Asexual reproduction induces a rapid and permanent loss of sexual reproduction capacity in the rice fungal pathogen Magnaporthe oryzae: results of in vitro experimental evolution assays

PubMed Central

2012-01-01

Background Sexual reproduction is common in eukaryotic microorganisms, with few species reproducing exclusively asexually. However, in some organisms, such as fungi, asexual reproduction alternates with episodic sexual reproduction events. Fungi are thus appropriate organisms for studies of the reasons for the selection of sexuality or clonality and of the mechanisms underlying this selection. Magnaporthe oryzae, an Ascomycete causing blast disease on rice, reproduces mostly asexually in natura. Sexual reproduction is possible in vitro and requires (i) two strains of opposite mating types including (ii) at least one female-fertile strain (i.e. a strain able to produce perithecia, the female organs in which meiosis occurs). Female-fertile strains are found only in limited areas of Asia, in which evidence for contemporary recombination has recently been obtained. We induced the forced evolution of four Chinese female-fertile strains in vitro by the weekly transfer of asexual spores (conidia) between Petri dishes. We aimed to determine whether female fertility was rapidly lost in the absence of sexual reproduction and whether this loss was controlled genetically or epigenetically. Results All the strains became female-sterile after 10 to 19 rounds of selection under asexual conditions. As no single-spore isolation was carried out, the observed decrease in the production of perithecia reflected the emergence and the invasion of female-sterile mutants. The female-sterile phenotype segregated in the offspring of crosses between female-sterile evolved strains and female-fertile wild-type strains. This segregation was maintained in the second generation in backcrosses. Female-sterile evolved strains were subjected to several stresses, but none induced the restoration of female fertility. This loss of fertility was therefore probably due to genetic rather than epigenetic mechanisms. In competition experiments, female-sterile mutants produced similar numbers of viable conidia to wild-type strains, but released them more efficiently. This advantage may account for the invasion of our populations by female-sterile mutants. Conclusions We show for the first time that, in the absence of sexual reproduction, female-sterile mutants of M. oryzae rice strains can arise and increase in abundance in asexual generations. This change in phenotype was frequent and probably caused by mutation. These results suggest that female fertility may have been lost rapidly during the dispersion of the fungus from Asia to the rest of the world. PMID:22458778

The Putative Protein Methyltransferase LAE1 of Trichoderma atroviride Is a Key Regulator of Asexual Development and Mycoparasitism

PubMed Central

Aghcheh, Razieh Karimi; Druzhinina, Irina S.; Kubicek, Christian P.

2013-01-01

In Ascomycota the protein methyltransferase LaeA is a global regulator that affects the expression of secondary metabolite gene clusters, and controls sexual and asexual development. The common mycoparasitic fungus Trichoderma atroviride is one of the most widely studied agents of biological control of plant-pathogenic fungi that also serves as a model for the research on regulation of asexual sporulation (conidiation) by environmental stimuli such as light and/or mechanical injury. In order to learn the possible involvement of LAE1 in these two traits, we assessed the effect of deletion and overexpression of lae1 gene on conidiation and mycoparasitic interaction. In the presence of light, conidiation was 50% decreased in a Δ lae1 and 30–50% increased in lae1-overexpressing (OElae1) strains. In darkness, Δ lae1 strains did not sporulate, and the OElae1 strains produced as much spores as the parent strain. Loss-of-function of lae1 also abolished sporulation triggered by mechanical injury of the mycelia. Deletion of lae1 also increased the sensitivity of T. atroviride to oxidative stress, abolished its ability to defend against other fungi and led to a loss of mycoparasitic behaviour, whereas the OElae1 strains displayed enhanced mycoparasitic vigor. The loss of mycoparasitic activity in the Δ lae1 strain correlated with a significant underexpressionn of several genes normally upregulated during mycoparasitic interaction (proteases, GH16 ß-glucanases, polyketide synthases and small cystein-rich secreted proteins), which in turn was reflected in the partial reduction of formation of fungicidal water soluble metabolites and volatile compounds. Our study shows T. atroviride LAE1 is essential for asexual reproduction in the dark and for defense and parasitism on other fungi. PMID:23826217

Epidemiology of Basil Downy Mildew.

PubMed

Cohen, Yigal; Ben Naim, Yariv; Falach, Lidan; Rubin, Avia E

2017-10-01

Basil downy mildew (BDM) caused by the oomycete Peronospora belbahrii is a destructive disease of sweet basil (Ocimum basilicum) worldwide. It originated in Uganda in the 1930s and recently spread to Europe, the Middle East, Americas, and the Far East. Seed transmission may be responsible for its quick global spread. The pathogen attacks leaf blades, producing chlorotic lesions with ample dark asexual spores on the lower leaf surface. Oospores may form in the mesophyll of infected leaves. The asexual spores germinate on a wet leaf surface within 2 h and penetrate into the epidermis within 4 h. Spore germination and infection occur at a wide range of temperatures from 5 to 28.5°C. Infection intensity depends on the length of dew period, leaf temperature, and inoculum dose. The duration of latent period (from infection to sporulation) extends from 5 to 10 days, depending on temperature and light regime. The shortest is 5 days at 25°C under continuous light. Sporulation requires high humidity but not free leaf wetness. Sporulation occurs at 10 to 26°C. At the optimum temperature of 18°C, the process of sporulation requires 7.5 h at relative humidity ≥ 85%, with 3 h for sporophores emergence from stomata and 4.5 h for spore formation. Sporophores can emerge under light or darkness, but spore formation occurs in the dark only. Limited data are available on spore dispersal. Spores dispersed from sporulating plants contaminate healthy plants within 2 h of exposure. Settled spores may survive on leaf surface of healthy plants for prolonged periods, depending on temperature. Seed transmission of the disease occurs in Europe, but not in Israel or the United States. P. belbahrii in Israel also attacks species belonging to Rosemarinus, Nepeta, Agastache, Micromeria, and Salvia but not Plectranthus (coleus). A Peronospora species that infects coleus does not infect sweet basil. Control of BDM includes chemical, physical, and genetic means. The fungicide mefenoxam was highly effective in controlling the disease but resistant populations were quickly selected for in Israel and Europe rendering it ineffective. A new compound oxathiapiprolin (OSBP inhibitor) is highly effective. Nocturnal illumination of basil crops controls the disease by preventing sporulation. Daytime solar heating suppressed the disease effectively by reducing spore and mycelium viability. The most effective physical means is fanning. Nocturnal fanning prevents or limits dew deposition on leaf surfaces, and as a result, infection and sporulation diminish and epidemics are prevented. Genetic resistance occurs in wild basil and its transfer to sweet basil is under way.

Substratum location and zoospore behaviour in the fouling alga Enteromorpha.

PubMed

Callow, M E; Callow, J A

2000-01-01

The green alga Enteromorpha is the most important macroalga that fouls ships, submarines and underwater structures. Major factors in its success in colonising new substrata are the production of enormous numbers of swimming spores and their ability to locate surfaces on which to settle. Factors facilitating the settlement and adhesion of asexual zoospores are examined in this article. Settlement and adhesion may be regulated by topographical, biological, chemical and physico-chemical cues, all of which are modified by the presence of microbial biofilm. The level of gregarious zoospore settlement is related to spore density and may be mediated by a number of external cues including fatty acids and 'detritus'.

Genetic Structure of Two Protist Species (Myxogastria, Amoebozoa) Suggests Asexual Reproduction in Sexual Amoebae

PubMed Central

Fiore-Donno, Anna Maria; Novozhilov, Yuri K.; Meyer, Marianne; Schnittler, Martin

2011-01-01

Plasmodial slime molds (Myxogastria or Myxomycetes) are common and widespread unicellular organisms that are commonly assumed to have a sexual life cycle culminating with the formation of often macroscopic fruiting bodies that efficiently disseminate spores. However, laboratory studies based on mating compatibility revealed the coexistence of asexual as well as sexual strains. To test this hypothesis in natural populations, we investigated the genetic variability of two species of the genus Lamproderma. Detailed ecological relevés were carried out in 2007 and 2009 in several deep ravines in the Elbsandsteingebirge (Saxony, south-eastern Germany). Morphological characters of 93 specimens of Lamproderma were recorded and genetic analyses, based on the small subunit ribosomal gene, the internal transcribed spacer 1 and partial elongation factor 1α sequences were carried out for 52 specimens. Genetic analyses showed the existence of two major clades, each composed of several discrete lineages. Most of these lineages were composed of several identical sequences (SSU, ITS 1 and EF-1α) which is explained best by an asexual mode of reproduction. Detrended Correspondence Analysis of morphological characters revealed two morphospecies that corresponded to the two major clades, except for one genotype (Lc6), thus challenging the morphospecies concept. Genetic patterns were not related to the geographical distribution: specimens belonging to the same genotype were found in distinct ravines, suggesting effective long-distance dispersal via spores, except for the Lc6 genotype which was found only in one ravine. Implications for the morphological and biological species concept are discussed. PMID:21829662

Characterization of oxylipins and dioxygenase genes in the asexual fungus Aspergillus niger

PubMed Central

2009-01-01

Background Aspergillus niger is an ascomycetous fungus that is known to reproduce through asexual spores, only. Interestingly, recent genome analysis of A. niger has revealed the presence of a full complement of functional genes related to sexual reproduction [1]. An example of such genes are the dioxygenase genes which in Aspergillus nidulans, have been shown to be connected to oxylipin production and regulation of both sexual and asexual sporulation [2-4]. Nevertheless, the presence of sex related genes alone does not confirm sexual sporulation in A. niger. Results The current study shows experimentally that A. niger produces the oxylipins 8,11-dihydroxy octadecadienoic acid (8,11-diHOD), 5,8-dihydroxy octadecadienoic acid (5,8-diHOD), lactonized 5,8-diHOD, 8-hydroxy octadecadienoic acid (8-HOD), 10-hydroxy octadecadienoic acid (10-HOD), small amounts of 8-hydroxy octadecamonoenoic acid (8-HOM), 9-hydroxy octadecadienoic acid (9-HOD) and 13-hydroxy octadecadienoic acid (13-HOD). Importantly, this study shows that the A. niger genome contains three putative dioxygenase genes, ppoA, ppoC and ppoD. Expression analysis confirmed that all three genes are indeed expressed under the conditions tested. Conclusion A. niger produces the same oxylipins and has similar dioxygenase genes as A. nidulans. Their presence could point towards the existence of sexual reproduction in A. niger or a broader role for the gene products in physiology, than just sexual development. PMID:19309517

Paleomycology of the Princeton Chert. III. Dictyosporic microfungi, Monodictysporites princetonensis gen. et sp. nov., associated with decayed rhizomes of an Eocene semi-aquatic fern.

PubMed

Klymiuk, Ashley A

2016-09-01

This study builds on previous investigations of paleomycological diversity within permineralized plants of a significant Eocene paleobotanical locality, the Princeton Chert. The fungal body fossils described here occur in decayed rhizomes of the extinct semi-aquatic fern Dennstaedtiopsis aerenchymata Fungi include vegetative hyphae throughout the plant tissue, as well as a dense assemblage of >100 dematiaceous spores. The spores occur in a discrete zone surrounding two extraneous rootlets of other plants, which penetrated the fern tissue post-mortem. Spores are obovoid and muriform, composed of 8-12 cells with constricted septa and produced from hyaline or slightly pigmented hyphae. The spores are morphologically similar to both asexual reproductive dictyospores of phylogenetically disparate microfungi attributed to the morphogenus Monodictys and perennating dictyochlamydospores that occur in the anamorph genus Phoma In addition to expanding the early Eocene fossil record for Ascomycota, these specimens also provide new insight into the rapidity of initial phases of the fossilization process in this important paleobotanical locality. © 2016 by The Mycological Society of America.

Termitomyces sp. associated with the termite Macrotermes natalensis has a heterothallic mating system and multinucleate cells.

PubMed

De Fine Licht, Henrik H; Andersen, Anders; Aanen, Duur K

2005-03-01

Fungi of the genus Termitomyces live in an obligate symbiosis with termites of the subfamily Macrotermitinae. Many species of Termitomyces frequently form fruit bodies, which develop from the fungus comb within the nest. In this study, we determined the mating system of a species of Termitomyces associated with the South African termite Macrotermes natalensis. Termite nests were excavated and a Termitomyces sp. was isolated into pure culture from the asexual fruit bodies (nodules) growing in the fungus gardens. For one strain, single basidiospore cultures were obtained from basidiomes growing from the fungus comb after incubation without termites. Using nuclear staining, we show that both comb cultures and single spore cultures have multinucleate cells and that the majority of spores has a single nucleus. However, DNA sequencing of the ITS region in the nuclear RNA gene revealed that the comb mycelium had two different ITS types that segregated in the single spore cultures, which consequently had only a single ITS type. These results unambiguously prove that the strain of Termitomyces studied here has a heterothallic mating system, with the fungus garden of the termite mound being in the heterokaryotic phase. This is the first time the mating system of a Termitomnyces species has been studied.

Storage Stability of Dried Microsclerotia of the Biological Control Pathogen Mycoleptodiscus Terrestris

DTIC Science & Technology

2009-09-01

asexual spores (sporogenic germination), or by sexual fruit bodies (carpogenic germination) (Webster and Weber 2007). Plating of dried microsclerotia...While drying the fungus does not appear to impact efficacy, it is unknown how prolonged storage might affect the viability and virulence of the organism...agar (Table 1). Warm water temperatures (25 ºC ± 1 ºC) and the presence of a host plant may have affected both germination and sporulation of the

Mate Finding, Sexual Spore Production, and the Spread of Fungal Plant Parasites.

PubMed

Hamelin, Frédéric M; Castella, François; Doli, Valentin; Marçais, Benoît; Ravigné, Virginie; Lewis, Mark A

2016-04-01

Sexual reproduction and dispersal are often coupled in organisms mixing sexual and asexual reproduction, such as fungi. The aim of this study is to evaluate the impact of mate limitation on the spreading speed of fungal plant parasites. Starting from a simple model with two coupled partial differential equations, we take advantage of the fact that we are interested in the dynamics over large spatial and temporal scales to reduce the model to a single equation. We obtain a simple expression for speed of spread, accounting for both sexual and asexual reproduction. Taking Black Sigatoka disease of banana plants as a case study, the model prediction is in close agreement with the actual spreading speed (100 km per year), whereas a similar model without mate limitation predicts a wave speed one order of magnitude greater. We discuss the implications of these results to control parasites in which sexual reproduction and dispersal are intrinsically coupled.

Bryophytes and lichens: Small but indispensable forest dwellers

USGS Publications Warehouse

Hutten, Martin; Woodward, Andrea

2002-01-01

* What is a Bryophyte? * Bryophytes are the small green plants commonly known as mosses, liverworts and hornworts. Compared to plants, they have primitive tissues for conducting food and water, and they lack a protective outer surface to maintain water balance. Most bryophytes, because they lack tissues such as roots, obtain their water through direct surface contact with their environment. During dry weather they have the capacity to withstand complete dehydration. Bryophytes that are dry may appear dead but will regain normal function when moisture is available. Instead of producing seeds, bryophytes can either reproduce sexually by means of spores, or asexually when small pieces break off and grow into new individuals. * What is a Lichen? * Lichens are dual organisms consisting of a fungus and an alga or a cyanobacterium. The fungus provides the alga with structure, protection, nutrients, and water absorbed from the atmosphere and the substrate (e.g., soil, rotten logs, tree branches). In return, the alga provides carbohydrates from photosynthesis to the fungus. Algae from some lichens grow independently of the fungus, but in lichen form, the algae can inhabit more challenging environments than when growing alone. Most lichens can reproduce asexually. They either produce specialized propagules containing both partners, or parts of the lichen simply break, allowing both the fungus and the alga to disperse together. In some lichens, the fungal partner reproduces sexually by releasing spores, but the partner alga must be present in order for a lichen to reform.

Extraction of High Molecular Weight DNA from Fungal Rust Spores for Long Read Sequencing.

PubMed

Schwessinger, Benjamin; Rathjen, John P

2017-01-01

Wheat rust fungi are complex organisms with a complete life cycle that involves two different host plants and five different spore types. During the asexual infection cycle on wheat, rusts produce massive amounts of dikaryotic urediniospores. These spores are dikaryotic (two nuclei) with each nucleus containing one haploid genome. This dikaryotic state is likely to contribute to their evolutionary success, making them some of the major wheat pathogens globally. Despite this, most published wheat rust genomes are highly fragmented and contain very little haplotype-specific sequence information. Current long-read sequencing technologies hold great promise to provide more contiguous and haplotype-phased genome assemblies. Long reads are able to span repetitive regions and phase structural differences between the haplomes. This increased genome resolution enables the identification of complex loci and the study of genome evolution beyond simple nucleotide polymorphisms. Long-read technologies require pure high molecular weight DNA as an input for sequencing. Here, we describe a DNA extraction protocol for rust spores that yields pure double-stranded DNA molecules with molecular weight of >50 kilo-base pairs (kbp). The isolated DNA is of sufficient purity for PacBio long-read sequencing, but may require additional purification for other sequencing technologies such as Nanopore and 10× Genomics.

Asexual Recombinants of Plasmopara halstedii Pathotypes from Dual Infection of Sunflower

PubMed Central

Spring, Otmar; Zipper, Reinhard

2016-01-01

Genetically homogenous strains of Plasmopara halstedii differing in host specificity and fungicide tolerance were used to test the hypothesis that asexual genetic recombination occurs and may account for the high genotype diversity of this homothallic reproducing oomycete, which causes downy mildew in sunflower. Dual inoculation of sunflower seedlings with single zoospore strains of complementary infection characteristics caused sporulation under conditions where inoculation with each strain alone failed to infect. PCR-based investigation with strain-specific primers proved the presence of genetic traits from both progenitors in single sporangia collected from sporangiophores of such infections. Sister zoospores released from these sporangia revealed the genotype of the one or the other parental strain thus indicating heterokaryology of sporangia. Moreover, some zoospores showed amplification products of both parents, which suggests that the generally mononucleic spores derived from genetic recombination. The possibility of parasexual genetic exchange in the host-independent stage of infection and the evolutionary consequences are discussed. PMID:27907026

Novel aspects in the life cycle and biotrophic interactions in Pezizomycetes (Ascomycota, Fungi).

PubMed

Tedersoo, Leho; Arnold, A Elizabeth; Hansen, Karen

2013-03-01

The ascomycete class Pezizomycetes (single order Pezizales)is known for its cup-shaped fruit bodies and the evolution of edible truffles and morels, but little is known about the ontogeny and ecology of this large and ecologically diverse fungal group. In this issue of Molecular Ecology, Healy et al. (2013) make a great leap forward by describing and identifying asexual, anamorphic structures that produce mitotic spores in many ectomycorrhiza-forming truffle and nontruffle species on soil surfaces worldwide(Fig. 1). Although such anamorphic forms have been reported sporadically from certain ectomycorrhizal and saprotrophic Pezizomycetes (e.g. Warcup 1990), Healy et al. (2013) demonstrate that these terricolous asexual forms are both taxonomically and geographically more widespread and, in fact, much more common than previously understood. We anticipate that deeper insight into other substrates, provided by molecular analyses of materials such as dead wood and seeds, is likely to reveal numerous anamorphs of saprotrophic and pathogenic Pezizomycetes as well (see Marek et al. 2009).

A Qualitative Exploration of the "Coming Out" Process for Asexual Individuals.

PubMed

Robbins, Nicolette K; Low, Kathryn Graff; Query, Anna N

2016-04-01

"Coming out" is an important process not only for identity formation in sexual minorities, but also for increasing access to romantic partners of similar identities (Vaughan & Waehler, 2010). It is unclear how asexuality and the variations within the asexual community are revealed and communicated in the coming out process. Some asexual individuals may find no practical value in coming out, as they do not seek romantic partnerships, while others pursue romantic relationships that are devoid of sexual activity. To date, virtually no psychological research has explored the "coming out" experience for those with an asexual identity. The current research analyzed the "coming out" narratives of 169 self-identified asexual individuals recruited from three online asexual communities using a phenomenological approach. Salient themes were extracted from narratives about the experience of developing an asexual identity. Themes included skepticism from family and friends, lack of acceptance and misunderstanding, non-disclosure of the asexual identity, relief upon discovering the asexual community, and the role of the internet in asexual identity discovery and expression. A theoretical model of asexual identity development is proposed based on these findings.

Strain H2-419-4 of Haematococcus pluvialis induced by ethyl methanesulphonate and ultraviolet radiation

NASA Astrophysics Data System (ADS)

Sun, Yanhong; Liu, Jianguo; Zhang, Xiaoli; Lin, Wei

2008-05-01

Two strains H2-410 and H2-419 were obtained from the chemically mutated survivors of wild Haematococcus pluvialis 2 by using ethyl methanesulphonate (EMS). Strains H2-410 and H2-419 showed a fast cell growth with 13% and 20% increase in biomass compared to wild type, respectively. Then H2-419-4, a fast cell growth and high astaxanthin accumulation strain, was obtained by exposing the strain H2-419 to ultraviolet radiation (UV) further. The total biomass, the astaxanthin content per cell, astaxanthin production of H2-419-4 showed 68%, 28%, and 120% increase compared to wild H. pluvialis 2, respectively. HPLC (High Performance Liquid Chromatography) data showed also an obvious proportional variation of different carotenoid compositions in the extracts of H2-419-4 and the wild type, although no peak of carotenoids appeared or disappeared. Therefore, the main compositions in strain H2-419-4, like its wild one, were free of astaxanthin, monoester, and diester of astaxanthin. The asexual reproduction in survivors after exposed to UV was not synchronous, and different from the normal synchronous asexual reproduction as the mother cells were motile instead of non-motile. Interestingly, some survivors from UV irradiation produced many mini-spores (or gamete?), the spores moved away from the mother cell gradually 4 or 5 days later. This is quite similar to sexual reproduction described by Elliot in 1934. However, whether this was sexual reproduction remains questionable, as no mating process has been observed.

Mitogen-Activated Protein Kinase Cascade Required for Regulation of Development and Secondary Metabolism in Neurospora crassa▿

PubMed Central

Park, Gyungsoon; Pan, Songqin; Borkovich, Katherine A.

2008-01-01

Mitogen-activated protein kinase (MAPK) signaling cascades are composed of MAPK kinase kinases (MAPKKKs), MAPK kinases (MAPKKs), and MAPKs. In this study, we characterize components of a MAPK cascade in Neurospora crassa (mik-1, MAPKKK; mek-1, MAPKK; and mak-1, MAPK) homologous to that controlling cell wall integrity in Saccharomyces cerevisiae. Growth of basal hyphae is significantly reduced in mik-1, mek-1, and mak-1 deletion mutants on solid medium. All three mutants formed short aerial hyphae and the formation of asexual macroconidia was reduced in Δmik-1 mutants and almost abolished in Δmek-1 and Δmak-1 strains. In contrast, the normally rare asexual spores, arthroconidia, were abundant in cultures of the three mutants. Δmik-1, Δmek-1, and Δmak-1 mutants were unable to form protoperithecia or perithecia when used as females in a sexual cross. The MAK-1 MAPK was not phosphorylated in Δmik-1 and Δmek-1 mutants, consistent with the involvement of MIK-1, MEK-1, and MAK-1 in the same signaling cascade. Interestingly, we observed increased levels of mRNA and protein for tyrosinase in the mutants under nitrogen starvation, a condition favoring sexual differentiation. Tyrosinase is an enzyme that catalyzes production of the secondary metabolite l-DOPA melanin. These results implicate the MAK-1 pathway in regulation of development and secondary metabolism in filamentous fungi. PMID:18849472

Zygomycetes in Human Disease

PubMed Central

Ribes, Julie A.; Vanover-Sams, Carolyn L.; Baker, Doris J.

2000-01-01

The Zygomycetes represent relatively uncommon isolates in the clinical laboratory, reflecting either environmental contaminants or, less commonly, a clinical disease called zygomycosis. There are two orders of Zygomycetes containing organisms that cause human disease, the Mucorales and the Entomophthorales. The majority of human illness is caused by the Mucorales. While disease is most commonly linked to Rhizopus spp., other organisms are also associated with human infection, including Mucor, Rhizomucor, Absidia, Apophysomyces, Saksenaea, Cunninghamella, Cokeromyces, and Syncephalastrum spp. Although Mortierella spp. do cause disease in animals, there is no longer sufficient evidence to suggest that they are true human pathogens. The spores from these molds are transmitted by inhalation, via a variety of percutaneous routes, or by ingestion of spores. Human zygomycosis caused by the Mucorales generally occurs in immunocompromised hosts as opportunistic infections. Host risk factors include diabetes mellitus, neutropenia, sustained immunosuppressive therapy, chronic prednisone use, iron chelation therapy, broad-spectrum antibiotic use, severe malnutrition, and primary breakdown in the integrity of the cutaneous barrier such as trauma, surgical wounds, needle sticks, or burns. Zygomycosis occurs only rarely in immunocompetent hosts. The disease manifestations reflect the mode of transmission, with rhinocerebral and pulmonary diseases being the most common manifestations. Cutaneous, gastrointestinal, and allergic diseases are also seen. The Mucorales are associated with angioinvasive disease, often leading to thrombosis, infarction of involved tissues, and tissue destruction mediated by a number of fungal proteases, lipases, and mycotoxins. If the diagnosis is not made early, dissemination often occurs. Therapy, if it is to be effective, must be started early and requires combinations of antifungal drugs, surgical intervention, and reversal of the underlying risk factors. The Entomophthorales are closely related to the Mucorales on the basis of sexual growth by production of zygospores and by the production of coenocytic hyphae. Despite these similarities, the Entomophthorales and Mucorales have dramatically different gross morphologies, asexual reproductive characteristics, and disease manifestations. In comparison to the floccose aerial mycelium of the Mucorales, the Entomophthorales produce a compact, glabrous mycelium. The asexually produced spores of the Entomophthorales may be passively released or actively expelled into the environment. Human disease with these organisms occurs predominantly in tropical regions, with transmission occurring by implantation of spores via minor trauma such as insect bites or by inhalation of spores into the sinuses. Conidiobolus typically infects mucocutaneous sites to produce sinusitis disease, while Basidiobolus infections occur as subcutaneous mycosis of the trunk and extremities. The Entomophthorales are true pathogens, infecting primarily immunocompetent hosts. They generally do not invade blood vessels and rarely disseminate. Occasional cases of disseminated and angioinvasive disease have recently been described, primarily in immunocompromised patients, suggesting a possible emerging role for this organism as an opportunist. PMID:10756000

Inter- and intra-specific cuticle variation between amphimictic and parthenogenetic species of root-knot nematode (Meloidogyne spp.) as revealed by a bacterial parasite (Pasteuria penetrans).

