreme lengths attained during the lifetime of these model flagella. We identify different parameter regimes of this model flagellum that mimic those of the wildtype and mutants of a well-known flagellated cell. By analysing our model in these different parameter regimes, we demonstrate how mutation can alter the level-crossing statistics even when the steady state length remains unaffected by the same mutation. Comparison of the theoretically predicted level crossing statistics, in addition to mean and variance of the length, in the steady state with the corresponding experimental data can be used in the near future as stringent tests for the validity of the models of flagellar length control. The experimental data required for this purpose, though never reported until now, can be collected, in principle, using a method developed very recently for flagellar length fluctuations.

The kinesin of the flagellum attachment zone in Leishmania is required for cell morphogenesis, cell division and virulence in the mammalian host

Leishmania parasites possess a unique and complex cytoskeletal structure termed flagellum attachment zone (FAZ) connecting the base of the flagellum to one side of the flagellar pocket (FP), an invagination of the cell body membrane and the sole site for endocytosis and exocytosis. This structure is involved in FP architecture and cell morphogenesis, but its precise role and molecular composition remain enigmatic. Here, we characterized Leishmania FAZ7, the only known FAZ protein containing a kinesin motor domain, and part of a clade of trypanosomatid-specific kinesins with unknown functions. The two paralogs of FAZ7, FAZ7A and FAZ7B, display different localizations and functions. FAZ7A localizes at the basal body, while FAZ7B localizes at the distal part of the FP, where the FAZ structure is present in Leishmania. While null mutants of FAZ7A displayed normal growth rates, the deletion of FAZ7B impaired cell growth in both promastigotes and amastigotes of Leishmania. The kinesin activity is crucial for its function. Deletion of FAZ7B resulted in altered cell division, cell morphogenesis (including flagellum length), and FP structure and function. Furthermore, knocking out FAZ7B induced a mis-localization of two of the FAZ proteins, and disrupted the molecular organization of the FP collar, affecting the localization of its components. Loss of the kinesin FAZ7B has important consequences in the insect vector and mammalian host by reducing proliferation in the sand fly and pathogenicity in mice. Our findings reveal the pivotal role of the only FAZ kinesin as part of the factors important for a successful life cycle of Leishmania.

Longer Sperm Swim More Slowly in the Canary Islands Chiffchaff.

Sperm swimming performance affects male fertilization success, particularly in species with high sperm competition. Understanding how sperm morphology impacts swimming performance is therefore important. Sperm swimming speed is hypothesized to increase with total sperm length, relative flagellum length (with the flagellum generating forward thrust), and relative midpiece length (as the midpiece contains the mitochondria). We tested these hypotheses and tested for divergence in sperm traits in five island populations of Canary Islands chiffchaff (Phylloscopus canariensis). We confirmed incipient mitochondrial DNA differentiation between Gran Canaria and the other islands. Sperm swimming speed correlated negatively with total sperm length, did not correlate with relative flagellum length, and correlated negatively with relative midpiece length (for Gran Canaria only). The proportion of motile cells increased with relative flagellum length on Gran Canaria only. Sperm morphology was similar across islands. We thus add to a growing number of studies on passerine birds that do not support sperm morphology–swimming speed hypotheses. We suggest that the swimming mechanics of passerine sperm are sufficiently different from mammalian sperm that predictions from mammalian hydrodynamic models should no longer be applied for this taxon. While both sperm morphology and sperm swimming speed are likely under selection in passerines, the relationship between them requires further elucidation.

Redistribution of FLAgellar Member 8 during the trypanosome life cycle: Consequences for cell fate prediction.

The single flagellum of African trypanosomes is essential in multiple aspects of the parasites' development. The FLAgellar Member 8 protein (FLAM8), localised to the tip of the flagellum in cultured insect forms of Trypanosoma brucei, was identified as a marker of the locking event that controls flagellum length. Here, we investigated whether FLAM8 could also reflect the flagellum maturation state in other parasite cycle stages. We observed that FLAM8 distribution extended along the entire flagellar cytoskeleton in mammalian‐infective forms. Then, a rapid FLAM8 concentration to the distal tip occurs during differentiation into early insect forms, illustrating the remodelling of an existing flagellum. In the tsetse cardia, FLAM8 further localises to the entire length of the new flagellum during an asymmetric division. Strikingly, in parasites dividing in the tsetse midgut and in the salivary glands, the amount and distribution of FLAM8 in the new flagellum were seen to predict the daughter cell fate. We propose and discuss how FLAM8 could be considered a meta‐marker of the flagellum stage and maturation state in trypanosomes.