PubMed

Davies, K G; Rowe, J A; Williamson, V M

2008-06-01

Specific host-parasite interactions exist between species and strains of plant parasitic root-knot nematodes and the Gram-positive bacterial hyperparasite Pasteuria penetrans. This bacterium produces endospores that adhere to the cuticle of migrating juveniles, germinate and colonise the developing female within roots. Endospore attachment of P. penetrans populations to second-stage juveniles of the root-knot nematode species Meloidogyne incognita and Meloidogyne hapla showed there were interactive differences between bacterial populations and nematode species. Infected females of M. incognita produced a few progeny which were used to establish two nematode lines from single infective juveniles encumbered with either three or 26 endospores. Single juvenile descent lines of each nematode species were produced to test whether cuticle variation was greater within M. hapla lines that reproduce by facultative meiotic parthenogenesis than within lines of M. incognita, which reproduces by obligate parthenogenesis. Assays revealed variability between broods of individual females derived from single second-stage juvenile descent lines of both M. incognita and M. hapla suggesting that progeny derived from a single individual can differ in spore adhesion in both sexual and asexual nematode species. These results suggest that special mechanisms that produced these functional differences in the cuticle surface may have evolved in both sexually and asexually reproducing nematodes as a strategy to circumvent infection by this specialised hyperparasite.

Conserved meiotic machinery in Glomus spp., a putatively ancient asexual fungal lineage.

PubMed

Halary, Sébastien; Malik, Shehre-Banoo; Lildhar, Levannia; Slamovits, Claudio H; Hijri, Mohamed; Corradi, Nicolas

2011-01-01

Arbuscular mycorrhizal fungi (AMF) represent an ecologically important and evolutionarily intriguing group of symbionts of land plants, currently thought to have propagated clonally for over 500 Myr. AMF produce multinucleate spores and may exchange nuclei through anastomosis, but meiosis has never been observed in this group. A provocative alternative for their successful and long asexual evolutionary history is that these organisms may have cryptic sex, allowing them to recombine alleles and compensate for deleterious mutations. This is partly supported by reports of recombination among some of their natural populations. We explored this hypothesis by searching for some of the primary tools for a sustainable sexual cycle--the genes whose products are required for proper completion of meiotic recombination in yeast--in the genomes of four AMF and compared them with homologs of representative ascomycete, basidiomycete, chytridiomycete, and zygomycete fungi. Our investigation used molecular and bioinformatic tools to identify homologs of 51 meiotic genes, including seven meiosis-specific genes and other "core meiotic genes" conserved in the genomes of the AMF Glomus diaphanum (MUCL 43196), Glomus irregulare (DAOM-197198), Glomus clarum (DAOM 234281), and Glomus cerebriforme (DAOM 227022). Homology of AMF meiosis-specific genes was verified by phylogenetic analyses with representative fungi, animals (Mus, Hydra), and a choanoflagellate (Monosiga). Together, these results indicate that these supposedly ancient asexual fungi may be capable of undergoing a conventional meiosis; a hypothesis that is consistent with previous reports of recombination within and across some of their populations.

Conserved Meiotic Machinery in Glomus spp., a Putatively Ancient Asexual Fungal Lineage

PubMed Central

Halary, Sébastien; Malik, Shehre-Banoo; Lildhar, Levannia; Slamovits, Claudio H.; Hijri, Mohamed; Corradi, Nicolas

2011-01-01

Arbuscular mycorrhizal fungi (AMF) represent an ecologically important and evolutionarily intriguing group of symbionts of land plants, currently thought to have propagated clonally for over 500 Myr. AMF produce multinucleate spores and may exchange nuclei through anastomosis, but meiosis has never been observed in this group. A provocative alternative for their successful and long asexual evolutionary history is that these organisms may have cryptic sex, allowing them to recombine alleles and compensate for deleterious mutations. This is partly supported by reports of recombination among some of their natural populations. We explored this hypothesis by searching for some of the primary tools for a sustainable sexual cycle—the genes whose products are required for proper completion of meiotic recombination in yeast—in the genomes of four AMF and compared them with homologs of representative ascomycete, basidiomycete, chytridiomycete, and zygomycete fungi. Our investigation used molecular and bioinformatic tools to identify homologs of 51 meiotic genes, including seven meiosis-specific genes and other “core meiotic genes” conserved in the genomes of the AMF Glomus diaphanum (MUCL 43196), Glomus irregulare (DAOM-197198), Glomus clarum (DAOM 234281), and Glomus cerebriforme (DAOM 227022). Homology of AMF meiosis-specific genes was verified by phylogenetic analyses with representative fungi, animals (Mus, Hydra), and a choanoflagellate (Monosiga). Together, these results indicate that these supposedly ancient asexual fungi may be capable of undergoing a conventional meiosis; a hypothesis that is consistent with previous reports of recombination within and across some of their populations. PMID:21876220

The putative guanine nucleotide exchange factor RicA mediates upstream signaling for growth and development in Aspergillus.

PubMed

Kwon, Nak-Jung; Park, Hee-Soo; Jung, Seunho; Kim, Sun Chang; Yu, Jae-Hyuk

2012-11-01

Heterotrimeric G proteins (G proteins) govern growth, development, and secondary metabolism in various fungi. Here, we characterized ricA, which encodes a putative GDP/GTP exchange factor for G proteins in the model fungus Aspergillus nidulans and the opportunistic human pathogen Aspergillus fumigatus. In both species, ricA mRNA accumulates during vegetative growth and early developmental phases, but it is not present in spores. The deletion of ricA results in severely impaired colony growth and the total (for A. nidulans) or near (for A. fumigatus) absence of asexual sporulation (conidiation). The overexpression (OE) of the A. fumigatus ricA gene (AfricA) restores growth and conidiation in the ΔAnricA mutant to some extent, indicating partial conservation of RicA function in Aspergillus. A series of double mutant analyses revealed that the removal of RgsA (an RGS protein of the GanB Gα subunit), but not sfgA, flbA, rgsB, or rgsC, restored vegetative growth and conidiation in ΔAnricA. Furthermore, we found that RicA can physically interact with GanB in yeast and in vitro. Moreover, the presence of two copies or OE of pkaA suppresses the profound defects caused by ΔAnricA, indicating that RicA-mediated growth and developmental signaling is primarily through GanB and PkaA in A. nidulans. Despite the lack of conidiation, brlA and vosA mRNAs accumulated to normal levels in the ΔricA mutant. In addition, mutants overexpressing fluG or brlA (OEfluG or OEbrlA) failed to restore development in the ΔAnricA mutant. These findings suggest that the commencement of asexual development requires unknown RicA-mediated signaling input in A. nidulans.

The Putative Guanine Nucleotide Exchange Factor RicA Mediates Upstream Signaling for Growth and Development in Aspergillus

PubMed Central

Kwon, Nak-Jung; Park, Hee-Soo; Jung, Seunho; Kim, Sun Chang

2012-01-01

Heterotrimeric G proteins (G proteins) govern growth, development, and secondary metabolism in various fungi. Here, we characterized ricA, which encodes a putative GDP/GTP exchange factor for G proteins in the model fungus Aspergillus nidulans and the opportunistic human pathogen Aspergillus fumigatus. In both species, ricA mRNA accumulates during vegetative growth and early developmental phases, but it is not present in spores. The deletion of ricA results in severely impaired colony growth and the total (for A. nidulans) or near (for A. fumigatus) absence of asexual sporulation (conidiation). The overexpression (OE) of the A. fumigatus ricA gene (AfricA) restores growth and conidiation in the ΔAnricA mutant to some extent, indicating partial conservation of RicA function in Aspergillus. A series of double mutant analyses revealed that the removal of RgsA (an RGS protein of the GanB Gα subunit), but not sfgA, flbA, rgsB, or rgsC, restored vegetative growth and conidiation in ΔAnricA. Furthermore, we found that RicA can physically interact with GanB in yeast and in vitro. Moreover, the presence of two copies or OE of pkaA suppresses the profound defects caused by ΔAnricA, indicating that RicA-mediated growth and developmental signaling is primarily through GanB and PkaA in A. nidulans. Despite the lack of conidiation, brlA and vosA mRNAs accumulated to normal levels in the ΔricA mutant. In addition, mutants overexpressing fluG or brlA (OEfluG or OEbrlA) failed to restore development in the ΔAnricA mutant. These findings suggest that the commencement of asexual development requires unknown RicA-mediated signaling input in A. nidulans. PMID:23002107

ICTV Virus Taxonomy Profile: Chrysoviridae.

PubMed

Ghabrial, Said A; Castón, José R; Coutts, Robert H A; Hillman, Bradley I; Jiang, Daohong; Kim, Dae-Hyun; Moriyama, Hiromitsu; Ictv Report Consortium

2018-01-01

The Chrysoviridae is a family of small, isometric, non-enveloped viruses (40 nm in diameter) with segmented dsRNA genomes (typically four segments). The genome segments are individually encapsidated and together comprise 11.5-12.8 kbp. The single genus Chrysovirus includes nine species. Chrysoviruses lack an extracellular phase to their life cycle; they are transmitted via intracellular routes within an individual during hyphal growth, in asexual or sexual spores, or between individuals via hyphal anastomosis. There are no known natural vectors for chrysoviruses. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Chrysoviridae, which is available at www.ictv.global/report/chrysoviridae.

Sex-specific gene expression during asexual development of Neurospora crassa.

PubMed

Wang, Zheng; Kin, Koryu; López-Giráldez, Francesc; Johannesson, Hanna; Townsend, Jeffrey P

2012-07-01

The impact of loci that determine sexual identity upon the asexual, dominant stage of fungal life history has been well studied. To investigate their impact, expression differences between strains of different mating type during asexual development were assayed, with RNA sampled from otherwise largely isogenic mat A and mat a strains of Neurospora crassa at early, middle, and late clonal stages of development. We observed significant differences in overall gene expression between mating types across clonal development, especially at late development stages. The expression levels of mating-type genes and pheromone genes were assayed by reverse transcription and quantitative PCR, revealing expression of pheromone and receptor genes in strains of both mating types in all development stages, and revealing that mating type (mat) genes were increasingly expressed over the course of asexual development. Interestingly, among differentially expressed genes, the mat A genotype more frequently exhibited a higher expression level than mat a, and demonstrated greater transcriptional regulatory dynamism. Significant up-regulation of expression was observed for many late light-responsive genes at late asexual development stages. Further investigation of the impact of light and the roles of light response genes in asexual development of both mating types are warranted. Copyright © 2012 Elsevier Inc. All rights reserved.

Understanding asexual identity as a means to facilitate culturally competent care: A systematic literature review.

PubMed

Jones, Catriona; Hayter, Mark; Jomeen, Julie

2017-12-01

To provide a contemporary overview of asexuality and the implications this has for healthcare practice. Individuals belonging to sexual minority groups face many barriers in accessing appropriate health care. The term "sexual minority group" is usually used to refer to lesbian women, gay, bisexual and transgender individuals. Anecdotal and research evidence suggests that those who identify as asexual have similar poor experiences. Systematic review and qualitative analysis. This work uses a systematic review and qualitative analysis of the existing interview data from self-identified asexuals, to construct features of the asexual identity. The findings will help practitioners and health professionals develop an understanding of this poorly understood construct. Ultimately this work is aimed at facilitating culturally competent care in the context of asexuality. Qualitative analysis produced three themes, which can be used, not only to frame asexuality in a positive and normalising way, but also to provide greater understanding of asexuality, "romantic differences coupled with sexual indifference," "validation through engagement with asexual communities" and "a diversity of subasexual identities." Having some understanding of what it means to identify as asexual, and respecting the choices made by asexuals can markedly improve the experiences of those who embrace an asexual identity when engaging with health care. Anecdotal evidence, taken from one of the largest asexual online forums, suggests that a number of self-identified asexuals choose not to disclose their identity to healthcare professionals through fear of their asexual status being pathologised, problematised or judged. Given that asexuality is a poorly understood concept, this may be due to lack of understanding on behalf of healthcare providers. The review provides health professionals and practitioners working in clinical settings with some insights of the features of an asexual identity to facilitate culturally competent care. © 2017 John Wiley & Sons Ltd.

Two members of the Ustilago maydis velvet family influence teliospore development and virulence on maize seedlings.

PubMed

Karakkat, Brijesh B; Gold, Scott E; Covert, Sarah F

2013-12-01

Members of the fungal-specific velvet protein family regulate sexual and asexual spore production in the Ascomycota. We predicted, therefore, that velvet homologs in the basidiomycetous plant pathogen Ustilago maydis would regulate sexual spore development, which is also associated with plant disease progression in this fungus. To test this hypothesis, we studied the function of three U. maydis velvet genes, umv1, umv2 and umv3. Using a gene replacement strategy, deletion mutants were made in all three genes in compatible haploid strains, and additionally for umv1 and umv2 in the solopathogenic strain, SG200. None of the mutants showed novel morphological phenotypes during yeast-like, in vitro growth. However, the Δumv1 mutants failed to induce galls or teliospores in maize. Chlorazol black E staining of leaves infected with Δumv1 dikaryons revealed that the Δumv1 hyphae did not proliferate normally and were blocked developmentally before teliospore formation. The Δumv2 mutants were able to induce galls and teliospores in maize, but were slow to do so and thus reduced in virulence. The Δumv3 mutants were not affected in teliospore formation or disease progression. Complementation of the Δumv1 and Δumv2 mutations in the SG200 background produced disease indices similar to those of SG200. These results indicate that two U. maydis velvet family members, umv1 and umv2, are important for normal teliospore development and disease progression in maize seedlings. Copyright © 2013 Elsevier Inc. All rights reserved.

The HEX1 Gene of Fusarium graminearum Is Required for Fungal Asexual Reproduction and Pathogenesis and for Efficient Viral RNA Accumulation of Fusarium graminearum Virus 1

PubMed Central

Son, Moonil; Lee, Kyung-Mi; Yu, Jisuk; Kang, Minji; Park, Jin Man; Kwon, Sun-Jung

2013-01-01

The accumulation of viral RNA depends on many host cellular factors. The hexagonal peroxisome (Hex1) protein is a fungal protein that is highly expressed when the DK21 strain of Fusarium graminearum virus 1 (FgV1) infects its host, and Hex1 affects the accumulation of FgV1 RNA. The Hex1 protein is the major constituent of the Woronin body (WB), which is a peroxisome-derived electron-dense core organelle that seals the septal pore in response to hyphal wounding. To clarify the role of Hex1 and the WB in the relationship between FgV1 and Fusarium graminearum, we generated targeted gene deletion and overexpression mutants. Although neither HEX1 gene deletion nor overexpression substantially affected vegetative growth, both changes reduced the production of asexual spores and reduced virulence on wheat spikelets in the absence of FgV1 infection. However, the vegetative growth of deletion and overexpression mutants was increased and decreased, respectively, upon FgV1 infection compared to that of an FgV1-infected wild-type isolate. Viral RNA accumulation was significantly decreased in deletion mutants but was significantly increased in overexpression mutants compared to the viral RNA accumulation in the virus-infected wild-type control. Overall, these data indicate that the HEX1 gene plays a direct role in the asexual reproduction and virulence of F. graminearum and facilitates viral RNA accumulation in the FgV1-infected host fungus. PMID:23864619

The effect of newly induced mutations on the fitness of genotypes and populations of yeast (Saccharomyces cerevisiae).

PubMed

Orthen, E; Lange, P; Wöhrmann, K

1984-12-01

This paper analyses the fate of artificially induced mutations and their importance to the fitness of populations of the yeast, Saccharomyces cerevisiae, an increasingly important model organism in population genetics. Diploid strains, treated with UV and EMS, were cultured asexually for approximately 540 generations and under conditions where the asexual growth was interrupted by a sexual phase. Growth rates of 100 randomly sampled diploid clones were estimated at the beginning and at the end of the experiment. After the induction of sporulation the growth rates of 100 randomly sampled spores were measured. UV and EMS treatment decreases the average growth rate of the clones significantly but increases the variability in comparison to the untreated control. After selection over approximately 540 generations, variability in growth rates was reduced to that of the untreated control. No increase in mean population fitness was observed. However, the results show that after selection there still exists a large amount of hidden genetic variability in the populations which is revealed when the clones are cultivated in environments other than those in which selection took place. A sexual phase increased the reduction of the induced variability.

Deletion and Complementation of the Mating Type (MAT) Locus of the Wheat Head Blight Pathogen Gibberella zeae

PubMed Central

Desjardins, A. E.; Brown, D. W.; Yun, S.-H.; Proctor, R. H.; Lee, T.; Plattner, R. D.; Lu, S.-W.; Turgeon, B. G.

2004-01-01

Gibberella zeae, a self-fertile, haploid filamentous ascomycete, causes serious epidemics of wheat (Triticum aestivum) head blight worldwide and contaminates grain with trichothecene mycotoxins. Anecdotal evidence dating back to the late 19th century indicates that G. zeae ascospores (sexual spores) are a more important inoculum source than are macroconidia (asexual spores), although the fungus can produce both during wheat head blight epidemics. To develop fungal strains to test this hypothesis, the entire mating type (MAT1) locus was deleted from a self-fertile (MAT1-1/MAT1-2), virulent, trichothecene-producing wild-type strain of G. zeae. The resulting MAT deletion (mat1-1/mat1-2) strains were unable to produce perithecia or ascospores and appeared to be unable to mate with the fertile strain from which they were derived. Complementation of a MAT deletion strain by transformation with a copy of the entire MAT locus resulted in recovery of production of perithecia and ascospores. MAT deletion strains and MAT-complemented strains retained the ability to produce macroconidia that could cause head blight, as assessed by direct injection into wheat heads in greenhouse tests. Availability of MAT-null and MAT-complemented strains provides a means to determine the importance of ascospores in the biology of G. zeae and perhaps to identify novel approaches to control wheat head blight. PMID:15066842

Proteomic analyses reveal the key roles of BrlA and AbaA in biogenesis of gliotoxin in Aspergillus fumigatus.

PubMed

Shin, Kwang-Soo; Kim, Young Hwan; Yu, Jae-Hyuk

2015-07-31

The opportunistic human pathogenic fungus Aspergillus fumigatus primarily reproduces by forming a large number of asexual spores (conidia). Sequential activation of the central regulators BrlA, AbaA and WetA is necessary for the fungus to undergo asexual development. In this study, to address the presumed roles of these key developmental regulators during proliferation of the fungus, we analyzed and compared the proteomes of vegetative cells of wild type (WT) and individual mutant strains. Approximately 1300 protein spots were detectable from 2-D electrophoresis gels. Among these, 13 proteins exhibiting significantly altered accumulation levels were further identified by ESI-MS/MS. Markedly, we found that the GliM and GliT proteins associated with gliotoxin (GT) biosynthesis and self-protection of the fungus from GT were significantly down-regulated in the ΔabaA and ΔbrlA mutants. Moreover, mRNA levels of other GT biosynthetic genes including gliM, gliP, gliT, and gliZ were significantly reduced in both mutant strains, and no and low levels of GT were detectable in the ΔbrlA and ΔabaA mutant strains, respectively. As GliT is required for the protection of the fungus from GT, growth of the ΔbrlA mutant with reduced levels of GliT was severely impaired by exogenous GT. Our studies demonstrate that AbaA and BrlA positively regulate expression of the GT biosynthetic gene cluster in actively growing vegetative cells, and likely bridge morphological and chemical development during the life-cycle of A. fumigatus. Copyright © 2015 Elsevier Inc. All rights reserved.

Gene discovery in EST sequences from the wheat leaf rust fungus Puccinia triticina sexual spores, asexual spores and haustoria, compared to other rust and corn smut fungi

PubMed Central

2011-01-01

Background Rust fungi are biotrophic basidiomycete plant pathogens that cause major diseases on plants and trees world-wide, affecting agriculture and forestry. Their biotrophic nature precludes many established molecular genetic manipulations and lines of research. The generation of genomic resources for these microbes is leading to novel insights into biology such as interactions with the hosts and guiding directions for breakthrough research in plant pathology. Results To support gene discovery and gene model verification in the genome of the wheat leaf rust fungus, Puccinia triticina (Pt), we have generated Expressed Sequence Tags (ESTs) by sampling several life cycle stages. We focused on several spore stages and isolated haustorial structures from infected wheat, generating 17,684 ESTs. We produced sequences from both the sexual (pycniospores, aeciospores and teliospores) and asexual (germinated urediniospores) stages of the life cycle. From pycniospores and aeciospores, produced by infecting the alternate host, meadow rue (Thalictrum speciosissimum), 4,869 and 1,292 reads were generated, respectively. We generated 3,703 ESTs from teliospores produced on the senescent primary wheat host. Finally, we generated 6,817 reads from haustoria isolated from infected wheat as well as 1,003 sequences from germinated urediniospores. Along with 25,558 previously generated ESTs, we compiled a database of 13,328 non-redundant sequences (4,506 singlets and 8,822 contigs). Fungal genes were predicted using the EST version of the self-training GeneMarkS algorithm. To refine the EST database, we compared EST sequences by BLASTN to a set of 454 pyrosequencing-generated contigs and Sanger BAC-end sequences derived both from the Pt genome, and to ESTs and genome reads from wheat. A collection of 6,308 fungal genes was identified and compared to sequences of the cereal rusts, Puccinia graminis f. sp. tritici (Pgt) and stripe rust, P. striiformis f. sp. tritici (Pst), and poplar leaf rust Melampsora species, and the corn smut fungus, Ustilago maydis (Um). While extensive homologies were found, many genes appeared novel and species-specific; over 40% of genes did not match any known sequence in existing databases. Focusing on spore stages, direct comparison to Um identified potential functional homologs, possibly allowing heterologous functional analysis in that model fungus. Many potentially secreted protein genes were identified by similarity searches against genes and proteins of Pgt and Melampsora spp., revealing apparent orthologs. Conclusions The current set of Pt unigenes contributes to gene discovery in this major cereal pathogen and will be invaluable for gene model verification in the genome sequence. PMID:21435244

Shifting from clonal to sexual reproduction in aphids: physiological and developmental aspects.

PubMed

Le Trionnaire, Gaël; Hardie, Jim; Jaubert-Possamai, Stéphanie; Simon, Jean-Christophe; Tagu, Denis

2008-08-01

Developmental biology is one of the fastest growing and fascinating research fields in life sciences. Among the wide range of embryonic development, a fundamental difference exists between organisms with sexual or asexual development. Aphids are unusual organisms which display alternative pathways of sexual and asexual development, the orientation of the pathway being determined by environmental conditions. These insects offer an adapted system in which to study developmental plasticity, because a side-by-side comparison of sexual and asexual development can be made in individuals with the same genotype. In this review, we describe the developmental mechanisms that have evolved in aphids for alternative sexual and asexual reproduction. In particular, we discuss how environmental cues orientate the reproductive mode of aphids from signal perception to endocrine regulation, and propose a comparative analysis of sexual and asexual gametogenesis and embryogenesis, which has been possible due to the development of molecular methods. As a result of the recent development of genomic resources in aphids, we expect these species will permit major advances in the study of the genomic basis underlying the choice of developmental fate and multiple reproduction strategies.

FgBud3, a Rho4-Interacting Guanine Nucleotide Exchange Factor, Is Involved in Polarity Growth, Cell Division and Pathogenicity of Fusarium graminearum.

PubMed

Zhang, Chengkang; Luo, Zenghong; He, Dongdong; Su, Li; Yin, Hui; Wang, Guo; Liu, Hong; Rensing, Christopher; Wang, Zonghua

2018-01-01

Rho GTPases are signaling macromolecules that are associated with developmental progression and pathogenesis of Fusarium graminearum . Generally, enzymatic activities of Rho GTPases are regulated by Rho GTPase guanine nucleotide exchange factors (RhoGEFs). In this study, we identified a putative RhoGEF encoding gene ( FgBUD3 ) in F. graminearum database and proceeded further by using a functional genetic approach to generate FgBUD3 targeted gene deletion mutant. Phenotypic analysis results showed that the deletion of FgBUD3 caused severe reduction in growth of FgBUD3 mutant generated during this study. We also observed that the deletion of FgBUD3 completely abolished sexual reproduction and triggered the production of abnormal asexual spores with nearly no septum in ΔFgbud3 strain. Further results obtained from infection assays conducted during this research revealed that the FgBUD3 defective mutant lost its pathogenicity on wheat and hence, suggests FgBud3 plays an essential role in the pathogenicity of F. graminearum . Additional, results derived from yeast two-hybrid assays revealed that FgBud3 strongly interacted with FgRho4 compared to the interaction with FgRho2, FgRho3, and FgCdc42. Moreover, we found that FgBud3 interacted with both GTP-bound and GDP-bound form of FgRho4. From these results, we subsequently concluded that, the Rho4-interacting GEF protein FgBud3 crucially promotes vegetative growth, asexual and sexual development, cell division and pathogenicity in F. graminearum .

The FgVps39-FgVam7-FgSso1 Complex Mediates Vesicle Trafficking and Is Important for the Development and Virulence of Fusarium graminearum.

PubMed

Li, Bing; Liu, Luping; Li, Ying; Dong, Xin; Zhang, Haifeng; Chen, Huaigu; Zheng, Xiaobo; Zhang, Zhengguang

2017-05-01

Vesicle trafficking is an important event in eukaryotic organisms. Many proteins and lipids transported between different organelles or compartments are essential for survival. These processes are mediated by soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins, Rab-GTPases, and multisubunit tethering complexes such as class C core vacuole or endosome tethering and homotypic fusion or vacuole protein sorting (HOPS). Our previous study has demonstrated that FgVam7, which encodes a SNARE protein involving in vesicle trafficking, plays crucial roles in growth, asexual or sexual development, deoxynivalenol production, and pathogenicity in Fusarium graminearum. Here, the affinity purification approach was used to identify FgVam7-interacting proteins to explore its regulatory mechanisms during vesicle trafficking. The orthologs of yeast Vps39, a HOPS tethering complex subunit, and Sso1, a SNARE protein localized to the vacuole or endosome, were identified and selected for further characterization. In yeast two-hybrid and glutathione-S-transferase pull-down assays, FgVam7, FgVps39, and FgSso1 interacted with each other as a complex. The ∆Fgvps39 mutant generated by targeted deletion was significantly reduced in vegetative growth and asexual development. It failed to produce sexual spores and was defective in plant infection and deoxynivalenol production. Further cellular localization and cytological examinations suggested that FgVps39 is involved in vesicle trafficking from early or late endosomes to vacuoles in F. graminearum. Additionally, the ∆Fgvps39 mutant was defective in vacuole morphology and autophagy, and it was delayed in endocytosis. Our results demonstrate that FgVam7 interacts with FgVps39 and FgSso1 to form a unique complex, which is involved in vesicle trafficking and modulating the proper development of infection-related morphogenesis in F. graminearum.

Compulsory sexuality.

PubMed

Emens, Elizabeth F

2014-02-01

Asexuality is an emerging identity category that challenges the common assumption that everyone is defined by some type of sexual attraction. Asexuals--those who report feeling no sexual attraction to others--constitute one percent of the population, according to one prominent study. In recent years, some individuals have begun to identify as asexual and to connect around their experiences interacting with a sexual society. Asexuality has also become a protected classification under the antidiscrimination law of one state and several localities, but legal scholarship has thus far neglected the subject. This Article introduces asexuality to the legal literature as a category of analysis, an object of empirical study, and a phenomenon of medical science. It then offers a close examination of the growing community of self-identified asexuals. Asexual identity has revealing intersections with the more familiar categories of gender, sexual orientation, and disability, and inspires new models for understanding sexuality. Thinking about asexuality also sheds light on our legal system. Ours is arguably a sexual law, predicated on the assumption that sex is important. This Article uses asexuality to develop a framework for identifying the ways that law privileges sexuality. Across various fields, these interactions include legal requirements of sexual activity, special carve-outs to shield sexuality from law, legal protections from others' sexuality, and legal protections for sexual identity. Applying this framework, the Article traces several ways that our sexual law burdens, and occasionally benefits, asexuals. This Article concludes by closely examining asexuality's prospects for broader inclusion into federal, state, and local antidiscrimination laws.

Asexuality Development among Middle Aged and Older Men

PubMed Central

Huang, Yan-Ping; Chen, Bin; Ping, Ping; Wang, Hong-Xiang; Hu, Kai; Yang, Hao; Zhang, Tao; Feng, Tan; Jin, Yan; Han, Yin-Fa; Wang, Yi-Xin; Huang, Yi-Ran

2014-01-01

Objectives To assess erectile function in middle-aged and older men with asexuality status and further analyze their specific reasons for this condition. Subjects and Methods Men who had regular sexual intercourse attempts (sex frequency≥1 time per month) were classified into mild erectile dysfunction (ED), moderate to severe ED and non-ED according to International Index of Erectile Function-5, and men having no sexual intercourse attempts for at least 6 months were defined as having an asexuality status. The risk factors associated with ED were collected in a sample of 1,531 Chinese men aged 40 to 80 years, and the self-report reasons for asexuality were recorded in asexual cohort individually. Comparative analyses and multivariate regression models were conducted among these groups. Results The prevalence rates of ED and asexuality status were 49.9% and 37.2%. The asexuality status group had higher risk factors than the moderate to severe ED group in terms of old age (age≥65, adjusted odds ratio (OR) 17.69 versus (Vs.) 7.19), diabetes (crude OR: 2.40 Vs. 2.36) and hypertension (crude OR: 1.78 Vs. 1.72). The specific reasons for the asexuality status were “erectile difficulty” (52.9%), “do not care about sexuality” (53.5%)”, “no longer necessary to have sexuality at this age” (47.7%), “severe stress” (44.4%), “severe fatigue” (26.3%) and “masturbation” (26.9%). Conclusions Men with an asexual status suffer from higher risk factors for ED than men with moderate to severe ED. The majority of this asexual status could be attributed to a full ED, although the reasons for this transient asexuality also involved sexual attitudes and interests, sexual partners and masturbation. PMID:24667838

Transcriptional Changes in the Transition from Vegetative Cells to Asexual Development in the Model Fungus Aspergillus nidulans

PubMed Central

Garzia, Aitor; Etxebeste, Oier; Rodríguez-Romero, Julio; Fischer, Reinhard; Espeso, Eduardo A.

2013-01-01

Morphogenesis encompasses programmed changes in gene expression that lead to the development of specialized cell types. In the model fungus Aspergillus nidulans, asexual development involves the formation of characteristic cell types, collectively known as the conidiophore. With the aim of determining the transcriptional changes that occur upon induction of asexual development, we have applied massive mRNA sequencing to compare the expression pattern of 19-h-old submerged vegetative cells (hyphae) with that of similar hyphae after exposure to the air for 5 h. We found that the expression of 2,222 (20.3%) of the predicted 10,943 A. nidulans transcripts was significantly modified after air exposure, 2,035 being downregulated and 187 upregulated. The activation during this transition of genes that belong specifically to the asexual developmental pathway was confirmed. Another remarkable quantitative change occurred in the expression of genes involved in carbon or nitrogen primary metabolism. Genes participating in polar growth or sexual development were transcriptionally repressed, as were those belonging to the HogA/SakA stress response mitogen-activated protein (MAP) kinase pathway. We also identified significant expression changes in several genes purportedly involved in redox balance, transmembrane transport, secondary metabolite production, or transcriptional regulation, mainly binuclear-zinc cluster transcription factors. Genes coding for these four activities were usually grouped in metabolic clusters, which may bring regulatory implications for the induction of asexual development. These results provide a blueprint for further stage-specific gene expression studies during conidiophore development. PMID:23264642

Loss of sexual recombination and segregation is associated with increased diversification in evening primroses.

PubMed

Johnson, Marc T J; Fitzjohn, Richard G; Smith, Stacey D; Rausher, Mark D; Otto, Sarah P

2011-11-01

The loss of sexual recombination and segregation in asexual organisms has been portrayed as an irreversible process that commits asexually reproducing lineages to reduced diversification. We test this hypothesis by estimating rates of speciation, extinction, and transition between sexuality and functional asexuality in the evening primroses. Specifically, we estimate these rates using the recently developed BiSSE (Binary State Speciation and Extinction) phylogenetic comparative method, which employs maximum likelihood and Bayesian techniques. We infer that net diversification rates (speciation minus extinction) in functionally asexual evening primrose lineages are roughly eight times faster than diversification rates in sexual lineages, largely due to higher speciation rates in asexual lineages. We further reject the hypothesis that a loss of recombination and segregation is irreversible because the transition rate from functional asexuality to sexuality is significantly greater than zero and in fact exceeded the reverse rate. These results provide the first empirical evidence in support of the alternative theoretical prediction that asexual populations should instead diversify more rapidly than sexual populations because they are free from the homogenizing effects of sexual recombination and segregation. Although asexual reproduction may often constrain adaptive evolution, our results show that the loss of recombination and segregation need not be an evolutionary dead end in terms of diversification of lineages. © 2011 The Author(s). Evolution© 2011 The Society for the Study of Evolution.

Development of crossbreeding high-yield-potential strains for commercial cultivation in the medicinal mushroom Wolfiporia cocos (Higher Basidiomycetes).

PubMed

Xiang, Xiaozhao; Wang, Xiaoxia; Bian, Yinbing; Xu, Zhangyi

2016-07-01

Wolfiporia cocos is a well-known medicinal mushroom, and its dried sclerotia has been widely used as a traditional medicine in China, Japan, and other Asian countries for centuries. However, long-term asexual reproduction of the breeding system in W. cocos results in a current universal degeneration of cultivated strains. To develop a W. cocos breeding program that will benefit commercial cultivation, we previously developed an optimum method for indoor induction of W. cocos fruiting bodies and clarified the nature of preponderant binuclear sexual basidiospores. In this paper, we first show that the majority of W. cocos single-spore isolates cannot form sclerotium in field cultivation. We then investigated the possibility of breeding new strains by crossbreeding. Three types of mating reactions were observed in both intra-strain pairings and inter-strain pairings, and a total of fifty-five hybrids were selected by antagonistic testing and allele-specific polymerase chain reaction (PCR). Field cultivation of hybrids demonstrated that some hybrids can form sclerotium via two cultivated methods. Two new high-yield strains were identified. This report will stimulate new thinking on W. cocos and promote further extensive studies on crossbreeding in W. cocos, a new topic related to the development of more efficient protocols for the discrimination of hybrids in W. cocos.

Genome Sequencing and Mapping Reveal Loss of Heterozygosity as a Mechanism for Rapid Adaptation in the Vegetable Pathogen Phytophthora capsici

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lamour, Kurt H.; Mudge, Joann; Gobena, Daniel

2012-02-07

The oomycete vegetable pathogen Phytophthora capsici has shown remarkable adaptation to fungicides and new hosts. Like other members of this destructive genus, P. capsici has an explosive epidemiology, rapidly producing massive numbers of asexual spores on infected hosts. In addition, P. capsici can remain dormant for years as sexually recombined oospores, making it difficult to produce crops at infested sites, and allowing outcrossing populations to maintain significant genetic variation. Genome sequencing, development of a high-density genetic map, and integrative genomic or genetic characterization of P. capsici field isolates and intercross progeny revealed significant mitotic loss of heterozygosity (LOH) in diversemore » isolates. LOH was detected in clonally propagated field isolates and sexual progeny, cumulatively affecting >30percent of the genome. LOH altered genotypes for more than 11,000 single-nucleotide variant sites and showed a strong association with changes in mating type and pathogenicity. Overall, it appears that LOH may provide a rapid mechanism for fixing alleles and may be an important component of adaptability for P. capsici.« less

Mutational activation of a Galphai causes uncontrolled proliferation of aerial hyphae and increased sensitivity to heat and oxidative stress in Neurospora crassa.

PubMed Central

Yang, Q; Borkovich, K A

1999-01-01

Heterotrimeric G proteins, consisting of alpha, beta, and gamma subunits, transduce environmental signals through coupling to plasma membrane-localized receptors. We previously reported that the filamentous fungus Neurospora crassa possesses a Galpha protein, GNA-1, that is a member of the Galphai superfamily. Deletion of gna-1 leads to defects in apical extension, differentiation of asexual spores, sensitivity to hyperosmotic media, and female fertility. In addition, Deltagna-1 strains have lower intracellular cAMP levels under conditions that promote morphological abnormalities. To further define the function of GNA-1 in signal transduction in N. crassa, we examined properties of strains with mutationally activated gna-1 alleles (R178C or Q204L) as the only source of GNA-1 protein. These mutations are predicted to inhibit the GTPase activity of GNA-1 and lead to constitutive signaling. In the sexual cycle, gna-1(R178C) and gna-1(Q204L) strains are female-fertile, but produce fewer and larger perithecia than wild type. During asexual development, gna-1(R178C) and gna-1(Q204L) strains elaborate abundant, long aerial hyphae, produce less conidia, and possess lower levels of carotenoid pigments in comparison to wild-type controls. Furthermore, gna-1(R178C) and gna-1(Q204L) strains are more sensitive to heat shock and exposure to hydrogen peroxide than wild-type strains, while Deltagna-1 mutants are more resistant. In contrast to Deltagna-1 mutants, gna-1(R178C) and gna-1(Q204L) strains have higher steady-state levels of cAMP than wild type. The results suggest that GNA-1 possesses several Gbetagamma-independent functions in N. crassa. We propose that GNA-1 mediates signal transduction pathway(s) that regulate aerial hyphae development and sensitivity to heat and oxidative stresses, possibly through modulation of cAMP levels. PMID:9872952

Ontology for the asexual development and anatomy of the colonial chordate Botryllus schlosseri.

PubMed

Manni, Lucia; Gasparini, Fabio; Hotta, Kohji; Ishizuka, Katherine J; Ricci, Lorenzo; Tiozzo, Stefano; Voskoboynik, Ayelet; Dauga, Delphine

2014-01-01

Ontologies provide an important resource to integrate information. For developmental biology and comparative anatomy studies, ontologies of a species are used to formalize and annotate data that are related to anatomical structures, their lineage and timing of development. Here, we have constructed the first ontology for anatomy and asexual development (blastogenesis) of a bilaterian, the colonial tunicate Botryllus schlosseri. Tunicates, like Botryllus schlosseri, are non-vertebrates and the only chordate taxon species that reproduce both sexually and asexually. Their tadpole larval stage possesses structures characteristic of all chordates, i.e. a notochord, a dorsal neural tube, and gill slits. Larvae settle and metamorphose into individuals that are either solitary or colonial. The latter reproduce both sexually and asexually and these two reproductive modes lead to essentially the same adult body plan. The Botryllus schlosseri Ontology of Development and Anatomy (BODA) will facilitate the comparison between both types of development. BODA uses the rules defined by the Open Biomedical Ontologies Foundry. It is based on studies that investigate the anatomy, blastogenesis and regeneration of this organism. BODA features allow the users to easily search and identify anatomical structures in the colony, to define the developmental stage, and to follow the morphogenetic events of a tissue and/or organ of interest throughout asexual development. We invite the scientific community to use this resource as a reference for the anatomy and developmental ontology of B. schlosseri and encourage recommendations for updates and improvements.

Ontology for the Asexual Development and Anatomy of the Colonial Chordate Botryllus schlosseri

PubMed Central

Manni, Lucia; Gasparini, Fabio; Hotta, Kohji; Ishizuka, Katherine J.; Ricci, Lorenzo; Tiozzo, Stefano; Voskoboynik, Ayelet; Dauga, Delphine

2014-01-01

Ontologies provide an important resource to integrate information. For developmental biology and comparative anatomy studies, ontologies of a species are used to formalize and annotate data that are related to anatomical structures, their lineage and timing of development. Here, we have constructed the first ontology for anatomy and asexual development (blastogenesis) of a bilaterian, the colonial tunicate Botryllus schlosseri. Tunicates, like Botryllus schlosseri, are non-vertebrates and the only chordate taxon species that reproduce both sexually and asexually. Their tadpole larval stage possesses structures characteristic of all chordates, i.e. a notochord, a dorsal neural tube, and gill slits. Larvae settle and metamorphose into individuals that are either solitary or colonial. The latter reproduce both sexually and asexually and these two reproductive modes lead to essentially the same adult body plan. The Botryllus schlosseri Ontology of Development and Anatomy (BODA) will facilitate the comparison between both types of development. BODA uses the rules defined by the Open Biomedical Ontologies Foundry. It is based on studies that investigate the anatomy, blastogenesis and regeneration of this organism. BODA features allow the users to easily search and identify anatomical structures in the colony, to define the developmental stage, and to follow the morphogenetic events of a tissue and/or organ of interest throughout asexual development. We invite the scientific community to use this resource as a reference for the anatomy and developmental ontology of B. schlosseri and encourage recommendations for updates and improvements. PMID:24789338

Patterns of Asexuality in China: Sexual Activity, Sexual and Romantic Attraction, and Sexual Desire.

PubMed

Zheng, Lijun; Su, Yanchen

2018-05-01

This study examined patterns of asexuality in Chinese asexual people in terms of sexual activities, sexual/romantic attraction, and sexual desire. The sample included 227 (64 men and 163 women) asexual participants and 57 (26 men and 31 women) uncertain asexual participants recruited from social networks for asexual people. The control group included 217 (115 men and 102 women) heterosexual participants recruited from general social networks. Participants scoring 40 or higher on the Asexuality Identification Scale were classified as asexual. Asexual participants reported having less frequent masturbation, sexual intercourse experience, and sexual and romantic attraction compared to heterosexual participants. Lower sexual attraction among asexuals indicated that "people who experience little or no sexual attraction" would be a more appropriate definition of asexuality. The pattern of uncertain asexual participants' sexual/romantic attraction and sexual desire was intermediate between heterosexual and asexual participants. Asexual participants scored significantly lower on dyadic sexual desire and slightly lower on solitary sexual desire than heterosexual participants. There were significant differences in sexual activities and solitary sexual desire among romantic orientation categories. Homoromantic participants showed higher dyadic sexual desire and were more likely to engage in masturbation, indicating the heterogeneity among asexual people. The findings indicated that Chinese asexual people showed similar patterns of asexuality as in Western nations. Specifically, asexual people have little or no sexual attraction, non-partner-orientated sexual desire, and are heterogeneous in sexual activities and sexual desire. This implies similar mechanisms underlying the etiology of asexuality across cultures.

A little bit of sex matters for genome evolution in asexual plants.

PubMed

Hojsgaard, Diego; Hörandl, Elvira

2015-01-01

Genome evolution in asexual organisms is theoretically expected to be shaped by various factors: first, hybrid origin, and polyploidy confer a genomic constitution of highly heterozygous genotypes with multiple copies of genes; second, asexuality confers a lack of recombination and variation in populations, which reduces the efficiency of selection against deleterious mutations; hence, the accumulation of mutations and a gradual increase in mutational load (Muller's ratchet) would lead to rapid extinction of asexual lineages; third, allelic sequence divergence is expected to result in rapid divergence of lineages (Meselson effect). Recent transcriptome studies on the asexual polyploid complex Ranunculus auricomus using single-nucleotide polymorphisms confirmed neutral allelic sequence divergence within a short time frame, but rejected a hypothesis of a genome-wide accumulation of mutations in asexuals compared to sexuals, except for a few genes related to reproductive development. We discuss a general model that the observed incidence of facultative sexuality in plants may unmask deleterious mutations with partial dominance and expose them efficiently to purging selection. A little bit of sex may help to avoid genomic decay and extinction.

Variation in the strength of male mate choice allows long-term coexistence of sperm-dependent asexuals and their sexual hosts.

PubMed

Mee, Jonathan A; Otto, Sarah P

2010-10-01

In several asexual taxa, reproduction requires mating with related sexual species to stimulate egg development, even though genetic material is not incorporated from the sexuals (gynogenesis). In cases in which gynogens do not invest in male function, they can potentially have a twofold competitive advantage over sexuals because the asexuals avoid the cost of producing males. If unmitigated, however, the competitive success of the asexuals would ultimately lead to their own demise, following the extinction of the sexual species that stimulate egg development. We have studied a model of mate choice among sexual individuals and asexual gynogens, where males of the sexual species preferentially mate with sexual females over gynogenetic females, to determine if such mating preferences can stably maintain both gynogenetic and sexual individuals within a community. Our model shows that stable coexistence of gynogens and their sexual hosts can occur when there is variation among males in the degree of preference for mating with sexual females and when pickier males pay a higher cost of preference. © 2010 The Author(s). Journal compilation © 2010 The Society for the Study of Evolution.

Coming to an Asexual Identity: Negotiating Identity, Negotiating Desire.

PubMed

Scherrer, Kristin S

2008-10-01

Sexuality is generally considered an important aspect of self-hood. Therefore, individuals who do not experience sexual attraction, and embrace an asexual identity are in a unique position to inform the social construction of sexuality. This study explores the experiences of asexual individuals utilizing open ended Internet survey data from 102 self-identified asexual people. In this paper I describe several distinct aspects of asexual identities: the meanings of sexual, and therefore, asexual behaviors, essentialist characterizations of asexuality, and lastly, interest in romance as a distinct dimension of sexuality. These findings have implications not only for asexual identities, but also for the connections of asexuality with other marginalized sexualities.

Gene expression analysis of parthenogenetic embryonic development of the pea aphid, Acyrthosiphon pisum, suggests that aphid parthenogenesis evolved from meiotic oogenesis.

PubMed

Srinivasan, Dayalan G; Abdelhady, Ahmed; Stern, David L

2014-01-01

Aphids exhibit a form of phenotypic plasticity, called polyphenism, in which genetically identical females reproduce sexually during one part of the life cycle and asexually (via parthenogenesis) during the remainder of the life cycle. The molecular basis for aphid parthenogenesis is unknown. Cytological observations of aphid parthenogenesis suggest that asexual oogenesis evolved either through a modification of meiosis or from a mitotic process. As a test of these alternatives, we assessed the expression levels and expression patterns of canonical meiotic recombination and germline genes in the sexual and asexual ovaries of the pea aphid, Acyrthosiphon pisum. We observed expression of all meiosis genes in similar patterns in asexual and sexual ovaries, with the exception that some genes encoding Argonaute-family members were not expressed in sexual ovaries. In addition, we observed that asexual aphid tissues accumulated unspliced transcripts of Spo11, whereas sexual aphid tissues accumulated primarily spliced transcripts. In situ hybridization revealed Spo11 transcript in sexual germ cells and undetectable levels of Spo11 transcript in asexual germ cells. We also found that an obligately asexual strain of pea aphid produced little spliced Spo11 transcript. Together, these results suggest that parthenogenetic oogenesis evolved from a meiosis-like, and not a mitosis-like, process and that the aphid reproductive polyphenism may involve a modification of Spo11 gene activity.

Gene Expression Analysis of Parthenogenetic Embryonic Development of the Pea Aphid, Acyrthosiphon pisum, Suggests That Aphid Parthenogenesis Evolved from Meiotic Oogenesis

PubMed Central

Srinivasan, Dayalan G.; Abdelhady, Ahmed; Stern, David L.

2014-01-01

Aphids exhibit a form of phenotypic plasticity, called polyphenism, in which genetically identical females reproduce sexually during one part of the life cycle and asexually (via parthenogenesis) during the remainder of the life cycle. The molecular basis for aphid parthenogenesis is unknown. Cytological observations of aphid parthenogenesis suggest that asexual oogenesis evolved either through a modification of meiosis or from a mitotic process. As a test of these alternatives, we assessed the expression levels and expression patterns of canonical meiotic recombination and germline genes in the sexual and asexual ovaries of the pea aphid, Acyrthosiphon pisum. We observed expression of all meiosis genes in similar patterns in asexual and sexual ovaries, with the exception that some genes encoding Argonaute-family members were not expressed in sexual ovaries. In addition, we observed that asexual aphid tissues accumulated unspliced transcripts of Spo11, whereas sexual aphid tissues accumulated primarily spliced transcripts. In situ hybridization revealed Spo11 transcript in sexual germ cells and undetectable levels of Spo11 transcript in asexual germ cells. We also found that an obligately asexual strain of pea aphid produced little spliced Spo11 transcript. Together, these results suggest that parthenogenetic oogenesis evolved from a meiosis-like, and not a mitosis-like, process and that the aphid reproductive polyphenism may involve a modification of Spo11 gene activity. PMID:25501006

Plant Growth-Promoting Rhizobacteria Stimulate Vegetative Growth and Asexual Reproduction of Kalanchoe daigremontiana.

PubMed

Park, Yong-Soon; Park, Kyungseok; Kloepper, Joseph W; Ryu, Choong-Min

2015-09-01

Certain bacterial species associate with plant roots in soil. The plant growth-promoting rhizobacteria (PGPR) stimulate plant growth and yield in greenhouse and field. Here, we examined whether application of known bacilli PGPR strains stimulated growth and asexual reproduction in the succulent plant Kalanchoe daigremontiana. Four PGPR strains B. amyloliquefaciens IN937a, B. cereus BS107, B. pumilus INR7, and B. subtilis GB03 were applied to young plantlets by soil-drenching, and plant growth and development was monitored for three months. Aerial growth was significantly stimulated in PGPR-inoculated plants, which was observed as increases in plant height, shoot weight, and stem width. The stimulated growth influenced plant development by increasing the total number of leaves per plant. Treatment with bacilli also increased the total root biomass compared with that of control plants, and led to a 2-fold increase in asexual reproduction and plantlet formation on the leaf. Collectively, our results firstly demonstrate that Bacillus spp. promote vegetative development of K. daigremontiana, and the enhanced growth stimulates asexual reproduction and plantlet formation.

On the Genealogy of Asexual Diploids

NASA Astrophysics Data System (ADS)

Lam, Fumei; Langley, Charles H.; Song, Yun S.

Given molecular genetic data from diploid individuals that, at present, reproduce mostly or exclusively asexually without recombination, an important problem in evolutionary biology is detecting evidence of past sexual reproduction (i.e., meiosis and mating) and recombination (both meiotic and mitotic). However, currently there is a lack of computational tools for carrying out such a study. In this paper, we formulate a new problem of reconstructing diploid genealogies under the assumption of no sexual reproduction or recombination, with the ultimate goal being to devise genealogy-based tools for testing deviation from these assumptions. We first consider the infinite-sites model of mutation and develop linear-time algorithms to test the existence of an asexual diploid genealogy compatible with the infinite-sites model of mutation, and to construct one if it exists. Then, we relax the infinite-sites assumption and develop an integer linear programming formulation to reconstruct asexual diploid genealogies with the minimum number of homoplasy (back or recurrent mutation) events. We apply our algorithms on simulated data sets with sizes of biological interest.