Characterization of Bottlenose Dolphin (Tursiops truncatus) Sperm Based on Morphometric Traits.

Simple SummaryDolphins are one of the best adapted aquatic mammalians in captivity. While these animals can reproduce naturally in aquaria, many aspects related to their reproduction are still unknown. Their behavior, sociability with humans and ability to learn make dolphins ideal subjects for investigating their reproduction features for conservation programs aimed at this aquatic species and others. It is known that dolphins use a multi-male mating strategy, in which sperm competition could play a fundamental role. This study aims to characterize the shape and dimensions of dolphin sperm from two mature males and putatively classifies them into subpopulations. Moreover, the influence of sex hormone levels (testosterone) and refrigeration (temperature and storage period) with sperm dimension was evaluated. The results indicated that sperm dimensions and shape differed between the two males studied and that the sperm of both males could be classified into two subpopulations depending on their dimensions. Moreover, both testosterone levels and refrigeration were seen to influence sperm dimensions. This investigation provides new insights into sperm competition in dolphin species, and the results could be extrapolated to other endangered aquatic mammalian species.Bottlenose dolphin (Tursiops truncatus) males follow many reproductive strategies to ensure their paternity. However, little is known about the sperm traits, including morphometric features, that contribute to their reproductive success. Our aim was to study dolphin sperm morphometry (a total of 13 parameters) in two adult males to evaluate (i) presumptive sperm subpopulations, (ii) the correlation of sperm morphometry with testosterone levels and (iii) the effect of refrigerated storage on the sperm morphometry. Sperm populations were classified into four principal components (PCs) based on morphometry (>94% of cumulative variance). The PCs clustered into two different sperm subpopulations, which differed between males. Furthermore, the levels of serum testosterone were positively correlated with the length of the midpiece but negatively correlated with head width and the principal piece, flagellum and total sperm lengths. Most of the sperm morphometric parameters changed during the storage period (day 1 vs. day 7), but only the principal piece length was affected by the storage temperature (5 °C vs. 15 °C). This is the first study to identify dolphin sperm subpopulations based on morphometry and the influence of serum testosterone and refrigeration on sperm morphometry.

A polo-like kinase modulates cytokinesis and flagella biogenesis in Giardia lamblia

BackgroundPolo-like kinases (PLKs) are conserved serine/threonine kinases that regulate the cell cycle. To date, the role of Giardia lamblia PLK (GlPLK) in cells has not been studied. Here, we report our investigation on the function of GlPLK to provide insight into the role of this PKL in Giardia cell division, especially during cytokinesis and flagella formation.MethodsTo assess the function of GIPLK, Giardia trophozoites were treated with the PLK-specific inhibitor GW843286X (GW). Using a putative open reading frame for the PLK identified in the Giardia genomic database, we generated a transgenic Giardia expressing hemagglutinin (HA)-tagged GlPLK and used this transgenic for immunofluorescence assays (IFAs). GlPLK expression was knocked down using an anti-glplk morpholino to observe its effect on the number of nuclei number and length of flagella. Giardia cells ectopically expressing truncated GlPLKs, kinase domain + linker (GlPLK-KDL) or polo-box domains (GlPLK-PBD) were constructed for IFAs. Mutant GlPLKs at Lys51, Thr179 and Thr183 were generated by site-directed mutagenesis and then used for the kinase assay. To elucidate the role of phosphorylated GlPLK, the phosphorylation residues were mutated and expressed in Giardia trophozoitesResultsAfter incubating trophozoites with 5 μM GW, the percentage of cells with > 4 nuclei and longer caudal and anterior flagella increased. IFAs indicated that GlPLK was localized to basal bodies and flagella and was present at mitotic spindles in dividing cells. Morpholino-mediated GlPLK knockdown resulted in the same phenotypes as those observed in GW-treated cells. In contrast to Giardia expressing GlPLK-PBD, Giardia expressing GlPLK-KDL was defective in terms of GIPLK localization to mitotic spindles and had altered localization of the basal bodies in dividing cells. Kinase assays using mutant recombinant GlPLKs indicated that mutation at Lys51 or at both Thr179 and Thr183 resulted in loss of kinase activity. Giardia expressing these mutant GlPLKs also demonstrated defects in cell growth, cytokinesis and flagella formation.ConclusionsThese data indicate that GlPLK plays a role in Giardia cell division, especially during cytokinesis, and that it is also involved in flagella formation.Graphical