Sexual versus Asexual Reproduction: Distinct Outcomes in Relative Abundance of Parthenogenetic Mealybugs following Recent Colonization.

PubMed

Tabata, Jun; Ichiki, Ryoko T; Tanaka, Hirotaka; Kageyama, Daisuke

2016-01-01

Asexual reproduction, including parthenogenesis in which embryos develop within a female without fertilization, is assumed to confer advantages over sexual reproduction, which includes a "cost of males." Sexual reproduction largely predominates in animals, however, indicating that this cost is outweighed by the genetic and/or ecological benefits of sexuality, including the acquisition of advantageous mutations occurring in different individuals and the elimination of deleterious mutations. But the evolution of sexual reproduction remains unclear, because we have limited examples that demonstrate the relative success of sexual lineages in the face of competition from asexual lineages in the same environment. Here we investigated a sympatric occurrence of sexual and asexual reproduction in the pineapple mealybug, Dysmicoccus brevipes. This pest invaded southwestern Japan, including Okinawa and Ishigaki Islands, in the 1930s in association with imported pineapple plants. Our recent censuses demonstrated that on Okinawa sexually reproducing individuals can coexist with and even dominate asexual individuals in the presence of habitat and resource competition, which is considered to be severe for this nearly immobile insect. Molecular phylogeny based on partial DNA sequences in the mitochondrial and nuclear genomes, as well as the endosymbiotic bacterial genome, revealed that the asexual lineage diverged from a common sexual ancestor in the relatively recent past. In contrast, only the asexual lineage exhibiting obligate apomictic thelytoky was discovered on Ishigaki. Co-existence of the two lineages cannot be explained by the results of laboratory experiments, which showed that the intrinsic rate of increase in the sexual lineage was not obviously superior to that of the asexual lineage. Differences in biotic and/or abiotic selective forces operating on the two islands might be the cause of this discrepancy. This biological system offers a unique opportunity to assess the relative success of sexual versus asexual lineages with an unusual morphology and life cycle.

Asexuality: a mixed-methods approach.

PubMed

Brotto, Lori A; Knudson, Gail; Inskip, Jess; Rhodes, Katherine; Erskine, Yvonne

2010-06-01

Current definitions of asexuality focus on sexual attraction, sexual behavior, and lack of sexual orientation or sexual excitation; however, the extent to which these definitions are accepted by self-identified asexuals is unknown. The goal of Study 1 was to examine relationship characteristics, frequency of sexual behaviors, sexual difficulties and distress, psychopathology, interpersonal functioning, and alexithymia in 187 asexuals recruited from the Asexuality Visibility and Education Network (AVEN). Asexual men (n = 54) and women (n = 133) completed validated questionnaires online. Sexual response was lower than normative data and was not experienced as distressing, and masturbation frequency in males was similar to available data for sexual men. Social withdrawal was the most elevated personality subscale; however, interpersonal functioning was in the normal range. Alexithymia was elevated in 12%. Social desirability was also in the normal range. Study 2 was designed to expand upon these quantitative findings with 15 asexuals from Study 1 through in-depth telephone interviews. The findings suggest that asexuality is best conceptualized as a lack of sexual attraction; however, asexuals varied greatly in their experience of sexual response and behavior. Asexuals partnered with sexuals acknowledged having to "negotiate" sexual activity. There were not higher rates of psychopathology among asexuals; however, a subset might fit the criteria for Schizoid Personality Disorder. There was also strong opposition to viewing asexuality as an extreme case of sexual desire disorder. Finally, asexuals were very motivated to liaise with sex researchers to further the scientific study of asexuality.

Temporal and Spatial Regulation of Gene Expression during Asexual Development of Neurospora crassa

USDA-ARS?s Scientific Manuscript database

In this study we profiled spatial and temporal transcriptional changes during asexual sporulation in the filamentous fungus Neurospora crassa. Aerial tissue was separated from the mycelium to allow detection of genes specific to each tissue. We identified 2641 genes that were differentially expresse...

Interactions between tafenoquine and artemisinin-combination therapy partner drug in asexual and sexual stage Plasmodium falciparum.

PubMed

Kemirembe, Karen; Cabrera, Mynthia; Cui, Liwang

2017-08-01

The 8-aminoquinoline tafenoquine (TFQ), a primaquine derivative, is currently in late-stage clinical development for the radical cure of P. vivax. Here drug interactions between TFQ and chloroquine and six artemisinin-combination therapy (ACT) partner drugs in P. falciparum asexual stages and gametocytes were investigated. TFQ was mostly synergistic with the ACT-partner drugs in asexual parasites regardless of genetic backgrounds. However, at fixed ratios of 1:3, 1:1 and 3:1, TFQ only interacted synergistically with naphthoquine, pyronaridine and piperaquine in gametocytes. This study indicated that TFQ and ACT-partner drugs will likely have increased potency against asexual stages of the malaria parasites, whereas some drugs may interfere with each other against the P. falciparum gametocytes. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Accelerated Mutation Accumulation in Asexual Lineages of a Freshwater Snail

PubMed Central

Neiman, Maurine; Hehman, Gery; Miller, Joseph T.; Logsdon, John M.; Taylor, Douglas R.

2010-01-01

Sexual reproduction is both extremely costly and widespread relative to asexual reproduction, meaning that it must also confer profound advantages in order to persist. One theorized benefit of sex is that it facilitates the clearance of harmful mutations, which would accumulate more rapidly in the absence of recombination. The extent to which ineffective purifying selection and mutation accumulation are direct consequences of asexuality and whether the accelerated buildup of harmful mutations in asexuals can occur rapidly enough to maintain sex within natural populations, however, remain as open questions. We addressed key components of these questions by estimating the rate of mutation accumulation in the mitochondrial genomes of multiple sexual and asexual representatives of Potamopyrgus antipodarum, a New Zealand snail characterized by mixed sexual/asexual populations. We found that increased mutation accumulation is associated with asexuality and occurs rapidly enough to be detected in recently derived asexual lineages of P. antipodarum. Our results demonstrate that increased mutation accumulation in asexuals can differentially affect coexisting and ecologically similar sexual and asexual lineages. The accelerated rate of mutation accumulation observed in asexual P. antipodarum provides some of the most direct evidence to date for a link between asexuality and mutation accumulation and implies that mutational buildup could be rapid enough to contribute to the short-term evolutionary mechanisms that favor sexual reproduction. PMID:19995828

No sex in fungus-farming ants or their crops.

PubMed

Himler, Anna G; Caldera, Eric J; Baer, Boris C; Fernández-Marín, Hermógenes; Mueller, Ulrich G

2009-07-22

Asexual reproduction imposes evolutionary handicaps on asexual species, rendering them prone to extinction, because asexual reproduction generates novel genotypes and purges deleterious mutations at lower rates than sexual reproduction. Here, we report the first case of complete asexuality in ants, the fungus-growing ant Mycocepurus smithii, where queens reproduce asexually but workers are sterile, which is doubly enigmatic because the clonal colonies of M. smithii also depend on clonal fungi for food. Degenerate female mating anatomy, extensive field and laboratory surveys, and DNA fingerprinting implicate complete asexuality in this widespread ant species. Maternally inherited bacteria (e.g. Wolbachia, Cardinium) and the fungal cultivars can be ruled out as agents inducing asexuality. M. smithii societies of clonal females provide a unique system to test theories of parent-offspring conflict and reproductive policing in social insects. Asexuality of both ant farmer and fungal crop challenges traditional views proposing that sexual farmer ants outpace coevolving sexual crop pathogens, and thus compensate for vulnerabilities of their asexual crops. Either the double asexuality of both farmer and crop may permit the host to fully exploit advantages of asexuality for unknown reasons or frequent switching between crops (symbiont reassociation) generates novel ant-fungus combinations, which may compensate for any evolutionary handicaps of asexuality in M. smithii.

Sexual Fantasy and Masturbation Among Asexual Individuals: An In-Depth Exploration.

PubMed

Yule, Morag A; Brotto, Lori A; Gorzalka, Boris B

2017-01-01

Human asexuality is generally defined as a lack of sexual attraction. We used online questionnaires to investigate reasons for masturbation, and explored and compared the contents of sexual fantasies of asexual individuals (identified using the Asexual Identification Scale) with those of sexual individuals. A total of 351 asexual participants (292 women, 59 men) and 388 sexual participants (221 women, 167 men) participated. Asexual women were significantly less likely to masturbate than sexual women, sexual men, and asexual men. Asexual women were less likely to report masturbating for sexual pleasure or fun than their sexual counterparts, and asexual men were less likely to report masturbating for sexual pleasure than sexual men. Both asexual women and men were significantly more likely than sexual women and men to report that they had never had a sexual fantasy. Of those who have had a sexual fantasy, asexual women and men were significantly more likely to endorse the response "my fantasies do not involve other people" compared to sexual participants, and consistently scored each sexual fantasy on a questionnaire as being less sexually exciting than did sexual participants. When using an open-ended format, asexual participants were more likely to report having fantasies about sexual activities that did not involve themselves, and were less likely to fantasize about topics such as group sex, public sex, and having an affair. Interestingly, there was a large amount of overlap between sexual fantasies of asexual and sexual participants. Notably, both asexual and sexual participants (both men and women) were equally likely to fantasize about topics such as fetishes and BDSM.

Agrobacterium-mediated transformation of the haploid liverwort Marchantia polymorpha L., an emerging model for plant biology.

PubMed

Ishizaki, Kimitsune; Chiyoda, Shota; Yamato, Katsuyuki T; Kohchi, Takayuki

2008-07-01

Agrobacterium-mediated transformation has not been practical in pteridophytes, bryophytes and algae to date, although it is commonly used in model plants including Arabidopsis and rice. Here we present a rapid Agrobacterium-mediated transformation system for the haploid liverwort Marchantia polymorpha L. using immature thalli developed from spores. Hundreds of hygromycin-resistant plants per sporangium were obtained by co-cultivation of immature thalli with Agrobacterium carrying the binary vector that contains a reporter, the beta-glucuronidase (GUS) gene with an intron, and a selection marker, the hygromycin phosphotransferase (hpt) gene. In this system, individual gemmae, which arise asexually from single initial cells, were analyzed as isogenic transformants. GUS activity staining showed that all hygromycin-resistant plants examined expressed the GUS transgene in planta. DNA analyses verified random integration of 1-5 copies of the intact T-DNA between the right and the left borders into the M. polymorpha genome. The efficient and rapid Agrobacterium-mediated transformation of M. polymorpha should provide molecular techniques to facilitate comparative genomics, taking advantage of this unique model plant that retains many features of the common ancestor of land plants.

Light inhibits spore germination through phytochrome in Aspergillus nidulans.

PubMed

Röhrig, Julian; Kastner, Christian; Fischer, Reinhard

2013-05-01

Aspergillus nidulans responds to light in several aspects. The balance between sexual and asexual development as well as the amount of secondary metabolites produced is controlled by light. Here, we show that germination is largely delayed by blue (450 nm), red (700 nm), and far-red light (740 nm). The largest effect was observed with far-red light. Whereas 60 % of the conidia produced a germ tube after 20 h in the dark, less than 5 % of the conidia germinated under far-red light conditions. Because swelling of conidia was not affected, light appears to act at the stage of germ-tube formation. In the absence of nutrients, far-red light even inhibited swelling of conidia, whereas in the dark, conidia did swell and germinated after prolonged incubation. The blue-light signaling components, LreA (WC-1) and LreB (WC-2), and also the cryptochrome/photolyase CryA were not required for germination inhibition. However, in the phytochrome mutant, ∆fphA, the germination delay was released, but germination was delayed in the dark in comparison to wild type. This suggests a novel function of phytochrome as far-red light sensor and as activator of polarized growth in the dark.

Genes necessary for expression of a virulence determinant and for transmission of Plasmodium falciparum are located on a 0.3-megabase region of chromosome 9.

PubMed Central

Day, K P; Karamalis, F; Thompson, J; Barnes, D A; Peterson, C; Brown, H; Brown, G V; Kemp, D J

1993-01-01

Virulence of the human malaria parasite Plasmodium falciparum is believed to relate to adhesion of parasitized erythrocytes to postcapillary venular endothelium (asexual cytoadherence). Transmission of malaria to the mosquito vector involves a switch from asexual to sexual development (gametocytogenesis). Continuous in vitro culture of P. falciparum frequently results in irreversible loss of asexual cytoadherence and gametocytogenesis. Field isolates and cloned lines differing in expression of these phenotypes were karyotyped by pulse-field gel electrophoresis. This analysis showed that expression of both phenotypes mapped to a 0.3-Mb subtelomeric deletion of chromosome 9. This deletion frequently occurs during adaptation of parasite isolates to in vitro culture. Parasites with this deletion did not express the variant surface agglutination phenotype and the putative asexual cytoadherence ligand designated P. falciparum erythrocyte membrane protein 1, which has recently been shown to undergo antigenic variation. The syntenic relationship between asexual cytoadherence and gametocytogenesis suggests that expression of these phenotypes is genetically linked. One explanation for this linkage is that both developmental pathways share a common cytoadherence mechanism. This proposed biological and genetic linkage between a virulence factor (asexual cytoadherence) and transmissibility (gametocytogenesis) would help explain why a high degree of virulence has evolved and been maintained in falciparum malaria. Images Fig. 1 Fig. 2 Fig. 3 PMID:8367496

The chastity of amoebae: re-evaluating evidence for sex in amoeboid organisms

PubMed Central

Lahr, Daniel J. G.; Parfrey, Laura Wegener; Mitchell, Edward A. D.; Katz, Laura A.; Lara, Enrique

2011-01-01

Amoebae are generally assumed to be asexual. We argue that this view is a relict of early classification schemes that lumped all amoebae together inside the ‘lower’ protozoa, separated from the ‘higher’ plants, animals and fungi. This artificial classification allowed microbial eukaryotes, including amoebae, to be dismissed as primitive, and implied that the biological rules and theories developed for macro-organisms need not apply to microbes. Eukaryotic diversity is made up of 70+ lineages, most of which are microbial. Plants, animals and fungi are nested among these microbial lineages. Thus, theories on the prevalence and maintenance of sex developed for macro-organisms should in fact apply to microbial eukaryotes, though the theories may need to be refined and generalized (e.g. to account for the variation in sexual strategies and prevalence of facultative sex in natural populations of many microbial eukaryotes). We use a revised phylogenetic framework to assess evidence for sex in several amoeboid lineages that are traditionally considered asexual, and we interpret this evidence in light of theories on the evolution of sex developed for macro-organisms. We emphasize that the limited data available for many lineages coupled with natural variation in microbial life cycles overestimate the extent of asexuality. Mapping sexuality onto the eukaryotic tree of life demonstrates that the majority of amoeboid lineages are, contrary to popular belief, anciently sexual, and that most asexual groups have probably arisen recently and independently. Additionally, several unusual genomic traits are prevalent in amoeboid lineages, including cyclic polyploidy, which may serve as alternative mechanisms to minimize the deleterious effects of asexuality. PMID:21429931

Proteomic analyses reveal the key roles of BrlA and AbaA in biogenesis of gliotoxin in Aspergillus fumigatus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shin, Kwang-Soo, E-mail: shinks@dju.kr; Kim, Young Hwan; Graduate School of Analytical Science and Technology, Chungnam National University, Daejeon, 305-764

2015-07-31

The opportunistic human pathogenic fungus Aspergillus fumigatus primarily reproduces by forming a large number of asexual spores (conidia). Sequential activation of the central regulators BrlA, AbaA and WetA is necessary for the fungus to undergo asexual development. In this study, to address the presumed roles of these key developmental regulators during proliferation of the fungus, we analyzed and compared the proteomes of vegetative cells of wild type (WT) and individual mutant strains. Approximately 1300 protein spots were detectable from 2-D electrophoresis gels. Among these, 13 proteins exhibiting significantly altered accumulation levels were further identified by ESI-MS/MS. Markedly, we found thatmore » the GliM and GliT proteins associated with gliotoxin (GT) biosynthesis and self-protection of the fungus from GT were significantly down-regulated in the ΔabaA and ΔbrlA mutants. Moreover, mRNA levels of other GT biosynthetic genes including gliM, gliP, gliT, and gliZ were significantly reduced in both mutant strains, and no and low levels of GT were detectable in the ΔbrlA and ΔabaA mutant strains, respectively. As GliT is required for the protection of the fungus from GT, growth of the ΔbrlA mutant with reduced levels of GliT was severely impaired by exogenous GT. Our studies demonstrate that AbaA and BrlA positively regulate expression of the GT biosynthetic gene cluster in actively growing vegetative cells, and likely bridge morphological and chemical development during the life-cycle of A. fumigatus. - Highlights: • Proteome analyses of WT and mutants reveal 13 differentially expressed proteins. • The GliT and GliM proteins are significantly down-regulated by ΔabaA and ΔbrlA. • Expression of other gliotoxin biosynthetic genes is lowered by ΔabaA and ΔbrlA. • Growth of ΔbrlA strain lacking GliT is completely inhibited by exogenous gliotoxin. • BrlA and AbaA play key roles in biogenesis of gliotoxin in Aspergillus fumigatus.« less

Physiological and subjective sexual arousal in self-identified asexual women.

PubMed

Brotto, Lori A; Yule, Morag A

2011-08-01

Asexuality can be defined as a lifelong lack of sexual attraction. Empirical research on asexuality reveals significantly lower self-reported sexual desire and arousal and lower rates of sexual activity; however, the speculation that there may also be an impaired psychophysiological sexual arousal response has never been tested. The aim of this study was to compare genital (vaginal pulse amplitude; VPA) and subjective sexual arousal in asexual and non-asexual women. Thirty-eight women between the ages of 19 and 55 years (10 heterosexual, 10 bisexual, 11 homosexual, and 7 asexual) viewed neutral and erotic audiovisual stimuli while VPA and self-reported sexual arousal and affect were measured. There were no significant group differences in the increased VPA and self-reported sexual arousal response to the erotic film between the groups. Asexuals showed significantly less positive affect, sensuality-sexual attraction, and self-reported autonomic arousal to the erotic film compared to the other groups; however, there were no group differences in negative affect or anxiety. Genital-subjective sexual arousal concordance was significantly positive for the asexual women and non-significant for the other three groups, suggesting higher levels of interoceptive awareness among asexuals. Taken together, the findings suggest normal subjective and physiological sexual arousal capacity in asexual women and challenge the view that asexuality should be characterized as a sexual dysfunction.

Development of an Aerosol Surface Inoculation Method for Bacillus Spores ▿

PubMed Central

Lee, Sang Don; Ryan, Shawn P.; Snyder, Emily Gibb

2011-01-01

A method was developed to deposit Bacillus subtilis spores via aerosolization onto various surface materials for biological agent decontamination and detection studies. This new method uses an apparatus coupled with a metered dose inhaler to reproducibly deposit spores onto various surfaces. A metered dose inhaler was loaded with Bacillus subtilis spores, a surrogate for Bacillus anthracis. Five different material surfaces (aluminum, galvanized steel, wood, carpet, and painted wallboard paper) were tested using this spore deposition method. This aerosolization method deposited spores at a concentration of more than 107 CFU per coupon (18-mm diameter) with less than a 50% coefficient of variation, showing that the aerosolization method developed in this study can deposit reproducible numbers of spores onto various surface coupons. Scanning electron microscopy was used to probe the spore deposition patterns on test coupons. The deposition patterns observed following aerosol impaction were compared to those of liquid inoculation. A physical difference in the spore deposition patterns was observed to result from the two different methods. The spore deposition method developed in this study will help prepare spore coupons via aerosolization fast and reproducibly for bench top decontamination and detection studies. PMID:21193670

Development of an aerosol surface inoculation method for bacillus spores.

PubMed

Lee, Sang Don; Ryan, Shawn P; Snyder, Emily Gibb

2011-03-01

A method was developed to deposit Bacillus subtilis spores via aerosolization onto various surface materials for biological agent decontamination and detection studies. This new method uses an apparatus coupled with a metered dose inhaler to reproducibly deposit spores onto various surfaces. A metered dose inhaler was loaded with Bacillus subtilis spores, a surrogate for Bacillus anthracis. Five different material surfaces (aluminum, galvanized steel, wood, carpet, and painted wallboard paper) were tested using this spore deposition method. This aerosolization method deposited spores at a concentration of more than 10(7) CFU per coupon (18-mm diameter) with less than a 50% coefficient of variation, showing that the aerosolization method developed in this study can deposit reproducible numbers of spores onto various surface coupons. Scanning electron microscopy was used to probe the spore deposition patterns on test coupons. The deposition patterns observed following aerosol impaction were compared to those of liquid inoculation. A physical difference in the spore deposition patterns was observed to result from the two different methods. The spore deposition method developed in this study will help prepare spore coupons via aerosolization fast and reproducibly for bench top decontamination and detection studies.

Spore-to-spore agar culture of the myxomycete Physarum globuliferum.

PubMed

Liu, Pu; Wang, Qi; Li, Yu

2010-02-01

The ontogeny of the myxomycete Physarum globuliferum was observed on corn meal agar and hanging drop cultures without adding sterile oat flakes, bacteria or other microorganisms. Its complete life cycle including spore germination, myxamoebae, swarm cells, plasmodial development, and maturity of fructifications was demonstrated. Details of spore-to-spore development are described and illustrated.

Asexuality: Sexual Orientation, Paraphilia, Sexual Dysfunction, or None of the Above?

PubMed

Brotto, Lori A; Yule, Morag

2017-04-01

Although lack of sexual attraction was first quantified by Kinsey, large-scale and systematic research on the prevalence and correlates of asexuality has only emerged over the past decade. Several theories have been posited to account for the nature of asexuality. The goal of this review was to consider the evidence for whether asexuality is best classified as a psychiatric syndrome (or a symptom of one), a sexual dysfunction, or a paraphilia. Based on the available science, we believe there is not sufficient evidence to support the categorization of asexuality as a psychiatric condition (or symptom of one) or as a disorder of sexual desire. There is some evidence that a subset of self-identified asexuals have a paraphilia. We also considered evidence supporting the classification of asexuality as a unique sexual orientation. We conclude that asexuality is a heterogeneous entity that likely meets conditions for a sexual orientation, and that researchers should further explore evidence for such a categorization.

Aspergillus fumigatus spore proteomics and genetics reveal that VeA represses DefA-mediated DNA damage response.

PubMed

Shin, Kwang-Soo; Park, Hee-Soo; Kim, Young; Heo, In-Beom; Kim, Young Hwan; Yu, Jae-Hyuk

2016-10-04

Aspergillus fumigatus reproduces and infects host by forming a high number of small asexual spores (conidia). The velvet proteins are global transcriptional regulators governing the complex process of conidiogenesis in this fungus. Here, to further understand the velvet-mediated regulation, we carried out comparative proteomic analyses of conidia of wild type (WT) and three velvet mutants (ΔveA, ΔvelB and ΔvosA). Cluster analysis of 184 protein spots showing at least 1.5-fold differential accumulation between WT and mutants reveal the clustering of WT- ΔveA and ΔvelB-ΔvosA. Among 43 proteins identified by Nano-LC-ESI-MS/MS, 23 including several heat shock proteins showed more than two-fold reduction in both the ∆velB and ∆vosA conidia. On the contrary, three proteins exhibited more than five-fold increase in ∆veA only, including the putative RNA polymerase II degradation factor DefA. The deletion of defA resulted in a reduced number of conidia and restricted colony growth. In addition, the defA deletion mutant conidia showed hypersensitivity against the DNA damaging agents NQO and MMS, while the ΔveA mutant conidia were more resistant against to NQO. Taken together, we propose that VeA controls protein level of DefA in conidia, which are dormant and equipped with multiple layers of protection against environmental cues. Copyright © 2016 Elsevier B.V. All rights reserved.

PpASCL, the Physcomitrella patens Anther-Specific Chalcone Synthase-Like Enzyme Implicated in Sporopollenin Biosynthesis, Is Needed for Integrity of the Moss Spore Wall and Spore Viability

PubMed Central

Daku, Rhys M.; Rabbi, Fazle; Buttigieg, Josef; Coulson, Ian M.; Horne, Derrick; Martens, Garnet; Ashton, Neil W.; Suh, Dae-Yeon

2016-01-01

Sporopollenin is the main constituent of the exine layer of spore and pollen walls. The anther-specific chalcone synthase-like (ASCL) enzyme of Physcomitrella patens, PpASCL, has previously been implicated in the biosynthesis of sporopollenin, the main constituent of exine and perine, the two outermost layers of the moss spore cell wall. We made targeted knockouts of the corresponding gene, PpASCL, and phenotypically characterized ascl sporophytes and spores at different developmental stages. Ascl plants developed normally until late in sporophytic development, when the spores produced were structurally aberrant and inviable. The development of the ascl spore cell wall appeared to be arrested early in microspore development, resulting in small, collapsed spores with altered surface morphology. The typical stratification of the spore cell wall was absent with only an abnormal perine recognisable above an amorphous layer possibly representing remnants of compromised intine and/or exine. Equivalent resistance of the spore walls of ascl mutants and the control strain to acetolysis suggests the presence of chemically inert, defective sporopollenin in the mutants. Anatomical abnormalities of late-stage ascl sporophytes include a persistent large columella and an air space incompletely filled with spores. Our results indicate that the evolutionarily conserved PpASCL gene is needed for proper construction of the spore wall and for normal maturation and viability of moss spores. PMID:26752629

Measuring Total and Germinable Spore Populations

NASA Technical Reports Server (NTRS)

Noell, A.C.; Yung, P.T.; Yang, W.; Lee, C.; Ponce, A.

2011-01-01

It has been shown that bacterial endospores can be enumerated using a microscopy based assay that images the luminescent halos from terbium ions bound to dipicolinic acid, a spore specific chemical marker released upon spore germination. Further development of the instrument has simplified it towards automation while at the same time improving image quality. Enumeration of total spore populations has also been developed allowing measurement of the percentage of viable spores in any population by comparing the germinable/culturable spores to the total. Percentage viability will allow a more quantitative comparison of the ability of spores to survive across a wide range of extreme environments.

Early development of fern gametophytes in microgravity

NASA Technical Reports Server (NTRS)

Roux, Stanley J.; Chatterjee, Ani; Hillier, Sheila; Cannon, Tom

2003-01-01

Dormant spores of the fern Ceratopteris richardii were flown on Shuttle mission STS-93 to evaluate the effects of micro-g on their development and on their pattern of gene expression. Prior to flight the spores were sterilized and sown into one of two environments: (1) Microscope slides in a video-microscopy module; and (2) Petri dishes. All spores were then stored in darkness until use. Spore germination was initiated on orbit after exposure to light. For the spores on microscope slides, cell level changes were recorded through the clear spore coat of the spores by video microscopy. After their exposure to light, spores in petri dishes were frozen in orbit at four different time points during which on earth gravity fixes the polarity of their development. Spores were then stored frozen in Biological Research in Canister units until recovery on earth. The RNAs from these cells and from 1-g control cells were extracted and analyzed on earth after flight to assay changes in gene expression. Video microscopy results revealed that the germinated spores developed normally in microgravity, although the polarity of their development, which is guided by gravity on earth, was random in space. Differential Display-PCR analyses of RNA extracted from space-flown cells showed that there was about a 5% change in the pattern of gene expression between cells developing in micro-g compared to those developing on earth. c2002 Published by Elsevier Science Ltd on behalf of COSPAR.

High-Throughput Assay and Discovery of Small Molecules that Interrupt Malaria Transmission

PubMed Central

Plouffe, David M.; Wree, Melanie; Du, Alan Y.; Meister, Stephan; Li, Fengwu; Patra, Kailash; Lubar, Aristea; Okitsu, Shinji L.; Flannery, Erika L.; Kato, Nobutaka; Tanaseichuk, Olga; Comer, Eamon; Zhou, Bin; Kuhen, Kelli; Zhou, Yingyao; Leroy, Didier; Schreiber, Stuart L.; Scherer, Christina A.; Vinetz, Joseph; Winzeler, Elizabeth A.

2016-01-01

Summary Preventing transmission is an important element of malaria control. However, most of the current available methods to assay for malaria transmission blocking are relatively low throughput and cannot be applied to large chemical libraries. We have developed a high-throughput and cost-effective assay, the Saponin-lysis Sexual Stage Assay (SaLSSA), for identifying small molecules with transmission-blocking capacity. SaLSSA analysis of 13,983 unique compounds uncovered that >90% of well-characterized antimalarials, including endoperoxides and 4-aminoquinolines, as well as compounds active against asexual blood stages, lost most of their killing activity when parasites developed into metabolically quiescent stage V gametocytes. On the other hand, we identified compounds with consistent low nanomolar transmission-blocking activity, some of which showed cross-reactivity against asexual blood and liver stages. The data clearly emphasize substantial physiological differences between sexual and asexual parasites and provide a tool and starting points for the discovery and development of transmission-blocking drugs. PMID:26749441

Bioforensics: Characterization of biological weapons agents by NanoSIMS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weber, P K; Ghosal, S; Leighton, T J

2007-02-26

The anthrax attacks of Fall 2001 highlight the need to develop forensic methods based on multiple identifiers to determine the origin of biological weapons agents. Genetic typing methods (i.e., DNA and RNA-based) provide one attribution technology, but genetic information alone is not usually sufficient to determine the provenance of the material. Non-genetic identifiers, including elemental and isotopic signatures, provide complementary information that can be used to identify the means, geographic location and date of production. Under LDRD funding, we have successfully developed the techniques necessary to perform bioforensic characterization with the NanoSIMS at the individual spore level. We have developedmore » methods for elemental and isotopic characterization at the single spore scale. We have developed methods for analyzing spore sections to map elemental abundance within spores. We have developed rapid focused ion beam (FIB) sectioning techniques for spores to preserve elemental and structural integrity. And we have developed a high-resolution depth profiling method to characterize the elemental distribution in individual spores without sectioning. We used these newly developed methods to study the controls on elemental abundances in spores, characterize the elemental distribution of in spores, and to study elemental uptake by spores. Our work under this LDRD project attracted FBI and DHS funding for applied purposes.« less

Coexistence of sexual individuals and genetically isolated asexual counterparts in a thrips.

PubMed

Kobayashi, Kazuya; Yoshimura, Jin; Hasegawa, Eisuke

2013-11-21

Sex is a paradoxical phenomenon because it is less efficient compared with asexual reproduction. To resolve this paradox we need a direct comparison between sexual and asexual forms. In many organisms, however, sexual and asexual forms do not occur in the same habitat, or at the same time. In a few cases where sexual and asexual forms are found in a single population, some (though rare) genetic exchange is usually detected between the two forms. When genetic exchange occurs a direct comparison is impossible. Here we investigate a thrips exhibiting both sexual and asexual forms (lineages) that are morphologically indistinguishable. We examine if the two forms are genetically isolated. Phylogeny based on nuclear genes confirms that the sexual and asexual lineages are genetically differentiated. Thus we demonstrate that the current system has certain advantages over existing and previously used model systems in the evolution of sexual reproduction.

Comparative toxicant sensitivity of sexual and asexual reproduction in the rotifer Brachionus calyciflorus

DOE Office of Scientific and Technical Information (OSTI.GOV)

Snell, T.W.; Carmona, M.J.

Cyclically parthenogenetic zooplankters like rotifers are important tools for assessing toxicity in aquatic environments. Sexual reproduction is an essential component of rotifer life cycles, but current toxicity tests utilize only asexual reproduction. The authors compared the effects of four toxicants on asexual and sexual reproduction of the rotifer Brachionus calyciflorus. Toxicants had a differential effect on sexual and asexual reproduction, with sexual reproduction consistently the most sensitive. Concentrations of 0.2 {mu}g/ml PCP (sodium pentachlorophenate) had no effect on the asexual reproductive rate, but significantly reduced sexual reproduction. Likewise, chlorpyrifos concentrations of 0.3 {mu}g/ml had no significant effect on asexual reproduction,more » but sexual reproduction was significantly reduced. There was no difference in NOECs, LOECs, and chronic values for asexual and sexual reproduction for cadmium and naphthol tests. However, comparison of toxicant effect levels revealed that sexual reproduction was more strongly reduced at each toxicant concentration. The four toxicants tested inhibited sexual reproduction 2 to 68 times more than asexual reproduction at the lowest observed effect concentrations. Toxicants inhibited sexual reproduction in its initial step: sexual female production. Because sexual reproduction is more sensitive, toxicity tests based exclusively on asexual reproduction may not be protective of rotifer life cycles.« less

Genetic Control of Contagious Asexuality in the Pea Aphid

PubMed Central

Jaquiéry, Julie; Stoeckel, Solenn; Larose, Chloé; Nouhaud, Pierre; Rispe, Claude; Mieuzet, Lucie; Bonhomme, Joël; Mahéo, Frédérique; Legeai, Fabrice; Gauthier, Jean-Pierre; Prunier-Leterme, Nathalie; Tagu, Denis; Simon, Jean-Christophe

2014-01-01

Although evolutionary transitions from sexual to asexual reproduction are frequent in eukaryotes, the genetic bases of such shifts toward asexuality remain largely unknown. We addressed this issue in an aphid species where both sexual and obligate asexual lineages coexist in natural populations. These sexual and asexual lineages may occasionally interbreed because some asexual lineages maintain a residual production of males potentially able to mate with the females produced by sexual lineages. Hence, this species is an ideal model to study the genetic basis of the loss of sexual reproduction with quantitative genetic and population genomic approaches. Our analysis of the co-segregation of ∼300 molecular markers and reproductive phenotype in experimental crosses pinpointed an X-linked region controlling obligate asexuality, this state of character being recessive. A population genetic analysis (>400-marker genome scan) on wild sexual and asexual genotypes from geographically distant populations under divergent selection for reproductive strategies detected a strong signature of divergent selection in the genomic region identified by the experimental crosses. These population genetic data confirm the implication of the candidate region in the control of reproductive mode in wild populations originating from 700 km apart. Patterns of genetic differentiation along chromosomes suggest bidirectional gene flow between populations with distinct reproductive modes, supporting contagious asexuality as a prevailing route to permanent parthenogenesis in pea aphids. This genetic system provides new insights into the mechanisms of coexistence of sexual and asexual aphid lineages. PMID:25473828

Studying molecular changes during gravity perception and response in a single cell.

PubMed

Cannon, Ashley E; Salmi, Mari L; Bushart, Thomas J; Roux, Stanley J

2015-01-01

Early studies revealed a highly predictable pattern of gravity-directed growth and development in Ceratopteris richardii spores. This makes the spores a valuable model system for the study of how a single cell senses and responds to the force of gravity. Gravity regulates both the direction and magnitude of a trans-cell calcium current in germinating spores, and the orientation of this current predicts the polarization of spore development. Molecular techniques have been developed to evaluate the transcriptomic and proteomic profiles of spores before and after gravity establishes the polarity of their development. Here we describe these techniques, along with protocols for sterilizing the spores, sowing them in a solid or liquid growth media, and evaluating germination.

Proteogenomic Analysis Greatly Expands the Identification of Proteins Related to Reproduction in the Apogamous Fern Dryopteris affinis ssp. affinis.

PubMed

Grossmann, Jonas; Fernández, Helena; Chaubey, Pururawa M; Valdés, Ana E; Gagliardini, Valeria; Cañal, María J; Russo, Giancarlo; Grossniklaus, Ueli

2017-01-01

Performing proteomic studies on non-model organisms with little or no genomic information is still difficult. However, many specific processes and biochemical pathways occur only in species that are poorly characterized at the genomic level. For example, many plants can reproduce both sexually and asexually, the first one allowing the generation of new genotypes and the latter their fixation. Thus, both modes of reproduction are of great agronomic value. However, the molecular basis of asexual reproduction is not well understood in any plant. In ferns, it combines the production of unreduced spores (diplospory) and the formation of sporophytes from somatic cells (apogamy). To set the basis to study these processes, we performed transcriptomics by next-generation sequencing (NGS) and shotgun proteomics by tandem mass spectrometry in the apogamous fern D. affinis ssp. affinis . For protein identification we used the public viridiplantae database (VPDB) to identify orthologous proteins from other plant species and new transcriptomics data to generate a "species-specific transcriptome database" (SSTDB). In total 1,397 protein clusters with 5,865 unique peptide sequences were identified (13 decoy proteins out of 1,410, protFDR 0.93% on protein cluster level). We show that using the SSTDB for protein identification increases the number of identified peptides almost four times compared to using only the publically available VPDB. We identified homologs of proteins involved in reproduction of higher plants, including proteins with a potential role in apogamy. With the increasing availability of genomic data from non-model species, similar proteogenomics approaches will improve the sensitivity in protein identification for species only distantly related to models.

Size matters for violent discharge height and settling speed of Sphagnum spores: important attributes for dispersal potential.

PubMed

Sundberg, Sebastian

2010-02-01

Initial release height and settling speed of diaspores are biologically controlled components which are key to modelling wind dispersal. Most Sphagnum (peat moss) species have explosive spore liberation. In this study, how capsule and spore sizes affect the height to which spores are propelled were measured, and how spore size and spore number of discharged particles relate to settling speed in the aspherical Sphagnum spores. Spore discharge and spore cloud development were filmed in a closed chamber (nine species). Measurements were taken from snapshots at three stages of cloud development. Settling speed of spores (14 species) and clusters were timed in a glass tube. The maximum discharge speed measured was 3.6 m s(-1). Spores reached a maximum height of 20 cm (average: 15 cm) above the capsule. The cloud dimensions at all stages were related positively to capsule size (R(2) = 0.58-0.65). Thus species with large shoots (because they have large capsules) have a dispersal advantage. Half of the spores were released as singles and the rest as clusters (usually two to four spores). Single spores settled at 0.84-1.86 cm s(-1), about 52 % slower than expected for spherical spores with the same diameters. Settling speed displayed a positive curvilinear relationship with spore size, close to predictions by Stokes' law for spherical spores with 68 % of the actual diameters. Light-coloured spores settled slower than dark spores. Settling speed of spore clusters agrees with earlier studies. Effective spore discharge and small, slowly settling spores appear particularly important for species in forested habitats. The spore discharge heights in Sphagnum are among the greatest for small, wind-dispersed propagules. The discharge heights and the slow settling of spores affect dispersal distances positively and may help to explain the wide distribution of most boreal Sphagnum species.

Size matters for violent discharge height and settling speed of Sphagnum spores: important attributes for dispersal potential

PubMed Central

Sundberg, Sebastian

2010-01-01

Background and Aims Initial release height and settling speed of diaspores are biologically controlled components which are key to modelling wind dispersal. Most Sphagnum (peat moss) species have explosive spore liberation. In this study, how capsule and spore sizes affect the height to which spores are propelled were measured, and how spore size and spore number of discharged particles relate to settling speed in the aspherical Sphagnum spores. Methods Spore discharge and spore cloud development were filmed in a closed chamber (nine species). Measurements were taken from snapshots at three stages of cloud development. Settling speed of spores (14 species) and clusters were timed in a glass tube. Key Results The maximum discharge speed measured was 3·6 m s−1. Spores reached a maximum height of 20 cm (average: 15 cm) above the capsule. The cloud dimensions at all stages were related positively to capsule size (R2 = 0·58–0·65). Thus species with large shoots (because they have large capsules) have a dispersal advantage. Half of the spores were released as singles and the rest as clusters (usually two to four spores). Single spores settled at 0·84–1·86 cm s−1, about 52 % slower than expected for spherical spores with the same diameters. Settling speed displayed a positive curvilinear relationship with spore size, close to predictions by Stokes' law for spherical spores with 68 % of the actual diameters. Light-coloured spores settled slower than dark spores. Settling speed of spore clusters agrees with earlier studies. Effective spore discharge and small, slowly settling spores appear particularly important for species in forested habitats. Conclusions The spore discharge heights in Sphagnum are among the greatest for small, wind-dispersed propagules. The discharge heights and the slow settling of spores affect dispersal distances positively and may help to explain the wide distribution of most boreal Sphagnum species. PMID:20123930

Cryptic sexual populations account for genetic diversity and ecological success in a widely distributed, asexual fungus-growing ant.

PubMed

Rabeling, Christian; Gonzales, Omar; Schultz, Ted R; Bacci, Maurício; Garcia, Marcos V B; Verhaagh, Manfred; Ishak, Heather D; Mueller, Ulrich G

2011-07-26

Sex and recombination are central processes in life generating genetic diversity. Organisms that rely on asexual propagation risk extinction due to the loss of genetic diversity and the inability to adapt to changing environmental conditions. The fungus-growing ant species Mycocepurus smithii was thought to be obligately asexual because only parthenogenetic populations have been collected from widely separated geographic localities. Nonetheless, M. smithii is ecologically successful, with the most extensive distribution and the highest population densities of any fungus-growing ant. Here we report that M. smithii actually consists of a mosaic of asexual and sexual populations that are nonrandomly distributed geographically. The sexual populations cluster along the Rio Amazonas and the Rio Negro and appear to be the source of independently evolved and widely distributed asexual lineages, or clones. Either apomixis or automixis with central fusion and low recombination rates is inferred to be the cytogenetic mechanism underlying parthenogenesis in M. smithii. Males appear to be entirely absent from asexual populations, but their existence in sexual populations is indicated by the presence of sperm in the reproductive tracts of queens. A phylogenetic analysis of the genus suggests that M. smithii is monophyletic, rendering a hybrid origin of asexuality unlikely. Instead, a mitochondrial phylogeny of sexual and asexual populations suggests multiple independent origins of asexual reproduction, and a divergence-dating analysis indicates that M. smithii evolved 0.5-1.65 million years ago. Understanding the evolutionary origin and maintenance of asexual reproduction in this species contributes to a general understanding of the adaptive significance of sex.

Freedom, invisibility, and community: a qualitative study of self-identification with asexuality.

PubMed

MacNeela, Pádraig; Murphy, Aisling

2015-04-01

A significant body of research is now emerging on the subjective meaning of asexuality. This study explored how self-identification as asexual is managed, both as a threat to the self-concept and a source of personal meaning. A total of 66 self-identified asexuals were recruited from an asexuality internet community and responded to open-ended questions on an online survey. Of these, 31 participants identified as female, 15 as male, 18 gave a different label such as genderqueer or androgynous, and two did not provide information on gender. A thematic analysis of the transcripts resulted in three themes. Socially, asexuality attracted denial and resistance due to incompatibility with heteronormative societal expectations. Despite the threat to self-integrity arising from asexuality being socially rejected, it was typically assimilated as a valued and meaningful orientation on an intra-personal level, aided by information and support from the online community. A second level of threat to self arose whereby other self-identifications, especially gender, had to be reconciled with a non-sexual persona. The accommodation made to other elements of the self was reflected in complex sub-identities. The findings were interpreted using identity process theory to understand how threats arising from self-identifying as asexual are managed. Although asexuality emerges as an orientation to sexuality that can be reconciled with the self, its invisibility or outright rejection in society constitute an on-going challenge.

Wnt affects symmetry and morphogenesis during post-embryonic development in colonial chordates.

PubMed

Di Maio, Alessandro; Setar, Leah; Tiozzo, Stefano; De Tomaso, Anthony W

2015-01-01

Wnt signaling is one of the earliest and most highly conserved regulatory pathways for the establishment of the body axes during regeneration and early development. In regeneration, body axes determination occurs independently of tissue rearrangement and early developmental cues. Modulation of the Wnt signaling in either process has shown to result in unusual body axis phenotypes. Botryllus schlosseri is a colonial ascidian that can regenerate its entire body through asexual budding. This processes leads to an adult body via a stereotypical developmental pathway (called blastogenesis), without proceeding through any embryonic developmental stages. In this study, we describe the role of the canonical Wnt pathway during the early stages of asexual development. We characterized expression of three Wnt ligands (Wnt2B, Wnt5A, and Wnt9A) by in situ hybridization and qRT-PCR. Chemical manipulation of the pathway resulted in atypical budding due to the duplication of the A/P axes, supernumerary budding, and loss of the overall cell apical-basal polarity. Our results suggest that Wnt signaling is used for equivalent developmental processes both during embryogenesis and asexual development in an adult organism, suggesting that patterning mechanisms driving morphogenesis are conserved, independent of embryonic, or regenerative development.

Recurrent loss of sex is associated with accumulation of deleterious mutations in Oenothera.

PubMed

Hollister, Jesse D; Greiner, Stephan; Wang, Wei; Wang, Jun; Zhang, Yong; Wong, Gane Ka-Shu; Wright, Stephen I; Johnson, Marc T J

2015-04-01

Sexual reproduction is nearly universal among eukaryotes. Theory predicts that the rarity of asexual eukaryotic species is in part caused by accumulation of deleterious mutations and heightened extinction risk associated with suppressed recombination and segregation in asexual species. We tested this prediction with a large data set of 62 transcriptomes from 29 species in the plant genus Oenothera, spanning ten independent transitions between sexual and a functionally asexual genetic system called permanent translocation heterozygosity. Illumina short-read sequencing and de novo transcript assembly yielded an average of 16.4 Mb of sequence per individual. Here, we show that functionally asexual species accumulate more deleterious mutations than sexual species using both population genomic and phylogenetic analysis. At an individual level, asexual species exhibited 1.8 × higher heterozygosity than sexual species. Within species, we detected a higher proportion of nonsynonymous polymorphism relative to synonymous variation within asexual compared with sexual species, indicating reduced efficacy of purifying selection. Asexual species also exhibited a greater proportion of transcripts with premature stop codons. The increased proportion of nonsynonymous mutations was also positively correlated with divergence time between sexual and asexual species, consistent with Muller's ratchet. Between species, we detected repeated increases in the ratio of nonsynonymous to synonymous divergence in asexual species compared with sexually reproducing sister taxa, indicating increased accumulation of deleterious mutations. These results confirm that an important advantage of sex is that it facilitates selection against deleterious alleles, which might help to explain the dearth of extant asexual species. © The Author 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Test method development to evaluate hot, humid air decontamination of materials contaminated with Bacillus anthracis ∆Sterne and B. thuringiensis Al Hakam spores.

PubMed

Buhr, T L; Young, A A; Minter, Z A; Wells, C M; McPherson, D C; Hooban, C L; Johnson, C A; Prokop, E J; Crigler, J R

2012-11-01

To develop test methods and evaluate the survival of Bacillus anthracis ∆Sterne and Bacillus thuringiensis Al Hakam spores after exposure to hot, humid air. Spores (>7 logs) of both strains were dried on six different test materials. Response surface methodology was employed to identify the limits of spore survival at optimal test combinations of temperature (60, 68, 77°C), relative humidity (60, 75, 90%) and time (1, 4, 7 days). No spores survived the harshest test run (77°C, 90% r.h., 7 days), while > 6·5 logs of spores survived the mildest test run (60°C, 60% r.h., 1 day). Spores of both strains inoculated on nylon webbing and polypropylene had greater survival rates at 68°C, 75% r.h., 4 days than spores on other materials. Electron microscopy showed no obvious physical damage to spores using hot, humid air, which contrasted with pH-adjusted bleach decontamination. Test methods were developed to show that hot, humid air effectively inactivates B. anthracis ∆Sterne and B. thuringiensis Al Hakam spores with similar kinetics. Hot, humid air is a potential alternative to conventional chemical decontamination. © 2012 The Authors Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.

Origination of asexual plantlets in three species of Crassulaceae.

PubMed

Guo, Jiansheng; Liu, Hailiang; He, Yangyang; Cui, Xianghuan; Du, Xiling; Zhu, Jian

2015-03-01

During asexual plant reproduction, cells from different organs can be reprogrammed to produce new individuals, a process that requires the coordination of cell cycle reactivation with the acquisition of other cellular morphological characteristics. However, the factors that influence the variety of asexual reproduction have not yet been determined. Here, we report on plantlet formation in Kalanchoe daigremontiana, Graptopetalum paraguayense, and Crassula portulacea (Crassulaceae) and analyse the effect of initiating cells on asexual reproduction in these three species. Additionally, the roles of WUSCHEL (WUS) and CUP-SHAPED COTYLEDON 1 (CUC1) in the asexual reproduction of these species were analysed through qRT-PCR. Our results indicated that pre-existing stem cell-like cells at the sites of asexual reproduction were responsible for the formation of plantlets. These cells were arrested in different phases of the cell cycle and showed different cell morphological characteristics and cell counts. The accumulation of auxin and cytokinin at the sites of asexual plantlet formation indicated their important functions, particularly for cell cycle reactivation. These differences may influence the pattern and complexity of asexual reproduction in these Crassulaceae species. Additionally, the dynamic expression levels of CUC1 and WUS may indicate that CUC1 functions in the formation of callus and shoot meristems; whereas, WUS was only associated with shoot induction.

Decontamination of materials contaminated with Bacillus anthracis and Bacillus thuringiensis Al Hakam spores using PES-Solid, a solid source of peracetic acid.

PubMed

Buhr, T L; Wells, C M; Young, A A; Minter, Z A; Johnson, C A; Payne, A N; McPherson, D C

2013-08-01

To develop test methods and evaluate survival of Bacillus anthracis Ames, B. anthracis ∆Sterne and B. thuringiensis Al Hakam spores after exposure to PES-Solid (a solid source of peracetic acid), including PES-Solid formulations with bacteriostatic surfactants. Spores (≥ 7 logs) were dried on seven different test materials and treated with three different PES-Solid formulations (or preneutralized controls) at room temperature for 15 min. There was either no spore survival or less than 1 log (<10 spores) of spore survival in 56 of 63 test combinations (strain, formulation and substrate). Less than 2.7 logs (<180 spores) survived in the remaining seven test combinations. The highest spore survival rates were seen on water-dispersible chemical agent resistant coating (CARC-W) and Naval ship topcoat (NTC). Electron microscopy and Coulter analysis showed that all spore structures were intact after spore inactivation with PES-Solid. Three PES-Solid formulations inactivated Bacillus spores that were dried on seven different materials. A test method was developed to show that PES-Solid formulations effectively inactivate Bacillus spores on different materials. Published 2013. This article is a U.S. Government work and is in the public domain in the USA.

Habitat heterogeneity favors asexual reproduction in natural populations of grassthrips

PubMed Central

Lavanchy, Guillaume; Strehler, Marie; Llanos Roman, Maria Noemi; Lessard‐Therrien, Malie; Humbert, Jean‐Yves; Dumas, Zoé; Jalvingh, Kirsten; Ghali, Karim; Fontcuberta García‐Cuenca, Amaranta; Zijlstra, Bart; Arlettaz, Raphaël; Schwander, Tanja

2016-01-01

Explaining the overwhelming success of sex among eukaryotes is difficult given the obvious costs of sex relative to asexuality. Different studies have shown that sex can provide benefits in spatially heterogeneous environments under specific conditions, but whether spatial heterogeneity commonly contributes to the maintenance of sex in natural populations remains unknown. We experimentally manipulated habitat heterogeneity for sexual and asexual thrips lineages in natural populations and under seminatural mesocosm conditions by varying the number of hostplants available to these herbivorous insects. Asexual lineages rapidly replaced the sexual ones, independently of the level of habitat heterogeneity in mesocosms. In natural populations, the success of sexual thrips decreased with increasing habitat heterogeneity, with sexual thrips apparently only persisting in certain types of hostplant communities. Our results illustrate how genetic diversity‐based mechanisms can favor asexuality instead of sex when sexual lineages co‐occur with genetically variable asexual lineages. PMID:27346066

Consequences of Asexuality in Natural Populations: Insights from Stick Insects.