Analysis of biological noise in the flagellar length control system

SummaryAny proposed mechanism for organelle size control should be able to account not only for average size but also for the variation in size. We analyzed cell-to-cell variation and within-cell variation of length for the two flagella in Chlamydomonas, finding that cell-to-cell variation is dominated by cell size, whereas within-cell variation results from dynamic fluctuations. Fluctuation analysis suggests tubulin assembly is not directly coupled with intraflagellar transport (IFT) and that the observed length fluctuations reflect tubulin assembly and disassembly events involving large numbers of tubulin dimers. Length variation is increased in long-flagella mutants, an effect consistent with theoretical models for flagellar length regulation. Cells with unequal flagellar lengths show impaired swimming but improved gliding, raising the possibility that cells have evolved mechanisms to tune biological noise in flagellar length. Analysis of noise at the level of organelle size provides a way to probe the mechanisms determining cell geometry.

New insights in Cercopithecinae spermatozoa.

Sperm morphometric and morphologic data have been shown to represent useful tools for monitoring fertility, improving assisted reproduction techniques and conservation of genetic material as well as detecting inbreeding of endangered primates. We provide here for the first time sperm morphologic and morphometric data from Cercopithecus neglectus, Cercopithecus cephus, Papio papio and critically endangered Cercopithecus roloway, as well as comparative data from other Cercopithecinae species, i.e. Allochrocebus lhoesti, Mandrillus sphinx and Papio anubis. Following collection from the epididymis, spermatozoa were measured for each species for the following parameters: head length, head width, head perimeter, head area, midpiece length and total flagellum length, and the head volume, ellipticity, elongation, roughness and regularity were then calculated. Our data are consistent with both the general morphology and the morphometric proportions of Cercopithecinae sperm. Some specificities were observed, with C. cephus displaying a narrow head (width = 2.76 ± 0.26 µM) and C. roloway displaying a short midpiece (6.65 ± 0.61 µM). This data set represents an important contribution, especially for Cercopithecus roloway, one of the most endangered monkeys in the world, and further data on additional specimens coupled to data on mating systems and reproductive ecology should allow a better understanding of the mechanisms underlying these morphological differences across primate species.

Defects in the cytoplasmic assembly of axonemal dynein arms cause morphological abnormalities and dysmotility in sperm cells leading to male infertility.

Axonemal protein complexes, such as outer (ODA) and inner (IDA) dynein arms, are responsible for the generation and regulation of flagellar and ciliary beating. Studies in various ciliated model organisms have shown that axonemal dynein arms are first assembled in the cell cytoplasm and then delivered into axonemes during ciliogenesis. In humans, mutations in genes encoding for factors involved in this process cause structural and functional defects of motile cilia in various organs such as the airways and result in the hereditary disorder primary ciliary dyskinesia (PCD). Despite extensive knowledge about the cytoplasmic assembly of axonemal dynein arms in respiratory cilia, this process is still poorly understood in sperm flagella. To better define its clinical relevance on sperm structure and function, and thus male fertility, further investigations are required. Here we report the fertility status in different axonemal dynein preassembly mutant males (DNAAF2/ KTU, DNAAF4/ DYX1C1, DNAAF6/ PIH1D3, DNAAF7/ZMYND10, CFAP300/C11orf70 and LRRC6). Besides andrological examinations, we functionally and structurally analyzed sperm flagella of affected individuals by high-speed video- and transmission electron microscopy as well as systematically compared the composition of dynein arms in sperm flagella and respiratory cilia by immunofluorescence microscopy. Furthermore, we analyzed the flagellar length in dynein preassembly mutant sperm. We found that the process of axonemal dynein preassembly is also critical in sperm, by identifying defects of ODAs and IDAs in dysmotile sperm of these individuals. Interestingly, these mutant sperm consistently show a complete loss of ODAs, while some respiratory cilia from the same individual can retain ODAs in the proximal ciliary compartment. This agrees with reports of solely one distinct ODA type in sperm, compared to two different ODA types in proximal and distal respiratory ciliary axonemes. Consistent with observations in model organisms, we also determined a significant reduction of sperm flagellar length in these individuals. These findings are relevant to subsequent studies on the function and composition of sperm flagella in PCD patients and non-syndromic infertile males. Our study contributes to a better understanding of the fertility status in PCD-affected males and should help guide genetic and andrological counselling for affected males and their families.