PubMed

Bast, Jens; Parker, Darren J; Dumas, Zoé; Jalvingh, Kirsten M; Tran Van, Patrick; Jaron, Kamil S; Figuet, Emeric; Brandt, Alexander; Galtier, Nicolas; Schwander, Tanja

2018-07-01

Recombination is a fundamental process with significant impacts on genome evolution. Predicted consequences of the loss of recombination include a reduced effectiveness of selection, changes in the amount of neutral polymorphisms segregating in populations, and an arrest of GC-biased gene conversion. Although these consequences are empirically well documented for nonrecombining genome portions, it remains largely unknown if they extend to the whole genome scale in asexual organisms. We identify the consequences of asexuality using de novo transcriptomes of five independently derived, obligately asexual lineages of stick insects, and their sexual sister-species. We find strong evidence for higher rates of deleterious mutation accumulation, lower levels of segregating polymorphisms and arrested GC-biased gene conversion in asexuals as compared with sexuals. Taken together, our study conclusively shows that predicted consequences of genome evolution under asexuality can indeed be found in natural populations.

Biological markers of asexuality: Handedness, birth order, and finger length ratios in self-identified asexual men and women.

PubMed

Yule, Morag A; Brotto, Lori A; Gorzalka, Boris B

2014-02-01

Human asexuality is defined as a lack of sexual attraction to anyone or anything and it has been suggested that it may be best conceptualized as a sexual orientation. Non-right-handedness, fraternal birth order, and finger length ratio (2D:4D) are early neurodevelopmental markers associated with sexual orientation. We conducted an Internet study investigating the relationship between self-identification as asexual, handedness, number of older siblings, and self-measured finger-lengths in comparison to individuals of other sexual orientation groups. A total of 325 asexuals (60 men and 265 women; M age, 24.8 years), 690 heterosexuals (190 men and 500 women; M age, 23.5 years), and 268 non-heterosexuals (homosexual and bisexual; 64 men and 204 women; M age, 29.0 years) completed online questionnaires. Asexual men and women were 2.4 and 2.5 times, respectively, more likely to be non-right-handed than their heterosexual counterparts and there were significant differences between sexual orientation groups in number of older brothers and older sisters, and this depended on handedness. Asexual and non-heterosexual men were more likely to be later-born than heterosexual men, and asexual women were more likely to be earlier-born than non-heterosexual women. We found no significant differences between sexual orientation groups on measurements of 2D:4D ratio. This is one of the first studies to test and provide preliminary empirical support for an underlying neurodevelopmental basis to account for the lack of sexual attraction characteristic of asexuality.

Mechanisms of Bacterial Spore Germination and Its Heterogeneity

DTIC Science & Technology

2015-01-10

mathematical model describing spore germination has been developed; 9) much of the work above has been extended to Clostridium spores; and 10) ~90...germination. C) Faeder lab, with Li and Setlow labs. We have developed a mathematical model of bacterial spore germination that accounts for...heterogeneity in both Tlag and commitment times. The model is built from three main mathematical components: a receptor distribution function

Origin, divergence, and phylogeny of asexual Epichloë endophyte in Elymus species from western China.

PubMed

Song, Hui; Nan, Zhibiao

2015-01-01

Asexual Epichloë species are likely derived directly from sexual Epichloë species that then lost their capacity for sexual reproduction or lost sexual reproduction because of interspecific hybridization between distinct lineages of sexual Epichloë and/or asexual Epichloë species. In this study we isolated asexual Epichloë endophytes from Elymus species in western China and sequenced intron-rich regions in the genes encoding β-tubulin (tubB) and translation elongation factor 1-α (tefA). Our results showed that there are no gene copies of tubB and tefA in any of the isolates. Phylogenetic analysis showed that sequences in this study formed a single clade with asexual Epichloë bromicola from Hordeum brevisubulatum, which implies asexual Epichloë endophytes that are symbionts in a western Chinese Elymus species likely share a common ancestor with asexual E. bromicola from European H. brevisubulatum. In addition, our results revealed that asexual E. bromicola isolates that are symbionts in a western Chinese Elymus species and sexual Epichloë species that are symbionts in a North American Elymus species have a different origin. Further analysis found that Epichloë species likely originated in Eurasia. In addition, the results support the hypothesis that migratory birds or humans might have aided the dispersal of these fungal endophytes to other continents.

Origin, Divergence, and Phylogeny of Asexual Epichloë Endophyte in Elymus Species from Western China

PubMed Central

Song, Hui; Nan, Zhibiao

2015-01-01

Asexual Epichloë species are likely derived directly from sexual Epichloë species that then lost their capacity for sexual reproduction or lost sexual reproduction because of interspecific hybridization between distinct lineages of sexual Epichloë and/or asexual Epichloë species. In this study we isolated asexual Epichloë endophytes from Elymus species in western China and sequenced intron-rich regions in the genes encoding β-tubulin (tubB) and translation elongation factor 1-α (tefA). Our results showed that there are no gene copies of tubB and tefA in any of the isolates. Phylogenetic analysis showed that sequences in this study formed a single clade with asexual Epichloë bromicola from Hordeum brevisubulatum, which implies asexual Epichloë endophytes that are symbionts in a western Chinese Elymus species likely share a common ancestor with asexual E. bromicola from European H. brevisubulatum. In addition, our results revealed that asexual E. bromicola isolates that are symbionts in a western Chinese Elymus species and sexual Epichloë species that are symbionts in a North American Elymus species have a different origin. Further analysis found that Epichloë species likely originated in Eurasia. In addition, the results support the hypothesis that migratory birds or humans might have aided the dispersal of these fungal endophytes to other continents. PMID:25970178

Planarian D-amino acid oxidase is involved in ovarian development during sexual induction.

PubMed

Maezawa, Takanobu; Tanaka, Hiroyuki; Nakagawa, Haruka; Ono, Mizuki; Aoki, Manabu; Matsumoto, Midori; Ishida, Tetsuo; Horiike, Kihachiro; Kobayashi, Kazuya

2014-05-01

To elucidate the molecular mechanisms underlying switching from asexual to sexual reproduction, namely sexual induction, we developed an assay system for sexual induction in the hermaphroditic planarian species Dugesia ryukyuensis. Ovarian development is the initial and essential step in sexual induction, and it is followed by the formation of other reproductive organs, including the testes. Here, we report a function of a planarian D-amino acid oxidase, Dr-DAO, in the control of ovarian development in planarians. Asexual worms showed significantly more widespread expression of Dr-DAO in the parenchymal space than did sexual worms. Inhibition of Dr-DAO by RNAi caused the formation of immature ovaries. In addition, we found that feeding asexual worms 5 specific D-amino acids could induce the formation of immature ovaries that are similar to those observed in Dr-DAO knockdown worms, suggesting that Dr-DAO inhibits the formation of immature ovaries by degrading these D-amino acids. Following sexual induction, Dr-DAO expression was observed in the ovaries. The knockdown of Dr-DAO during sexual induction delayed the maturation of the other reproductive organs, as well as ovary. These findings suggest that Dr-DAO acts to promote ovarian maturation and that complete sexual induction depends on the production of mature ovaries. We propose that Dr-DAO produced in somatic cells prevents the onset of sexual induction in the asexual state, and then after sexual induction, the female germ cells specifically produce Dr-DAO to induce full maturation. Therefore, Dr-DAO produced in somatic and female germline cells may play different roles in sexual induction. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Fungal microcolonies on indoor surfaces — an explanation for the base-level fungal spore counts in indoor air

NASA Astrophysics Data System (ADS)

Pasanen, A.-L.; Heinonen-Tanski, H.; Kalliokoski, P.; Jantunen, M. J.

In the subarctic winter, fungal spores are found in indoor air even when outdoor spore levels are very low. The results of this study support an explanation that some indoor airborne fungal spores are derived from unnoticeable fungal microcolonies, which may develop on temporarily wet surfaces. Laboratory experiments on Penicillium verrucosum indicated that the fungus germinated on new wallpaper very quickly (about half an hour) under moist conditions. Hyphal growth and sporulation of the fungus on moist wallpaper occured within one day of incubation. In gravity-settling tape samples from occasionally wet surfaces in a suburban home, large spore aggregates, hyphal fragments with some spores and spores in the germination stage were found, indicating fungal growth. These experiments showed that fungal microcolonies can develop within a week on occasionally wet indoor surfaces.

Cyclical parthenogenesis and viviparity in aphids as evolutionary novelties.

PubMed

Davis, Gregory K

2012-09-01

Evolutionary novelties represent challenges to biologists, particularly those who would like to understand the developmental and genetic changes responsible for their appearance. Most modern aphids possess two apparent evolutionary novelties: cyclical parthenogenesis (a life cycle with both sexual and asexual phases) and viviparity (internal development and live birth of progeny) in their asexual phase. Here I discuss the evolution of these apparent novelties from a developmental standpoint. Although a full understanding of the evolution of cyclical parthenogenesis and viviparity in aphids can seem a daunting task, these complex transitions can at least be broken down into a handful of steps. I argue that these should include the following: a differentiation of two developmentally distinct oocytes; de novo synthesis of centrosomes and modification of meiosis during asexual oogenesis; a loss or bypass of any cell cycle arrest and changes in key developmental events during viviparous oogenesis; and a change in how mothers specify the sexual vs. asexual fates of their progeny. Grappling with the nature of such steps and the order in which they occurred ought to increase our understanding and reduce the apparent novelty of complex evolutionary transitions. © 2012 Wiley Periodicals, Inc.

Effects of plant sex on range distributions and allocation to reproduction.

PubMed

Johnson, Marc T J; Smith, Stacey D; Rausher, Mark D

2010-05-01

Despite an abundance of theory, few empirical studies have explored the ecological and evolutionary consequences of sex. We used a comparative phylogenetic approach to examine whether transitions between sexual and asexual reproduction are associated with changes in the size and distribution of species' geographical ranges, and their investment in reproduction. Here, we reconstructed the phylogeny of the genus Oenothera sections Oenothera and Calylophus (Onagraceae), which contain 35 sexual and 30 functionally asexual species. From each species, we collected data on the geographical distribution and variation in plant traits related to reproduction. Functionally asexual species occurred at higher latitudes, but did not differ in range size, compared with sexual species. Transitions to asexuality were associated with decreased investment in floral structures, including the length of petals, floral tubes and styles. Decreased anther size and increased seed size within asexual species also suggest altered allocation to male and female fitness. The observed range shifts are consistent with superior colonization of environments by asexual species following glaciation, and the observed changes in reproductive allocation support predictions made by models relating to the evolution of selfing. Our results suggest that the evolutionary consequences of asexual reproduction might be less restrictive than previously thought.

Population dynamics with a mixed type of sexual and asexual reproduction in a fluctuating environment

PubMed Central

2012-01-01

Background Carassius gibelio, a cyprinid fish from Eurasia, has the ability to reproduce both sexually and asexually. This fish is also known as an invasive species which colonized almost all continental Europe, most likely originating from Asia and Eastern Europe. Populations of both sexually and asexually reproducing individuals exist in sympatry. In this study we try to elucidate the advantages of such a mixed type of reproduction. We investigate the dynamics of two sympatric populations with sexual and asexual reproduction in a periodically fluctuating environment. We define an individual-based computational model in which genotypes are represented by L loci, and the environment is composed of L resources for which the two populations compete. Results Our model demonstrates advantageous population dynamics where the optimal percentage of asexual reproduction depends on selection strength, on the number of selected loci and on the timescale of environmental fluctuations. We show that the sexual reproduction is necessary for "generating" fit genotypes, while the asexual reproduction is suitable for "amplifying" them. The simulations show that the optimal percentage of asexual reproduction increases with the length of the environment stability period and decrease with the strength of the selection and the number of loci. Conclusions In this paper we addressed the advantages of a mixed type of sexual and asexual reproduction in a changing environment and explored the idea that a species that is able to adapt itself to environmental fluctuation can easily colonize a new habitat. Our results could provide a possible explanation for the rapid and efficient invasion of species with a variable ratio of sexual and asexual reproduction such as Carassius gibelio. PMID:22489797

Requirements for the Development of Bacillus Anthracis Spore Reference Materials Used to Test Detection Systems

DTIC Science & Technology

2006-01-01

the sporangium) contributes the com- plex layers of the spore coats that encase the spore DNA. The mother cell dies and begins to fall apart at the end...spores. Bacillus spores contain a number of coat layers and some species posses an additional outermost layer called the exosporium. BA, B. cereus, and B...exosporium is the outermost layer of the BA spores, it likely contains important protein and carbohydrate markers that are recognized by antibodies

Microbial Burden Approach : New Monitoring Approach for Measuring Microbial Burden

NASA Technical Reports Server (NTRS)

Venkateswaran, Kasthuri; Vaishampayan, Parag; Barmatz, Martin

2013-01-01

Advantages of new approach for differentiating live cells/ spores from dead cells/spores. Four examples of Salmonella outbreaks leading to costly destruction of dairy products. List of possible collaboration activities between JPL and other industries (for future discussion). Limitations of traditional microbial monitoring approaches. Introduction to new approach for rapid measurement of viable (live) bacterial cells/spores and its areas of application. Detailed example for determining live spores using new approach (similar procedure for determining live cells). JPL has developed a patented approach for measuring amount of live and dead cells/spores. This novel "molecular" method takes less than 5 to 7 hrs. compared to the seven days required using conventional techniques. Conventional "molecular" techniques can not discriminate live cells/spores among dead cells/spores. The JPL-developed novel method eliminates false positive results obtained from conventional "molecular" techniques that lead to unnecessary delay in the processing and to unnecessary destruction of food products.

Pharmacological Properties of Biocompounds from Spores of the Lingzhi or Reishi Medicinal Mushroom Ganoderma lucidum (Agaricomycetes): A Review.

PubMed

Soccol, Carlos Ricardo; Bissoqui, Lucas Yamasaki; Rodrigues, Cristine; Rubel, Rosalia; Sella, Sandra R B R; Leifa, Fan; de Souza Vandenberghe, Luciana Porto; Soccol, Vanete Thomaz

2016-01-01

Ganoderma lucidum is a well-known representative of mushrooms that have been used in traditional Chinese medicine for centuries. New discoveries related to this medicinal mushroom and its biological properties are frequently reported. However, only recently have scientists started to pay special attention to G. lucidum spores. This is in part because of the recent development of methods for breaking the spore wall and extracting biocompounds from the spore. Although some research groups are working with G. lucidum spores, data in the literature are still limited, and the methods used have not been systematized. This review therefore describes the main advances in techniques for breaking the spore wall and extracting biocompounds from the spore. In addition, the major active components identified and their biological properties, such as neurological activity and antiaging and cell-protective effects, are investigated because these are of importance for potential drug development.

A putative APSES transcription factor is necessary for normal growth and development of Aspergillus nidulans.

PubMed

Lee, Ji-Yeon; Kim, Lee-Han; Kim, Ha-Eun; Park, Jae-Sin; Han, Kap-Hoon; Han, Dong-Min

2013-12-01

The nsdD gene encoding a GATA type transcription factor positively controls sexual development in Aspergillus nidulans. According to microarray data, 20 genes that were upregulated by deleting nsdD during various life cycle stages were randomly selected and deleted for functional analysis. None of the mutants showed apparent changes in growth or development compared with those of the wild-type except the AN3154 gene that encodes a putative APSES transcription factor and is an ortholog of Saccharomyces cerevisiae swi4. Deleting AN3154 resulted in retarded growth and development, and the gene was named rgdA (retared growth and development). The rgdA deletion mutant developed a reduced number of conidia even under favorable conditions for asexual development. The retarded growth and development was partially suppressed by the veA1 mutation. The conidial heads of the mutant aborted, showing reduced and irregular shaped phialides. Fruiting body development was delayed compared with that in the wild-type. The mutant did not respond to various nutritional or environmental factors that affected the development patterns. The rgdA gene was expressed at low levels throughout the life cycle and was not significantly affected by several regulators of sexual and asexual development such as nsdD, veA, stuA, or brlA. However, the rgdA gene affected brlA and abaA expression, which function as key regulators of asexual sporulation, suggesting that rgdA functions upstream of those genes.

Sexual conflict and the evolution of asexuality at low population densities.

PubMed

Gerber, Nina; Kokko, Hanna

2016-10-26

Theories for the evolution of sex rarely include facultatively sexual reproduction. Sexual harassment by males is an underappreciated factor: it should at first sight increase the relative advantage of asexual reproduction by increasing the cost of sex. However, if the same females can perform either sexual or asexual life cycles, then females trying to reproduce asexually may not escape harassment. If resisting male harassment is costly, it might be beneficial for a female to accept a mating and undertake a sexual life cycle rather than 'insist' on an asexual one. We investigate the effects of sexual harassment on the maintenance of sex under different population densities. Our model shows that resisting matings pays off at low population densities, which leads to the complete extinction of males, and thus to the evolution of completely asexual populations. Facultative sex persists in a narrow range of slightly higher densities. At high densities, selection favours giving up resisting male mating attempts and thus sexual reproduction takes over. These interactions between the outcomes of sexual conflict and population density suggest an explanation for the rarity of facultative sex and also patterns of geographical parthenogenesis, where marginal environments with potentially low densities are associated with asexuality. © 2016 The Author(s).

A cytoplasmically transmissible hypovirulence phenotype associated with mitochondrial DNA mutations in the chestnut blight fungus Cryphonectria parasitica.

PubMed Central

Monteiro-Vitorello, C B; Bell, J A; Fulbright, D W; Bertrand, H

1995-01-01

Mutations causing mitochondrial defects were induced in a virulent strain of the chestnut blight fungus Cryphonectria parasitica (Murr.) Barr. Virulence on apples and chestnut trees was reduced in four of six extensively characterized mutants. Relative to the virulent progenitor, the attenuated mutants had reduced growth rates, abnormal colony morphologies, and few asexual spores, and they resembled virus-infected strains. The respiratory defects and attenuated virulence phenotypes (hypovirulence) were transmitted from two mutants to a virulent strain by hyphal contact. The infectious transmission of hypovirulence occurred independently of the transfer of nuclei, did not involve a virus, and dynamically reflects fungal diseases caused by mitochondrial mutations. In these mutants, mitochondrial mutations are further implicated in generation of the attenuated state by (i) uniparental (maternal) inheritance of the trait, (ii) presence of high levels of cyanide-insensitive mitochondrial alternative oxidase activity, (iii) cytochrome deficiencies, and (iv) structural abnormalities in the mtDNA. Hence, cytoplasmically transmissible hypovirulence phenotypes found in virus-free strains of C. parasitica from recovering trees may be caused by mutant forms of mtDNA. Images Fig. 2 Fig. 4 PMID:11607549

Diversity of Marine-Derived Aspergillus from Tidal Mudflats and Sea Sand in Korea

PubMed Central

Lee, Seobihn; Park, Myung Soo

2016-01-01

Aspergillus (Trichocomaceae, Eurotiales, and Ascomycota) is a genus of well-defined asexual spore-forming fungi that produce valuable compounds such as secondary metabolites and enzymes; however, some species are also responsible for diseases in plants and animals, including humans. To date, 26 Aspergillus species have been reported in Korea, with most species located in terrestrial environments. In our study, Aspergillus species were isolated from mudflats and sea sand along the western and southern coasts of Korea. A total of 84 strains were isolated and identified as 17 Aspergillus species in 11 sections on the basis of both morphological characteristics and sequence analysis of the calmodulin gene (CaM) locus. Commonly isolated species were A. fumigatus (26 strains), A. sydowii (14 strains), and A. terreus (10 strains). The diversity of Aspergillus species isolated from mudflats (13 species) was higher than the diversity of those from sea sand (five species). Four identified species—A. caesiellus, A. montenegroi, A. rhizopodus, and A. tabacinus—are in the first records in Korea. Here, we provide detailed descriptions of the morphological characteristics of these four species. PMID:28154481

Diapause and maintenance of facultative sexual reproductive strategies

PubMed Central

Lehtonen, Jussi

2016-01-01

Facultative sex combines sexual and asexual reproduction in the same individual (or clone) and allows for a large diversity of life-history patterns regarding the timing, frequency and intensity of sexual episodes. In addition, other life-history traits such as a diapause stage may become linked to sex. Here, we develop a matrix modelling framework for addressing the cost of sex in facultative sexuals, in constant, periodic and stochastically fluctuating environments. The model is parametrized using life-history data from Brachionus calyciflorus, a facultative sexual rotifer in which sex and diapause are linked. Sexual propensity was an important driver of costs in constant environments, in which high costs (always > onefold, and sometimes > twofold) indicated that asexuals should outcompete facultative sexuals. By contrast, stochastic environments with high temporal autocorrelation favoured facultative sex over obligate asex, in particular, if the penalty to fecundity in ‘bad’ environments was large. In such environments, obligate asexuals were constrained by their life cycle length (i.e. time from birth to last reproductive adult age class), which determined an upper limit to the number of consecutive bad periods they could tolerate. Nevertheless, when facultative asexuals with different sexual propensities competed simultaneously against each other and asex, the lowest sex propensity was the most successful in stochastic environments with positive autocorrelation. Our results suggest that a highly specific mechanism (i.e. diapause linked to sex) can alone stabilize facultative sex in these animals, and protect it from invasion of both asexual and pure sexual strategies. This article is part of the themed issue ‘Weird sex: the underappreciated diversity of sexual reproduction’. PMID:27619700

Benefits of a Recombination-Proficient Escherichia coli System for Adaptive Laboratory Evolution.

PubMed

Peabody, George; Winkler, James; Fountain, Weston; Castro, David A; Leiva-Aravena, Enzo; Kao, Katy C

2016-11-15

Adaptive laboratory evolution typically involves the propagation of organisms asexually to select for mutants with the desired phenotypes. However, asexual evolution is prone to competition among beneficial mutations (clonal interference) and the accumulation of hitchhiking and neutral mutations. The benefits of horizontal gene transfer toward overcoming these known disadvantages of asexual evolution were characterized in a strain of Escherichia coli engineered for superior sexual recombination (genderless). Specifically, we experimentally validated the capacity of the genderless strain to reduce the mutational load and recombine beneficial mutations. We also confirmed that inclusion of multiple origins of transfer influences both the frequency of genetic exchange throughout the chromosome and the linkage of donor DNA. We built a simple kinetic model to estimate recombination frequency as a function of transfer size and relative genotype enrichment in batch transfers; the model output correlated well with the experimental data. Our results provide strong support for the advantages of utilizing the genderless strain over its asexual counterpart during adaptive laboratory evolution for generating beneficial mutants with reduced mutational load. Over 80 years ago Fisher and Muller began a debate on the origins of sexual recombination. Although many aspects of sexual recombination have been examined at length, experimental evidence behind the behaviors of recombination in many systems and the means to harness it remain elusive. In this study, we sought to experimentally validate some advantages of recombination in typically asexual Escherichia coli and determine if a sexual strain of E. coli can become an effective tool for strain development. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

Benefits of a Recombination-Proficient Escherichia coli System for Adaptive Laboratory Evolution

PubMed Central

Peabody, George; Winkler, James; Fountain, Weston; Castro, David A.; Leiva-Aravena, Enzo

2016-01-01

ABSTRACT Adaptive laboratory evolution typically involves the propagation of organisms asexually to select for mutants with the desired phenotypes. However, asexual evolution is prone to competition among beneficial mutations (clonal interference) and the accumulation of hitchhiking and neutral mutations. The benefits of horizontal gene transfer toward overcoming these known disadvantages of asexual evolution were characterized in a strain of Escherichia coli engineered for superior sexual recombination (genderless). Specifically, we experimentally validated the capacity of the genderless strain to reduce the mutational load and recombine beneficial mutations. We also confirmed that inclusion of multiple origins of transfer influences both the frequency of genetic exchange throughout the chromosome and the linkage of donor DNA. We built a simple kinetic model to estimate recombination frequency as a function of transfer size and relative genotype enrichment in batch transfers; the model output correlated well with the experimental data. Our results provide strong support for the advantages of utilizing the genderless strain over its asexual counterpart during adaptive laboratory evolution for generating beneficial mutants with reduced mutational load. IMPORTANCE Over 80 years ago Fisher and Muller began a debate on the origins of sexual recombination. Although many aspects of sexual recombination have been examined at length, experimental evidence behind the behaviors of recombination in many systems and the means to harness it remain elusive. In this study, we sought to experimentally validate some advantages of recombination in typically asexual Escherichia coli and determine if a sexual strain of E. coli can become an effective tool for strain development. PMID:27613685

Gravity regulation in tuber-bearing moss Leptobryum pyriforme (Hedw.) Wilson

NASA Astrophysics Data System (ADS)

Lobachevska, Oksana

Considerable number of moss species is propagated asexually, and asexual reproduction is the key factor of their life strategy and effective mechanism of rapid population and attaching plants to habitats with great environmental fluctuations (Velde et al., 2001; Frey, Kűrshner, 2010). It has been shown for the first time for gravisensitive species Leptobryum pyriforme (Hedw.) Wilson that the development of propagules as organs of vegetative reproduction and accumulation of nutrient substances is gravidependent phenomenon. L. pyriforme differs from other moss species in higher growth and development rate. In darkness the greatest bundle of gravisensitive negatively gravitropic filaments (above 50 filaments) of both caulonemal and chloronemal type arised from 1 protonemal ball of moss. Perhaps, it is caused by high protonema gravisensitivity and morphogenetic effectiveness of gravitation force. It has been shown that propagules of L. pyriforme are formed much faster in darkness and their number is twice higher than on light. After five-day clinorotation of the L. pyriforme turfs the number of propagules is lower in darkness compared to gravistimulated turfs and higher than on the light. Thus, vegetative reproduction of L. pyriforme is the gravidependent process and gravitation force has stimulating influence on the formation of propagula. In L. pyriforme rhizoid tubers from round to oval (93-116 x ({) } (x) 120-148 muμm) are formed from 5-6 big cells (70 x ({) } (x) 80 muμm). Due to small capsules, L{it pyriforme }does not have a lot of big spores which are spread to insignificant distances, the mass formation of brood organs promotes moss survival and its preservation. The results of investigation prove the participation of rhizoids and rhizoid tubers as imperceptible but important phase of vital cycle of moss species - settlers in realization of vital tolerance strategy to extreme conditions of temporarily available habitats: due to rapid method of spatial distribution, to store considerable genetic variability and long term preservation of viable diasporas bank as a “genetic memory”. It also has been shown that spiral growth, which still had been found only for Ceratodon purpureus (Hedw.) Brid. under the conditions of microgravitation and for Barbula unguiculata Hedw. for 1 g (Kern et al., 2005; Demkiv et al., 2006) is also characteristic of protonemata turf L. pyriforme. Considering the fact that spirality is generally accepted biological phenomenon, it is possible to find the growth spiral movements under certain ecological conditions and various stages of moss development. Thus, the space orientation of moss organs is gravidependent process, and gravimorphogenesis is precondition to change growth model at various ontogenesis stages.