Flagellar arrangements in elongated peritrichous bacteria: bundle formation and swimming properties

The surface distribution of flagella in peritrichous bacterial cells has been traditionally assumed to be random. Recently, the presence of a regular grid-like pattern of basal bodies has been suggested. Experimentally, the manipulation of the anchoring points of flagella in the cell membrane is difficult, and thus, elucidation of the consequences of a particular pattern on bacterial locomotion is challenging. We analyze the bundle formation process and swimming properties of Bacillus subtilis-like cells considering random, helical, and ring-like arrangements of flagella by means of mesoscale hydrodynamics simulations. Helical and ring patterns preferentially yield configurations with a single bundle, whereas configurations with no clear bundles are most likely for random anchoring. For any type of pattern, there is an almost equally low probability to form V-shaped bundle configurations with at least two bundles. Variation of the flagellum length yields a clear preference for a single major bundle in helical and ring patterns as soon as the flagellum length exceeds the body length. The average swimming speed of cells with a single or two bundles is rather similar, and approximately 50% larger than that of cells of other types of flagellar organization. Considering the various anchoring patterns, rings yield the smallest average swimming speed independent of the type of bundle, followed by helical arrangements, and largest speeds are observed for random anchoring. Hence, a regular pattern provides no advantage in terms of swimming speed compared to random anchoring of flagella, but yields more likely single-bundle configurations.Graphic abstract

A CRISPR interference platform for selective downregulation of gene expression in Borrelia burgdorferi.

The spirochete Borrelia burgdorferi causes Lyme disease, an increasingly prevalent infection. While previous studies have provided important insight into B. burgdorferi biology, many aspects, including basic cellular processes, remain underexplored. To help speed up the discovery process, we adapted a CRISPR interference (CRISPRi) platform for use in B. burgdorferi For efficiency and flexibility of use, we generated various CRISPRi template constructs that produce different basal and induced levels of dcas9 and carry different antibiotic resistance markers. We characterized the effectiveness of our CRISPRi platform by targeting the motility and cell morphogenesis genes flaB, mreB, rodA, and ftsI, whose native expression levels span two orders of magnitude. For all four genes, we obtained gene repression efficiencies of at least 95%. We showed by darkfield microscopy and cryo-electron tomography that flagellin (FlaB) depletion reduced the length and number of periplasmic flagella, which impaired cellular motility and resulted in cell straightening. Depletion of FtsI caused cell filamentation, implicating this protein in cell division in B. burgdorferi Finally, localized cell bulging in MreB- and RodA-depleted cells matched the locations of new peptidoglycan insertion specific to spirochetes of the Borrelia genus. These results therefore implicate MreB and RodA in the particular mode of cell wall elongation of these bacteria. Collectively, our results demonstrate the efficiency and ease of use of our B. burgdorferi CRISPRi platform, which should facilitate future genetic studies of this important pathogen.IMPORTANCE Gene function studies are facilitated by the availability of rapid and easy-to-use genetic tools. Homologous recombination-based methods traditionally used to genetically investigate gene function remain cumbersome to perform in B. burgdorferi, as they often are relatively inefficient. In comparison, our CRISPRi platform offers an easy and fast method to implement as it only requires a single plasmid transformation step and IPTG addition to obtain potent (>95%) downregulation of gene expression. To facilitate studies of various genes in wild-type and genetically modified strains, we provide over 30 CRISPRi plasmids that produce distinct levels of dcas9 expression and carry different antibiotic resistance markers. Our CRISPRi platform represents a useful and efficient complement to traditional genetic and chemical methods to study gene function in B. burgdorferi.

Some recent data on sperm morphology and motility kinetics in Atlantic cod (Gadus morhua L.).

Studying biology of sperm provides valuable information to optimize artificial reproduction and is crucial for sustainable aquaculture. Here, we investigated morphology of spermatozoon in Atlantic cod (Gadus morhua) using transmission and scanning electron microscopy. Furthermore, spermatozoa motility kinetics at different osmolalities were studied using computer-assisted sperm analysis software. The spermatozoon lacked an acrosome and consisted of a head, midpiece, and flagellum. The head of spermatozoa was round, oval, and rather elongated in shape, showing high variations in dimensions. There were up to 6 mitochondria that encircled the proximal part of the flagellum. The proximal and distal centrioles were located within the nuclear notch and arranged orthogonal to each other. The axoneme had a typical 9 + 2 microtubule structure. The flagellar length of spermatozoon was 66.94 ± 0.46μm. Spermatozoa were immotile in the seminal plasma. Dilution of sperm with natural seawater (1100mOsmol/kg) resulted in initiation of motility for 91.0 ± 3.4% of spermatozoa with average velocity of 86.2 ±2.3μm/s and beating frequency of 52Hz. The duration of spermatozoa motility was > 6min; however, the percentage of motile spermatozoa decreased at 60s post-activation. When osmolality of natural seawater was modified using distilled water or NaCl, spermatozoa motility was not initiated at ≤ 400 and ≥ 2500mOsmol/kg, and the highest percentage of motility was observed at 730-1580mOsmol/kg. In a sucrose solution, spermatozoa motility was initiated and suppressed at 600 and 1500mOsmol/kg, respectively, and highest percentage of motility was observed at 800-1100mOsmol/kg. Spermatozoon morphology comparisons within Gadiformes showed differences in dimensions of head and mitochondria, flagellar length, and number of mitochondria. The present study provides valuable data that can be used for phylogenetic implications based on spermatozoon morphology and for development of artificial fertilization and sperm cryopreservation protocols based on sperm motility.

Review of Chilean Cyrtinae (Diptera: Acroceridae) with the Description of Three New Species and the First Record of Villalus inanis from Argentina.

Acrocerid flies are endoparasitoids of spiders that are distributed in all ecosystems except deserts and the poles. This group was historically classified into three subfamilies. However, molecular analyses have indicated two new subfamilies. One of these is Cyrtinae Newman, which is made up of two clades: the north, with Palearctic species, and the south, with Chilean species (except for the fossil genus Villalites Hennig). In the southern clade, endemic genera Holops Philippi and Villalus Cole have been poorly described over time by different authors. The aim of this study is to re-describe the known species of Holops and Villalus and describe three new Holops species based on collected material: Holops grezi sp. nov., Holops anarayae sp. nov., and Holops pullomen sp. nov. In addition, an identification species key with updated distributions for all species and the first record of Villalus inanis from Argentina is also provided. New species are mainly characterized by the length of flagellum; colors of the postocellar lobe and thoracic segments, pilosity and legs; wing cells and veins, and shape of the abdomen. Holops and Villalus do not share morphological traits with any other known extant genus of Acroceridae in the southern hemisphere. The real biodiversity of this group must be evaluated with a molecular phylogeny in the future. In addition, the new species in the Pehuén and Valdivian forest provinces, areas considered to have been sheltered during the last glacial period, promoting speciation in several taxa. New and cryptic Holops species open the doors to disentangling the real biodiversity of spider flies in Chile.

Effects of breed and ejaculate volume on sperm morphology and semen parameters of boars.