A Standard Method To Inactivate Bacillus anthracis Spores to Sterility via Gamma Irradiation

PubMed Central

Cote, Christopher K.; Buhr, Tony; Bernhards, Casey B.; Bohmke, Matthew D.; Calm, Alena M.; Esteban-Trexler, Josephine S.; Hunter, Melissa; Katoski, Sarah E.; Kennihan, Neil; Klimko, Christopher P.; Miller, Jeremy A.; Minter, Zachary A.; Pfarr, Jerry W.; Prugh, Amber M.; Quirk, Avery V.; Rivers, Bryan A.; Shea, April A.; Shoe, Jennifer L.; Sickler, Todd M.; Young, Alice A.; Fetterer, David P.; Welkos, Susan L.; McPherson, Derrell; Fountain, Augustus W.

2018-01-01

ABSTRACT In 2015, a laboratory of the United States Department of Defense (DoD) inadvertently shipped preparations of gamma-irradiated spores of Bacillus anthracis that contained live spores. In response, a systematic evidence-based method for preparing, concentrating, irradiating, and verifying the inactivation of spore materials was developed. We demonstrate the consistency of spore preparations across multiple biological replicates and show that two different DoD institutions independently obtained comparable dose-inactivation curves for a monodisperse suspension of B. anthracis spores containing 3 × 1010 CFU. Spore preparations from three different institutions and three strain backgrounds yielded similar decimal reduction (D10) values and irradiation doses required to ensure sterility (DSAL) to the point at which the probability of detecting a viable spore is 10−6. Furthermore, spores of a genetically tagged strain of B. anthracis strain Sterne were used to show that high densities of dead spores suppress the recovery of viable spores. Together, we present an integrated method for preparing, irradiating, and verifying the inactivation of spores of B. anthracis for use as standard reagents for testing and evaluating detection and diagnostic devices and techniques. IMPORTANCE The inadvertent shipment by a U.S. Department of Defense (DoD) laboratory of live Bacillus anthracis (anthrax) spores to U.S. and international destinations revealed the need to standardize inactivation methods for materials derived from biological select agents and toxins (BSAT) and for the development of evidence-based methods to prevent the recurrence of such an event. Following a retrospective analysis of the procedures previously employed to generate inactivated B. anthracis spores, a study was commissioned by the DoD to provide data required to support the production of inactivated spores for the biodefense community. The results of this work are presented in this publication, which details the method by which spores can be prepared, irradiated, and tested, such that the chance of finding residual living spores in any given preparation is 1/1,000,000. These irradiated spores are used to test equipment and methods for the detection of agents of biological warfare and bioterrorism. PMID:29654186

Coalescence with Background and Balancing Selection in Systems with Bi- and Uniparental Reproduction: Contrasting Partial Asexuality and Selfing

PubMed Central

Agrawal, Aneil F.; Hartfield, Matthew

2016-01-01

Uniparental reproduction in diploids, via asexual reproduction or selfing, reduces the independence with which separate loci are transmitted across generations. This is expected to increase the extent to which a neutral marker is affected by selection elsewhere in the genome. Such effects have previously been quantified in coalescent models involving selfing. Here we examine the effects of background selection and balancing selection in diploids capable of both sexual and asexual reproduction (i.e., partial asexuality). We find that the effect of background selection on reducing coalescent time (and effective population size) can be orders of magnitude greater when rates of sex are low than when sex is common. This is because asexuality enhances the effects of background selection through both a recombination effect and a segregation effect. We show that there are several reasons that the strength of background selection differs between systems with partial asexuality and those with comparable levels of uniparental reproduction via selfing. Expectations for reductions in Ne via background selection have been verified using stochastic simulations. In contrast to background selection, balancing selection increases the coalescence time for a linked neutral site. With partial asexuality, the effect of balancing selection is somewhat dependent upon the mode of selection (e.g., heterozygote advantage vs. negative frequency-dependent selection) in a manner that does not apply to selfing. This is because the frequency of heterozygotes, which are required for recombination onto alternative genetic backgrounds, is more dependent on the pattern of selection with partial asexuality than with selfing. PMID:26584901

Coalescence with Background and Balancing Selection in Systems with Bi- and Uniparental Reproduction: Contrasting Partial Asexuality and Selfing.

PubMed

Agrawal, Aneil F; Hartfield, Matthew

2016-01-01

Uniparental reproduction in diploids, via asexual reproduction or selfing, reduces the independence with which separate loci are transmitted across generations. This is expected to increase the extent to which a neutral marker is affected by selection elsewhere in the genome. Such effects have previously been quantified in coalescent models involving selfing. Here we examine the effects of background selection and balancing selection in diploids capable of both sexual and asexual reproduction (i.e., partial asexuality). We find that the effect of background selection on reducing coalescent time (and effective population size) can be orders of magnitude greater when rates of sex are low than when sex is common. This is because asexuality enhances the effects of background selection through both a recombination effect and a segregation effect. We show that there are several reasons that the strength of background selection differs between systems with partial asexuality and those with comparable levels of uniparental reproduction via selfing. Expectations for reductions in Ne via background selection have been verified using stochastic simulations. In contrast to background selection, balancing selection increases the coalescence time for a linked neutral site. With partial asexuality, the effect of balancing selection is somewhat dependent upon the mode of selection (e.g., heterozygote advantage vs. negative frequency-dependent selection) in a manner that does not apply to selfing. This is because the frequency of heterozygotes, which are required for recombination onto alternative genetic backgrounds, is more dependent on the pattern of selection with partial asexuality than with selfing. Copyright © 2016 by the Genetics Society of America.

A Perspective on the Importance of Reproductive Mode in Astrobiology

NASA Astrophysics Data System (ADS)

Van Doninck, Karine; Schön, Isa; Martens, Koen

2003-12-01

Reproduction is a vital characteristic of life, and sex is the most common reproductive mode in the eukaryotic world. Sex and reproduction are not necessarily linked mechanisms: Sexuality without reproduction exists, while several forms of asexual reproduction are known. The occurrence of sexuality itself is paradoxical, as it is very costly in evolutionary terms. Most of the hypotheses (more than 20) attempting to explain the prevalence of sex fall into two categories: Sex either creates good gene combinations for adaptation to environments or eliminates bad gene combinations counteracting the accumulation of mutations. In spite of this apparent wealth of beneficial effects of sex, asexuality is not rare. Most eukaryotic, asexual lineages are short-lived and can only persist through the presence of sexual roots, but at least two animal groups, bdelloid rotifers and darwinulid ostracods, seem to claim the status of ancient asexuals. Research on (a)sexuality is relevant to astrobiology in a number of ways. First, strong relationships between the origin and persistence of life in extreme environments and reproductive mode are known. Second, the "habitability" of nonterrestrial environments to life greatly depends on reproductive mode. Whereas asexuals can do equally well or better in harsh environments, they fail to adapt fast enough to changing abiotic and biotic environments. Third, it has been shown that plants reproduce mainly asexually in space, and sperm production and motility in some vertebrates are hampered. Both findings indicate that extraterrestrial life under conditions different from Earth might be dominated by asexual reproduction. Finally, for exchange of biological material between planets, the choice of reproductive mode will be important.

Asexual Reproduction Does Not Apparently Increase the Rate of Chromosomal Evolution: Karyotype Stability in Diploid and Triploid Clonal Hybrid Fish (Cobitis, Cypriniformes, Teleostei)

PubMed Central

Majtánová, Zuzana; Choleva, Lukáš; Symonová, Radka; Ráb, Petr; Kotusz, Jan; Pekárik, Ladislav; Janko, Karel

2016-01-01

Interspecific hybridization, polyploidization and transitions from sexuality to asexuality considerably affect organismal genomes. Especially the last mentioned process has been assumed to play a significant role in the initiation of chromosomal rearrangements, causing increased rates of karyotype evolution. We used cytogenetic analysis and molecular dating of cladogenetic events to compare the rate of changes of chromosome morphology and karyotype in asexually and sexually reproducing counterparts in European spined loach fish (Cobitis). We studied metaphases of three sexually reproducing species and their diploid and polyploid hybrid clones of different age of origin. The material includes artificial F1 hybrid strains, representatives of lineage originated in Holocene epoch, and also individuals of an oldest known age to date (roughly 0.37 MYA). Thereafter we applied GISH technique as a marker to differentiate parental chromosomal sets in hybrids. Although the sexual species accumulated remarkable chromosomal rearrangements after their speciation, we observed no differences in chromosome numbers and/or morphology among karyotypes of asexual hybrids. These hybrids possess chromosome sets originating from respective parental species with no cytogenetically detectable recombinations, suggesting their integrity even in a long term. The switch to asexual reproduction thus did not provoke any significant acceleration of the rate of chromosomal evolution in Cobitis. Asexual animals described in other case studies reproduce ameiotically, while Cobitis hybrids described here produce eggs likely through modified meiosis. Therefore, our findings indicate that the effect of asexuality on the rate of chromosomal change may be context-dependent rather than universal and related to particular type of asexual reproduction. PMID:26808475

Asexual Reproduction Does Not Apparently Increase the Rate of Chromosomal Evolution: Karyotype Stability in Diploid and Triploid Clonal Hybrid Fish (Cobitis, Cypriniformes, Teleostei).

PubMed

Majtánová, Zuzana; Choleva, Lukáš; Symonová, Radka; Ráb, Petr; Kotusz, Jan; Pekárik, Ladislav; Janko, Karel

2016-01-01

Interspecific hybridization, polyploidization and transitions from sexuality to asexuality considerably affect organismal genomes. Especially the last mentioned process has been assumed to play a significant role in the initiation of chromosomal rearrangements, causing increased rates of karyotype evolution. We used cytogenetic analysis and molecular dating of cladogenetic events to compare the rate of changes of chromosome morphology and karyotype in asexually and sexually reproducing counterparts in European spined loach fish (Cobitis). We studied metaphases of three sexually reproducing species and their diploid and polyploid hybrid clones of different age of origin. The material includes artificial F1 hybrid strains, representatives of lineage originated in Holocene epoch, and also individuals of an oldest known age to date (roughly 0.37 MYA). Thereafter we applied GISH technique as a marker to differentiate parental chromosomal sets in hybrids. Although the sexual species accumulated remarkable chromosomal rearrangements after their speciation, we observed no differences in chromosome numbers and/or morphology among karyotypes of asexual hybrids. These hybrids possess chromosome sets originating from respective parental species with no cytogenetically detectable recombinations, suggesting their integrity even in a long term. The switch to asexual reproduction thus did not provoke any significant acceleration of the rate of chromosomal evolution in Cobitis. Asexual animals described in other case studies reproduce ameiotically, while Cobitis hybrids described here produce eggs likely through modified meiosis. Therefore, our findings indicate that the effect of asexuality on the rate of chromosomal change may be context-dependent rather than universal and related to particular type of asexual reproduction.

Anthrax Spores under a microscope

NASA Technical Reports Server (NTRS)

2003-01-01

Anthrax spores are inactive forms of Bacillus anthracis. They can survive for decades inside a spore's tough protective coating; they become active when inhaled by humans. A result of NASA- and industry-sponsored research to develop small greenhouses for space research is the unique AiroCide TiO2 system that kills anthrax spores and other pathogens.

Surface tension propulsion of fungal spores by use of microdroplets

NASA Astrophysics Data System (ADS)

Noblin, Xavier; Yang, Sylvia; Dumais, Jacques

2010-11-01

Most basidiomycete fungi (such as edible mushrooms) actively eject their spores. The process begins with the condensation of a water droplet at the base of the spore. The fusion of the droplet onto the spore creates a momentum that propels the spore forward. The use of surface tension for spore ejection offers a new paradigm to perform work at small length scales. However, this mechanism of force generation remains poorly understood. To elucidate how fungal spores make effective use of surface tension, we performed high-speed video imaging of spore ejection in Auricularia auricula and Sporobolomyces yeast, along with a detailed mechanical analysis of the spore ejection. We developed an explicit relation for the conversion of surface energy into kinetic energy during the coalescence process. The relation was validated with a simple artificial system.

FigA, a putative homolog of low-affinity calcium system member Fig1 in Saccharomyces cerevisiae, is involved in growth and asexual and sexual development in Aspergillus nidulans.

PubMed

Zhang, Shizhu; Zheng, Hailin; Long, Nanbiao; Carbó, Natalia; Chen, Peiying; Aguilar, Pablo S; Lu, Ling

2014-02-01

Calcium-mediated signaling pathways are widely employed in eukaryotes and are implicated in the regulation of diverse biological processes. In Saccharomyces cerevisiae, at least two different calcium uptake systems have been identified: the high-affinity calcium influx system (HACS) and the low-affinity calcium influx system (LACS). Compared to the HACS, the LACS in fungi is not well known. In this study, FigA, a homolog of the LACS member Fig1 from S. cerevisiae, was functionally characterized in the filamentous fungus Aspergillus nidulans. Loss of figA resulted in retardant hyphal growth and a sharp reduction of conidial production. Most importantly, FigA is essential for the homothallic mating (self-fertilization) process; further, FigA is required for heterothallic mating (outcrossing) in the absence of HACS midA. Interestingly, in a figA deletion mutant, adding extracellular Ca(2+) rescued the hyphal growth defects but could not restore asexual and sexual reproduction. Furthermore, quantitative PCR results revealed that figA deletion sharply decreased the expression of brlA and nsdD, which are known as key regulators during asexual and sexual development, respectively. In addition, green fluorescent protein (GFP) tagging at the C terminus of FigA (FigA::GFP) showed that FigA localized to the center of the septum in mature hyphal cells, to the location between vesicles and metulae, and between the junctions of metulae and phialides in conidiophores. Thus, our findings suggest that FigA, apart from being a member of a calcium uptake system in A. nidulans, may play multiple unexplored roles during hyphal growth and asexual and sexual development.

Efficacy of propidium iodide and FUN-1 stains for assessing viability in basidiospores of Rhizopogon roseolus.

PubMed

Fernández-Miranda, Elena; Majada, Juan; Casares, Abelardo

2017-01-01

The use of spores in applications of ectomycorrhizal fungi requires information regarding spore viability and germination, especially in genera such as Rhizopogon with high rates of spore dormancy. The authors developed a protocol to assess spore viability of Rhizopogon roseolus using four vital stains to quantify spore viability and germination and to optimize storage procedures. They showed that propidium iodide is an excellent stain for quantifying nonviable spores. Observing red fluorescent intravacuolar structures following staining with 2-chloro-4-(2,3-dihydro-3-methyl-(benzo-1,3-thiazol-2-yl)-methylidene)-1-phenylquinolinium iodide (FUN-1) can help identify viable spores that are activated. At 6 mo and 1 y, the spores kept in a water suspension survived better than those left within intact, dry gasterocarps. Our work highlights the importance of temperature, nutrients, and vitamins for maturation and germination of spores of R. roseolus during 1 y of storage.

Tests of the relative roles of calcium channels and calcium pumps in controlling gravity-directed development in single spore cells of the fern Ceratopteris richardii

NASA Astrophysics Data System (ADS)

Roux, Stanley; Porterfield, D. Marshall; Haque, Aeraj Ul; Bushart, Thomas

The vector of gravity sets the direction of polarized development of single spore cells of the fern Ceratopteris richardii after light initiates their germination. Gravity also sets the direction of a trans-cell calcium current, which enters the cell along its bottom and exits it from its top. The direction of this current predicts the subsequent direction of spore development, and blocking this current with calcium channel blockers randomizes the direction of subsequent development. Recently the laboratory of D. Marshall Porterfield (Purdue University) developed a microchip device that can measure the direction and magnitude of the trans-spore calcium current in real time. Our laboratory in collaboration with Porterfield's recently found that this current inverts rapidly when the cells are turned upside down and that the magnitude of the current rises and falls with the magnitude of the g-force when these cells are tested in parabolic flight on the DC-9 aircraft. We assume that the gravity-directed entry of calcium into these cells is through calcium channels and its exit is through calcium pumps. Here we report our studies of a calcium pump that is highly expressed in the spores during the period when gravity is setting the direction of the calcium current, and we describe pharmacological tests of the relative importance of calcium pumps in maintaining the calcium current and in controlling the direction of subsequent spore development. We found that inhibitors that block the activity of calcium pumps also greatly depress the trans-cell current, but, surprisingly, have little effect on the ability of gravity to set the direction of spore development. These results, in combination with earlier findings, indicate that the gravity-directed opening of calcium channels along the bottom of spore cells plays a more important role in directing subsequent spore development than the activity of calcium pumps, despite the importance of these pumps in maintaining the trans-cell calcium current. Supported by NASA grants NAG2-1586 and NAG10-295 to S. J. R.

Does the silver moss Bryum argenteum exhibit sex-specific patterns in vegetative growth rate, asexual fitness or prezygotic reproductive investment?

PubMed

Horsley, Kimberly; Stark, Lloyd R; McLetchie, D Nicholas

2011-05-01

Expected life history trade-offs associated with sex differences in reproductive investment are often undetected in seed plants, with the difficulty arising from logistical issues of conducting controlled experiments. By controlling genotype, age and resource status of individuals, a bryophyte was assessed for sex-specific and location-specific patterns of vegetative, asexual and sexual growth/reproduction across a regional scale. Twelve genotypes (six male, six female) of the dioecious bryophyte Bryum argenteum were subcultured to remove environmental effects, regenerated asexually to replicate each genotype 16 times, and grown over a period of 92 d. Plants were assessed for growth rates, asexual and sexual reproductive traits, and allocation to above- and below-ground regenerative biomass. The degree of sexual versus asexual reproductive investment appears to be under genetic control, with three distinct ecotypes found in this study. Protonemal growth rate was positively correlated with asexual reproduction and sexual reproduction, whereas asexual reproduction was negatively correlated (appeared to trade-off) with vegetative growth (shoot production). No sex-specific trade-offs were detected. Female sex-expressing shoots were longer than males, but the sexes did not differ in growth traits, asexual traits, sexual induction times, or above- and below-ground biomass. Males, however, had much higher rates of inflorescence production than females, which translated into a significantly higher (24x) prezygotic investment for males relative to females. Evidence for three distinct ecotypes is presented for a bryophyte based on regeneration traits. Prior to zygote production, the sexes of this bryophyte did not differ in vegetative growth traits but significantly differed in reproductive investment, with the latter differences potentially implicated in the strongly biased female sex ratio. The disparity between males and females for prezygotic reproductive investment is the highest known for bryophytes.

Requirements for the Development of Bacillus Anthracis Spore Reference Materials Used to Test Detection Systems

PubMed Central

Almeida, Jamie L.; Wang, Lili; Morrow, Jayne B.; Cole, Kenneth D.

2006-01-01

Bacillus anthracis spores have been used as biological weapons and the possibility of their further use requires surveillance systems that can accurately and reliably detect their presence in the environment. These systems must collect samples from a variety of matrices, process the samples, and detect the spores. The processing of the sample may include removal of inhibitors, concentration of the target, and extraction of the target in a form suitable for detection. Suitable reference materials will allow the testing of each of these steps to determine the sensitivity and specificity of the detection systems. The development of uniform and well-characterized reference materials will allow the comparison of different devices and technologies as well as assure the continued performance of detection systems. This paper discusses the special requirements of reference materials for Bacillus anthracis spores that could be used for testing detection systems. The detection of Bacillus anthracis spores is based on recognition of specific characteristics (markers) on either the spore surface or in the nucleic acids (DNA). We have reviewed the specific markers and their relevance to characterization of reference materials. We have also included the approach for the characterization of candidate reference materials that we are developing at the NIST laboratories. Additional applications of spore reference materials would include testing sporicidal treatments, techniques for sampling the environment, and remediation of spore-contaminated environments. PMID:27274929

Phenotypic divergence along geographic gradients reveals potential for rapid adaptation of the White-nose Syndrome pathogen, Pseudogymnoascus destructans, in North America.

PubMed

Forsythe, Adrian; Giglio, Victoria; Asa, Jonathan; Xu, Jianping

2018-06-18

White-nose Syndrome (WNS) is an ongoing epizootic affecting multiple species of North American bats, caused by epidermal infections of the psychrophilic filamentous fungus, Pseudogymnoascus destructans Since its introduction from Europe, WNS has spread rapidly across eastern North America and resulted in high mortality rates in bats. At present, the mechanisms behind its spread and the extent of its adaptation to different geographic and ecological niches remain unknown. The objective of this study was to examine the geographic patterns of phenotypic variation and the potential evidence for adaptation among strains representing broad geographic locations in eastern North America. The morphological features of these strains were evaluated on artificial medium, and the viability of asexual arthroconidia of representative strains were investigated after storage at high (23°C), moderate (14°C), and low (4°C) temperatures at different lengths of times. Our analyses identified evidence for a geographic pattern of colony morphology changes among the clonal descendants of the fungus, with trait values correlated with increased distance from the epicenter of WNS. Our genomic comparisons of three representative isolates revealed novel genetic polymorphisms and suggested potential candidate mutations that might be related to some of the phenotypic changes. These results show that even though this pathogen arrived in North America only recently and reproduces asexually, there has been substantial evolution and phenotypic diversification during its rapid clonal expansion. Importance The causal agent of White-nose Syndrome in bats is Pseudogymnoascus destructans , a filamentous fungus recently introduced from its native range in Europe. Infections caused by P. destructans have progressed across the eastern parts of Canada and the United States over the last ten years. It is not clear how the disease is spread as the pathogen is unable to grow above 23°C and ambient temperature can act as a barrier when hosts disperse. Here, we explore the patterns of phenotypic diversity and the germination of the fungal asexual spores, arthroconidia, from strains across a sizeable area of the epizootic range. Our analyses revealed evidence of adaptation along geographic gradients during its expansion. The results have implications for understanding the diversification of P. destructans and the limits of WNS spread in North America. Given the rapidly expanding distribution of WNS, a detailed understanding of the genetic bases for phenotypic variations in growth, reproduction, and dispersal of P. destructans are urgently needed in order to help control this disease. Copyright © 2018 American Society for Microbiology.

Bacillus spore-based oral carriers loading curcumin for the therapy of colon cancer.

PubMed

Yin, Liang; Meng, Zhan; Zhang, Yuxiao; Hu, Kaikai; Chen, Wuya; Han, Kaibin; Wu, Bao-Yan; You, Rong; Li, Chu-Hua; Jin, Ying; Guan, Yan-Qing

2018-02-10

Oral drug delivery has attracted substantial attention due to its advantages over other administration routes. Bacillus spores, as oral probiotic agents, are applied widely. In this paper, a novel Bacillus spore-based oral colon targeted carrier loading curcumin was developed for colon cancer treatment. Curcumin was linked covalently with the outer coat of Bacillus spore and folate, respectively (SPORE-CUR-FA). Bacillus spores are capable of delivering drugs to the colon area through gastric barrier, taking the advantage of its tolerance to the harsh conditions and disintegration of the outer coat of spores after germination in the colon. The drug release in vitro and in vivo of SPORE-CUR-FA was investigated. Results showed that SPORE-CUR-FA had the characteristics of colon-targeted drug release. Pharmacokinetic studies confirmed that Bacillus spore-based carriers could efficiently improve the oral bioavailability of curcumin. In vitro and in vivo anti-tumor studies showed that SPORE-CUR-FA had substantial ability for inhibiting colon cancer cells. These findings suggest that this Bacillus spore-based oral drug delivery system has a great potential for the treatment of colon cancer. Copyright © 2017 Elsevier B.V. All rights reserved.

Gonad establishment during asexual reproduction in the annelid Pristina leidyi.