The aim of the study was to determine the relation between the semen quality, frequency of sperm defects, sperm dimensions and shape, and the ejaculate volume of Large White and Landrace boars. A total of 648 ejaculates collected from 31 Large White and 30 Landrace boars were divided into three groups according to the criterion of the ejaculate volume. In this study Landrace boars produced ejaculates with higher volume, sperm concentration, and total numbers of spermatozoa than Large White boars. Landrace boars also showed a lower frequency of sperm with morphological abnormalities (P < 0.05). Landrace boars sperm had larger heads, which were by 0.15 μm longer, and by a larger perimeter and area (P < 0.05). Landrace boar spermatozoa also had a longer flagellum and were generally larger and by 2.07 μm longer than Large White boar sperm (P < 0.05). Significant differences were also found in the shape of sperm of the two breeds (P < 0.05). Landrace boars sperm had more elongated heads, and the ratio of head size to flagellum length was lower than in Large White boars sperm (P < 0.05). Sperm from ejaculates with low volume had a shorter flagellum and a greater head length/flagellum length ratio than sperm from medium- and high-volume ejaculates (P < 0.05).

Energy sources that fuel metabolic processes in protruding finger‐like organelles

Cells possess a variety of organelles with characteristic structure and subcellular localization intimately linked to their specific function. While most are intracellular and found in virtually all eukaryotic cells, there is a small group of organelles of elongated cylindrical shapes in highly specialized cells that protrude into the extracellular space, such as cilia, flagella, and microvilli. The ATP required by intracellular organelles is amply available in the cytosol, largely generated by mitochondria. However, such is not the case for cilia and flagella, whose slender structures cannot accommodate mitochondria. These organelles consume massive amounts of ATP to carry out high energy-demanding functions, such as sensory transduction or motility. ATP from the nearest mitochondria or other reactions within the cell body is severely limited by diffusion and generally insufficient to fuel the entire length of cilia and flagella. These organelles overcome this fuel restriction by local generation of ATP, using mechanisms that vary depending on the nutrients that are available in their particular external environment. Here, we review, with emphasis in mammals, the remarkable adaptations that cilia and flagella use to fuel their metabolic needs. Additionally, we discuss how a decrease in nutrients surrounding olfactory cilia might impair olfaction in COVID-19 patients.

Cell size influences inorganic carbon acquisition in artificially selected phytoplankton.

Cell size influences the rate at which phytoplankton assimilate dissolved inorganic carbon (DIC), but it is unclear whether volume-specific carbon uptake should be greater in smaller or larger cells. On the one hand, Fick's Law predicts smaller cells to have a superior diffusive CO2 supply. On the other, larger cells may have greater scope to invest metabolic energy to upregulate active transport per unit area through CO2 -concentrating mechanisms (CCMs). Previous studies have focused on among-species comparisons, which complicates disentangling the role of cell size from other covarying traits. In this study, we investigated the DIC assimilation of the green alga Dunaliella tertiolecta after using artificial selection to evolve a 9.3-fold difference in cell volume. We compared CO2 affinity, external carbonic anhydrase (CAext ), isotopic signatures (δ13 C) and growth among size-selected lineages. Evolving cells to larger sizes led to an upregulation of CCMs that improved the DIC uptake of this species, with higher CO2 affinity, higher CAext and higher δ13 C. Larger cells also achieved faster growth and higher maximum biovolume densities. We showed that evolutionary shifts in cell size can alter the efficiency of DIC uptake systems to influence the fitness of a phytoplankton species.

Production of common carp donor-derived offspring from goldfish surrogate broodstock

Common carp (Cyprinus carpio) is the fourth most-produced species in worldwide aquaculture. Significant efforts are invested in breeding and preservation of genetic integrity of this important species. However, maintaining a carp gene bank in situ can be demanding due to the large body size of the fish. Moreover, ex situ gene banking in common carp is as in other fish species limited to sperm cryopreservation. Recent progress in reproductive biotechnology in fishes has allowed the transfer of germ stem cells (gamete precursors) between individuals or even species. After maturation, the recipients are producing gametes of the donor. Surrogacy can serve as a valuable tool in germplasm banking to recover cryopreserved germ stem cells. Some unfavourable characteristics of the donor species such as long maturation time or large body size hampering its reproduction can be overcome by choosing a surrogate with more convenient characteristics. Efficient protocols for cryopreservation of common carp male and female germ stem cells have been recently developed in our laboratory. The next step has been to assess the potential of smaller goldfish (Carassius auratus) surrogates to produce donor-derived gametes of common carp after intraperitoneal transplantation of testicular cells.High transplantation success was achieved when 44% of the surviving goldfish produced pure donor-derived gametes of common carp at the age of 3 years giving rise to viable progeny. Donor-derived identity of the offspring was confirmed by genotyping and the production of a typical phenotype corresponding to the donor species. Reproductive performance of chimeras was similar to goldfish controls. Assessment of gamete characteristics showed that the size of donor-derived eggs is between control common carp and goldfish eggs. Interestingly, flagellum length in donor-derived spermatozoa was comparable to the common carp flagellum and significantly shorter than goldfish flagellum. Genotyping of the Y chromosome locus in chimeric goldfish provided a novel insight on the fate of transplanted cells, where only genotypic females were capable of producing eggs, while both genotypic males and females were capable of producing sperm. The present technology can be combined with cryopreservation procedures to ease needs for common carp breeds preservation and their recovery using many times smaller goldfish surrogates but also serve as a convenient model to study interspecific surrogacy in cyprinids.

Testicular histology of Anurans that deposit eggs out of water

Anuran amphibians show the largest diversity of amphibian reproductive modes, many of which imply egg deposition out of the water. This kind of egg deposition requires specialisations to avoid egg desiccation. Physiological, anatomical and ethological traits integrate to define these modes. In particular, morphological features of the urogenital system correlate with these reproductive modes and the environmental conditions where egg-laying occurs. In this study, we describe the testicular histology and spermatogenesis of the nest building frog Leptodactylus latinasus, and we compare it with other species that breed out of water. We found variations in tes tis size, the thickness of interstitial tissue, tunica albuginea, and peritubular tunics, flagellum length, and in the shape and size of the spermatozoal nucleus. Certain specifics’ characters differed at the species level, but not between families. Such variation could be an indicator of spermatozoal performance and environmental constraints under which fertilisation takes place.

Flagellar length control in monoflagellates by motorized transport: Growth kinetics and correlations of length fluctuations

How does a cell self-organize so that its appendages attain specific lengths that are convenient for their respective functions? What kind of ‘rulers’ does a cell use to measure the length of these appendages? How does a cell transport structure building materials between the cell body and distal tips of these appendages so as to regulate their dynamic lengths during various stages of its lifetime? Some of these questions are addressed here in the context of a specific cell appendage called flagellum (also called cilium). A “time of flight” (ToF) mechanism, adapted from the pioneering idea of Galileo, has been used successfully very recently to explain the length control of flagella by a biflagellate green algae. Using the same ToF mechanism, here we develop a stochastic model for the dynamics of flagella in two different types of monoflagellate unicellular organisms. A unique feature of these monoflagellates is that these become transiently multi-flagellated during a short span of their life time. The mean length of the flagella in our model reproduce the trend of their temporal variation observed in experiments. Moreover, for probing the intracellular molecular communication between the dynamic flagella of a given cell, we have computed the correlation in the fluctuations of their lengths during the multiflagellated stage of the cell cycle by Monte Carlo simulation.

Control of filament length by a depolymerizing gradient

Cells assemble microns-long filamentous structures from protein monomers that are nanometers in size. These structures are often highly dynamic, yet in order for them to function properly, cells maintain them at a precise length. Here we investigate length-dependent depolymerization as a mechanism of length control. This mechanism has been recently proposed for flagellar length control in the single cell organisms Chlamydomonas and Giardia. Length dependent depolymerization can arise from a concentration gradient of a depolymerizing protein, such as kinesin-13 in Giardia, along the length of the flagellum. Two possible scenarios are considered: a linear and an exponential gradient of depolymerizing proteins. We compute analytically the probability distributions of filament lengths for both scenarios and show how these distributions are controlled by key biochemical parameters through a dimensionless number that we identify. In Chlamydomonas cells, the assembly dynamics of its two flagella are coupled via a shared pool of molecular components that are in limited supply, and so we investigate the effect of a limiting monomer pool on the length distributions. Finally, we compare our calculations to experiments. While the computed mean lengths are consistent with observations, the noise is two orders of magnitude smaller than the observed length fluctuations.