PubMed

Özpolat, B Duygu; Bely, Alexandra E

2015-09-01

Animals that can reproduce by both asexual agametic reproduction and sexual reproduction must transmit or re-establish their germ line post-embryonically. Although such a dual reproductive mode has evolved repeatedly among animals, how asexually produced individuals establish their germ line remains poorly understood in most groups. We investigated germ line development in the annelid Pristina leidyi, a species that typically reproduces asexually by paratomic fission, intercalating a new tail and head in the middle of the body followed by splitting. We found that in fissioning individuals, gonads occur in anterior segments in the anterior-most individual as well as in new heads forming within fission zones. Homologs of the germ line/multipotency genes piwi, vasa, and nanos are expressed in the gonads, as well as in proliferative tissues including the posterior growth zone, fission zone, and regeneration blastema. In fissioning animals, certain cells on the ventral nerve cord express a homolog of piwi, are abundant near fission zones, and sometimes make contact with gonads. Such cells are typically undetectable near the blastema and posterior growth zone. Time-lapse imaging provides direct evidence that cells on the ventral nerve cord migrate preferentially towards fission zones. Our findings indicate that gonads form routinely in fissioning individuals, that a population of piwi-positive cells on the ventral nerve cord is associated with fission and gonads, and that cells resembling these piwi-positive cells migrate along the ventral nerve cord. We suggest that the piwi-positive ventral cells are germ cells that transmit the germ line across asexually produced individuals via migration along the ventral nerve cord. Copyright © 2015 Elsevier Inc. All rights reserved.

Adaptations to different habitats in sexual and asexual populations of parasitoid wasps: a meta-analysis.

PubMed

Amat, Isabelle; van Alphen, Jacques J M; Kacelnik, Alex; Desouhant, Emmanuel; Bernstein, Carlos

2017-01-01

Coexistence of sexual and asexual populations remains a key question in evolutionary ecology. We address the question how an asexual and a sexual form of the parasitoid Venturia canescens can coexist in southern Europe. We test the hypothesis that both forms are adapted to different habitats within their area of distribution. Sexuals inhabit natural environments that are highly unpredictable, and where density of wasps and their hosts is low and patchily distributed. Asexuals instead are common in anthropic environments (e.g., grain stores) where host outbreaks offer periods when egg-load is the main constraint on reproductive output. We present a meta-analysis of known adaptations to these habitats. Differences in behavior, physiology and life-history traits between sexual and asexual wasps were standardized in term of effect size (Cohen's d value; Cohen, 1988). Seeking consilience from the differences between multiple traits, we found that sexuals invest more in longevity at the expense of egg-load, are more mobile, and display higher plasticity in response to thermal variability than asexual counterparts. Thus, each form has consistent multiple adaptations to the ecological circumstances in the contrasting environments.

Neutral and selection-driven decay of sexual traits in asexual stick insects

PubMed Central

Schwander, Tanja; Crespi, Bernard J.; Gries, Regine; Gries, Gerhard

2013-01-01

Environmental shifts and lifestyle changes may result in formerly adaptive traits becoming non-functional or maladaptive. The subsequent decay of such traits highlights the importance of natural selection for adaptations, yet its causes have rarely been investigated. To study the fate of formerly adaptive traits after lifestyle changes, we evaluated sexual traits in five independently derived asexual lineages, including traits that are specific to males and therefore not exposed to selection. At least four of the asexual lineages retained the capacity to produce males that display normal courtship behaviours and are able to fertilize eggs of females from related sexual species. The maintenance of male traits may stem from pleiotropy, or from these traits only regressing via drift, which may require millions of years to generate phenotypic effects. By contrast, we found parallel decay of sexual traits in females. Asexual females produced altered airborne and contact signals, had modified sperm storage organs, and lost the ability to fertilize their eggs, impeding reversals to sexual reproduction. Female sexual traits were decayed even in recently derived asexuals, suggesting that trait changes following the evolution of asexuality, when they occur, proceed rapidly and are driven by selective processes rather than drift. PMID:23782880

Adaptations to different habitats in sexual and asexual populations of parasitoid wasps: a meta-analysis

PubMed Central

van Alphen, Jacques J.M.; Kacelnik, Alex; Desouhant, Emmanuel

2017-01-01

Background Coexistence of sexual and asexual populations remains a key question in evolutionary ecology. We address the question how an asexual and a sexual form of the parasitoid Venturia canescens can coexist in southern Europe. We test the hypothesis that both forms are adapted to different habitats within their area of distribution. Sexuals inhabit natural environments that are highly unpredictable, and where density of wasps and their hosts is low and patchily distributed. Asexuals instead are common in anthropic environments (e.g., grain stores) where host outbreaks offer periods when egg-load is the main constraint on reproductive output. Methods We present a meta-analysis of known adaptations to these habitats. Differences in behavior, physiology and life-history traits between sexual and asexual wasps were standardized in term of effect size (Cohen’s d value; Cohen, 1988). Results Seeking consilience from the differences between multiple traits, we found that sexuals invest more in longevity at the expense of egg-load, are more mobile, and display higher plasticity in response to thermal variability than asexual counterparts. Discussion Thus, each form has consistent multiple adaptations to the ecological circumstances in the contrasting environments. PMID:28924495

Asexual Reproduction in Holothurians

PubMed Central

Dolmatov, Igor Yu.

2014-01-01

Aspects of asexual reproduction in holothurians are discussed. Holothurians are significant as fishery and aquaculture items and have high commercial value. The last review on holothurian asexual reproduction was published 18 years ago and included only 8 species. An analysis of the available literature shows that asexual reproduction has now been confirmed in 16 holothurian species. Five additional species are also most likely capable of fission. The recent discovery of new fissiparous holothurian species indicates that this reproduction mode is more widespread in Holothuroidea than previously believed. New data about the history of the discovery of asexual reproduction in holothurians, features of fission, and regeneration of anterior and posterior fragments are described here. Asexual reproduction is obviously controlled by the integrated systems of the organism, primarily the nervous system. Special molecular mechanisms appear to determine the location where fission occurs along the anterior-posterior axis of the body. Alteration of the connective tissue strength of the body wall may play an important role during fission of holothurians. The basic mechanism of fission is the interaction of matrix metalloproteinases, their inhibitors, and enzymes forming cross-link complexes between fibrils of collagen. The population dynamics of fissiparous holothurians are discussed. PMID:25405228

Trace detection of meglumine and diatrizoate from Bacillus spore samples using liquid chromatography/mass spectrometry.

PubMed

Swider, Catherine; Maguire, Kelly; Rickenbach, Michael; Montgomery, Madeline; Ducote, Matthew J; Marhefka, Craig A

2012-07-01

Following the September 11, 2001 terrorist attacks, letters containing Bacillus anthracis were distributed through the United States postal system killing five people. A complex forensic investigation commenced to identify the perpetrator of these mailings. A novel liquid chromatography/mass spectrometry protocol for the qualitative detection of trace levels of meglumine and diatrizoate in dried spore preparations of B. anthracis was developed. Meglumine and diatrizoate are components of radiographic imaging products that have been used to purify bacterial spores. Two separate chromatographic assays using multiple mass spectrometric analyses were developed for the detection of meglumine and diatrizoate. The assays achieved limits of detection for meglumine and diatrizoate of 1.00 and 10.0 ng/mL, respectively. Bacillus cereus T strain spores were effectively used as a surrogate for B. anthracis spores during method development and validation. This protocol was successfully applied to limited evidentiary B. anthracis spore material, providing probative information to the investigators. 2012 American Academy of Forensic Sciences. Published 2012. This article is a U.S. Government work and is in the public domain in the U.S.A.

The autophagy-related genes BbATG1 and BbATG8 have different functions in differentiation, stress resistance and virulence of mycopathogen Beauveria bassiana

PubMed Central

Ying, Sheng-Hua; Liu, Jing; Chu, Xin-Ling; Xie, Xue-Qin; Feng, Ming-Guang

2016-01-01

Autophagy-related proteins play significantly different roles in eukaryotes. In the entomopathogenic fungus Beauveria bassiana, autophagy is associated with fungal growth and development. BbATG1 (a serine/threonine protein kinase) and BbATG8 (a ubiquitin-like protein) have similar roles in autophagy, but different roles in other processes. Disruption mutants of BbATG1 and BbATG8 had impaired conidial germination under starvation stress. The mutant ΔBbATG8 exhibited enhanced sensitivity to oxidative stress, while a ΔBbATG1 mutant did not. BbATG1 and BbATG8 showed different roles in spore differentiation. The blastospore yield was reduced by 70% and 92% in ΔBbATG1 and ΔBbATG8 mutants, respectively, and the double mutant had a reduction of 95%. Conidial yield was reduced by approximately 90% and 50% in ΔBbATG1 and ΔBbATG8 mutants, respectively. A double mutant had a reduction similar to ΔBbATG1. Additionally, both BbATG1 and BbATG8 affected the levels of conidial protein BbCP15p required for conidiation. The virulence of each autophagy-deficient mutant was considerably weakened as indicated in topical and intrahemocoel injection assays, and showed a greater reduction in topical infection. However, BbATG1 and BbATG8 had different effects on fungal virulence. Our data indicate that these autophagy-related proteins have different functions in fungal stress response, asexual development and virulence. PMID:27197558

Near-infrared surface-enhanced-Raman-scattering (SERS) mediated identification of single optically trapped, bacterial spores

NASA Astrophysics Data System (ADS)

Alexander, Troy A.; Gillespie, James B.; Pellegrino, Paul M.; Fell, Nicholas F., Jr.; Wood, Gary L.; Salamo, Gregory J.

2003-03-01

A novel methodology has been developed for the investigation of bacterial spores. Specifically, this method has been used to probe the spore coat composition of several Bacillus species. This technique may be useful in many applications; most notably, development of novel detection schemes toward potentially harmful biological agents. This method would also be useful as an ancillary environmental monitoring system where sterility is of importance (i.e., food preparation areas as well as invasive and minimally invasive medical applications). This unique detection scheme is based on the near-infrared (NIR) Surface-Enhanced-Raman-Scattering (SERS) from single, optically trapped, bacterial spores. The SERS spectra of several bacterial spores in aqueous media have been measured using SERS substrates based on 60-nm diameter gold colloids bound to 3-Aminopropyltriethoxysilane derivatized glass. The light from a 785-nm laser diode was used to capture/manipulate as well as simultaneously excite the SERS of an individual bacterial spore. The collected SERS spectra were examined for uniqueness and the applicability of this technique for the species identification of bacterial spores.

Impact of electromagnetic microwaves on the germination of spores of Streptomyces xanthochromogenes in a peat soil and in a liquid nutrient medium

NASA Astrophysics Data System (ADS)

Komarova, A. S.; Likhacheva, A. A.; Lapygina, E. V.; Maksimova, I. A.; Pozdnyakov, A. I.

2010-01-01

The impact of microwaves on the germination of spores of Streptomyces xanthochromogenes in a liquid nutrient medium and in a peat soil was studied. The treatment of inoculums with microwave radiation affected the development of the microorganisms from the stage of spore germination to the stage of the formation of microcolonies of actinomycetes upon the spore cultivation in the liquid medium. Typical hypnum-herbaceous peat was used to study the rate of germination of the actinomycetal spores in soil. The study of the dynamics of the Streptomyces xanthochromogenes population in the control soil (without treatment with microwaves) showed that the most active development of the culture took place in the soil moistened to 60% of the maximum water capacity. When the soil was moistened to the minimum adsorption capacity, the streptomyces did not complete their full cycle of development. The stimulation of the spore germination and mycelium growth with microwaves in the soil medium required a longer period in comparison with that for the liquid medium. The stimulation of the spore germination was observed in the liquid nutrient medium in the case of 30-s treatment and in the soil in the case of 60-s treatment.

Bacterial spore inactivation induced by cold plasma.

PubMed

Liao, Xinyu; Muhammad, Aliyu Idris; Chen, Shiguo; Hu, Yaqin; Ye, Xingqian; Liu, Donghong; Ding, Tian

2018-04-05

Cold plasma has emerged as a non-thermal technology for microbial inactivation in the food industry over the last decade. Spore-forming microorganisms pose challenges for microbiological safety and for the prevention of food spoilage. Inactivation of spores induced by cold plasma has been reported by several studies. However, the exact mechanism of spore deactivation by cold plasma is poorly understood; therefore, it is difficult to control this process and to optimize cold plasma processing for efficient spore inactivation. In this review, we summarize the factors that affect the resistance of spores to cold plasma, including processing parameters, environmental elements, and spore properties. We then describe possible inactivation targets in spore cells (e.g., outer structure, DNA, and metabolic proteins) that associated with inactivation by cold plasma according to previous studies. Kinetic models of the sporicidal activity of cold plasma have also been described here. A better understanding of the interaction between spores and cold plasma is essential for the development and optimization of cold plasma technology in food the industry.

Novel Fungitoxicity Assays for Inhibition of Germination-Associated Adhesion of Botrytis cinerea and Puccinia recondita Spores

PubMed Central

Slawecki, Richard A.; Ryan, Eileen P.; Young, David H.

2002-01-01

Botrytis cinerea and Puccinia recondita spores adhere strongly to polystyrene microtiter plates coincident with germination. We developed assays for inhibition of spore adhesion in 96-well microtiter plates by using sulforhodamine B staining to quantify the adherent spores. In both organisms, fungicides that inhibited germination strongly inhibited spore adhesion, with 50% effective concentrations (EC50s) comparable to those for inhibition of germination. In contrast, fungicides that acted after germination in B. cinerea inhibited spore adhesion to microtiter plates only at concentrations much higher than their EC50s for inhibition of mycelial growth. Similarly, in P. recondita the ergosterol biosynthesis inhibitors myclobutanil and fenbuconazole acted after germination and did not inhibit spore adhesion. The assays provide a rapid, high-throughput alternative to traditional spore germination assays and may be applicable to other fungi. PMID:11823196

Ontogenetic characterization of sporangium and spore of Huperzia serrata: an anti-aging disease fern.

PubMed

Long, Hua; Li, Jing; Li, You-You; Xie, De-Yu; Peng, Qing-Zhong; Li, Li

2016-12-01

Huperzia serrata is a medicinal plant used in Traditional Chinese Medicine, which has been used to prevent against aging diseases. It is mainly propagated by spores and grows extremely slowly. Due to severe harvest, it is a highly endangered species. In this report, we characterize ontogenesis of sporangia and spores that are associated with propagation. A wild population of H. serrata plants is localized in western Hunan province, China and protected by Chinese Government to study its development (e.g. sporangia and spores) and ecology. Both field and microscopic observations were conducted for a few of years. The development of sporangia from their initiation to maturation took nearly 1 year. Microscopic observations showed that the sporangial walls were developed from epidermal cells via initiation, cell division, and maturation. The structure of the mature sporangial wall is composed of one layer of epidermis, two middle layers of cells, and one layer of tapetum. Therefore, the sporangium is the eusporangium type. Spore development is characterized into six stages, initiation from epidermal cell and formation of sporogenous cells, primary sporogenous cell, secondary sporogenous cell, spore mother cell, tetrad, and maturation. The sporangial development of H. serrata belongs to the eusporangium type. The development takes approximately 1 year period from the initiation to the maturation. These data are useful for improving propagation of this medicinal plant in the future.

Venturing in coral larval chimerism: a compact functional domain with fostered genotypic diversity

NASA Astrophysics Data System (ADS)

Rinkevich, Baruch; Shaish, Lee; Douek, Jacob; Ben-Shlomo, Rachel

2016-01-01

The globally distributed coral species Pocillopora damicornis is known to release either sexual or asexual derived planula-larvae in various reef locations. Using microsatellite loci as markers, we documented the release of asexually derived chimeric larvae (CL), originating from mosaicked maternal colonies that were also chimeras, at Thai and Philippines reefs. The CL, each presenting different combinations of maternal genotypic constituents, create genetically-complex sets of asexual propagules. This novel mode of inheritance in corals challenges classical postulations of sexual/asexual reproduction traits, as asexual derived CL represent an alliance between genotypes that significantly sways the recruits’ absolute fitness. This type of inherited chimerism, while enhancing intra-entity genetic heterogeneity, is an evolutionary tactic used to increase genetic-heterogeneity, primarily in new areas colonized by a limited number of larvae. Chimerism may also facilitate combat global change impacts by exhibiting adjustable genomic combinations of within-chimera traits that could withstand alterable environmental pressures, helping Pocillopora become a successful cosmopolitan species.

Development of size-selective sampling of Bacillus anthracis surrogate spores from simulated building air intake mixtures for analysis via laser-induced breakdown spectroscopy.

PubMed

Gibb-Snyder, Emily; Gullett, Brian; Ryan, Shawn; Oudejans, Lukas; Touati, Abderrahmane

2006-08-01

Size-selective sampling of Bacillus anthracis surrogate spores from realistic, common aerosol mixtures was developed for analysis by laser-induced breakdown spectroscopy (LIBS). A two-stage impactor was found to be the preferential sampling technique for LIBS analysis because it was able to concentrate the spores in the mixtures while decreasing the collection of potentially interfering aerosols. Three common spore/aerosol scenarios were evaluated, diesel truck exhaust (to simulate a truck running outside of a building air intake), urban outdoor aerosol (to simulate common building air), and finally a protein aerosol (to simulate either an agent mixture (ricin/anthrax) or a contaminated anthrax sample). Two statistical methods, linear correlation and principal component analysis, were assessed for differentiation of surrogate spore spectra from other common aerosols. Criteria for determining percentages of false positives and false negatives via correlation analysis were evaluated. A single laser shot analysis of approximately 4 percent of the spores in a mixture of 0.75 m(3) urban outdoor air doped with approximately 1.1 x 10(5) spores resulted in a 0.04 proportion of false negatives. For that same sample volume of urban air without spores, the proportion of false positives was 0.08.

Detecting invisible bacillus spores on surfaces using a portable surface-enhanced Raman analyzer

NASA Astrophysics Data System (ADS)

Farquharson, Stuart; Inscore, Frank; Sperry, Jay F.

2006-10-01

Since the distribution of anthrax causing spores through the U.S. Postal System in the autumn of 2001, numerous methods have been developed to detect spores with the goal of minimizing casualties. During and following an attack it is also important to detect spores on surfaces, to assess extent of an attack, to quantify risk of infection by contact, as well as to evaluate post-attack clean-up. To perform useful measurements, analyzers and/or methods must be capable of detecting as few as 10 spores/cm2, in under 5-minutes, with little or no sample preparation or false-positive responses, using a portable device. In an effort to develop such a device, we have been investigating the ability of surfaceenhanced Raman spectroscopy (SERS) to detect dipicolinic acid (DPA) as a chemical signature of bacilli spores. In 2003 we employed SERS to measure DPA extracted from a 10,000 spores per μL sample using hot dodecylamine. Although the entire measurement was performed in 2 minutes, the need to heat the dodecylamine limits field portability of the method. Here we describe the use of a room temperature digesting agent in combination with SERS to detect 220 spores collected from a surface in a 1 μL sample within 3 minutes.

The roles of sexual and asexual reproduction in the origin and dissemination of strains causing fungal infectious disease outbreaks.

PubMed

Ashu, Eta Ebasi; Xu, Jianping

2015-12-01

Sexual reproduction commonly refers to the reproductive process in which genomes from two sources are combined into a single cell through mating and then the zygote genomes are partitioned to progeny cells through meiosis. Reproduction in the absence of mating and meiosis is referred to as asexual or clonal reproduction. One major advantage of sexual reproduction is that it generates genetic variation among progeny which may allow for faster adaptation of the population to novel and/or stressful environments. However, adaptation to stressful or new environments can still occur through mutation, in the absence of sex. In this review, we analyzed the relative contributions of sexual and asexual reproduction in the origin and spread of strains causing fungal infectious diseases outbreaks. The necessity of sex and the ability of asexual fungi to initiate outbreaks are discussed. We propose a framework that relates the modes of reproduction to the origin and propagation of fungal disease outbreaks. Our analyses suggest that both sexual and asexual reproduction can play critical roles in the origin of outbreak strains and that the rapid spread of outbreak strains is often accomplished through asexual expansion. Copyright © 2015 Elsevier B.V. All rights reserved.

Mutator dynamics in sexual and asexual experimental populations of yeast.

PubMed

Raynes, Yevgeniy; Gazzara, Matthew R; Sniegowski, Paul D

2011-06-07

In asexual populations, mutators may be expected to hitchhike with associated beneficial mutations. In sexual populations, recombination is predicted to erode such associations, inhibiting mutator hitchhiking. To investigate the effect of recombination on mutators experimentally, we compared the frequency dynamics of a mutator allele (msh2Δ) in sexual and asexual populations of Saccharomyces cerevisiae. Mutator strains increased in frequency at the expense of wild-type strains in all asexual diploid populations, with some approaching fixation in 150 generations of propagation. Over the same period of time, mutators declined toward loss in all corresponding sexual diploid populations as well as in haploid populations propagated asexually. We report the first experimental investigation of mutator dynamics in sexual populations. We show that a strong mutator quickly declines in sexual populations while hitchhiking to high frequency in asexual diploid populations, as predicted by theory. We also show that the msh2Δ mutator has a high and immediate realized cost that is alone sufficient to explain its decline in sexual populations. We postulate that this cost is indirect; namely, that it is due to a very high rate of recessive lethal or strongly deleterious mutation. However, we cannot rule out the possibility that msh2Δ also has unknown directly deleterious effects on fitness, and that these effects may differ between haploid asexual and sexual populations. Despite these reservations, our results prompt us to speculate that the short-term cost of highly deleterious recessive mutations can be as important as recombination in preventing mutator hitchhiking in sexual populations.

Heritable gene expression differences between apomictic clone members in Taraxacum officinale: Insights into early stages of evolutionary divergence in asexual plants.

PubMed

Ferreira de Carvalho, Julie; Oplaat, Carla; Pappas, Nikolaos; Derks, Martijn; de Ridder, Dick; Verhoeven, Koen J F

2016-03-08

Asexual reproduction has the potential to enhance deleterious mutation accumulation and to constrain adaptive evolution. One source of mutations that can be especially relevant in recent asexuals is activity of transposable elements (TEs), which may have experienced selection for high transposition rates in sexual ancestor populations. Predictions of genomic divergence under asexual reproduction therefore likely include a large contribution of transposable elements but limited adaptive divergence. For plants empirical insight into genome divergence under asexual reproduction remains limited. Here, we characterize expression divergence between clone members of a single apomictic lineage of the common dandelion (Taraxacum officinale) to contribute to our knowledge of genome evolution under asexuality. Using RNA-Seq, we show that about one third of heritable divergence within the apomictic lineage is driven by TEs and TE-related gene activity. In addition, we identify non-random transcriptional differences in pathways related to acyl-lipid and abscisic acid metabolisms which might reflect functional divergence within the apomictic lineage. We analyze SNPs in the transcriptome to assess genetic divergence between the apomictic clone members and reveal that heritable expression differences between the accessions are not explained simply by genome-wide genetic divergence. The present study depicts a first effort towards a more complete understanding of apomictic plant genome evolution. We identify abundant TE activity and ecologically relevant functional genes and pathways affecting heritable within-lineage expression divergence. These findings offer valuable resources for future work looking at epigenetic silencing and Cis-regulation of gene expression with particular emphasis on the effects of TE activity on asexual species' genome.

Hybridization and polyploidy enable genomic plasticity without sex in the most devastating plant-parasitic nematodes

PubMed Central

Perfus-Barbeoch, Laetitia; Da Rocha, Martine; Sallet, Erika; Bailly-Bechet, Marc; Castagnone-Sereno, Philippe; Flot, Jean-François; Kozlowski, Djampa K.; Cazareth, Julie; Couloux, Arnaud; Da Silva, Corinne; Guy, Julie; Kim-Jo, Yu-Jin; Rancurel, Corinne; Abad, Pierre; Wincker, Patrick

2017-01-01

Root-knot nematodes (genus Meloidogyne) exhibit a diversity of reproductive modes ranging from obligatory sexual to fully asexual reproduction. Intriguingly, the most widespread and devastating species to global agriculture are those that reproduce asexually, without meiosis. To disentangle this surprising parasitic success despite the absence of sex and genetic exchanges, we have sequenced and assembled the genomes of three obligatory ameiotic and asexual Meloidogyne. We have compared them to those of relatives able to perform meiosis and sexual reproduction. We show that the genomes of ameiotic asexual Meloidogyne are large, polyploid and made of duplicated regions with a high within-species average nucleotide divergence of ~8%. Phylogenomic analysis of the genes present in these duplicated regions suggests that they originated from multiple hybridization events and are thus homoeologs. We found that up to 22% of homoeologous gene pairs were under positive selection and these genes covered a wide spectrum of predicted functional categories. To biologically assess functional divergence, we compared expression patterns of homoeologous gene pairs across developmental life stages using an RNAseq approach in the most economically important asexually-reproducing nematode. We showed that >60% of homoeologous gene pairs display diverged expression patterns. These results suggest a substantial functional impact of the genome structure. Contrasting with high within-species nuclear genome divergence, mitochondrial genome divergence between the three ameiotic asexuals was very low, signifying that these putative hybrids share a recent common maternal ancestor. Transposable elements (TE) cover a ~1.7 times higher proportion of the genomes of the ameiotic asexual Meloidogyne compared to the sexual relative and might also participate in their plasticity. The intriguing parasitic success of asexually-reproducing Meloidogyne species could be partly explained by their TE-rich composite genomes, resulting from allopolyploidization events, and promoting plasticity and functional divergence between gene copies in the absence of sex and meiosis. PMID:28594822

Does the silver moss Bryum argenteum exhibit sex-specific patterns in vegetative growth rate, asexual fitness or prezygotic reproductive investment?

PubMed Central

Horsley, Kimberly; Stark, Lloyd R.; McLetchie, D. Nicholas

2011-01-01

Background and Aims Expected life history trade-offs associated with sex differences in reproductive investment are often undetected in seed plants, with the difficulty arising from logistical issues of conducting controlled experiments. By controlling genotype, age and resource status of individuals, a bryophyte was assessed for sex-specific and location-specific patterns of vegetative, asexual and sexual growth/reproduction across a regional scale. Methods Twelve genotypes (six male, six female) of the dioecious bryophyte Bryum argenteum were subcultured to remove environmental effects, regenerated asexually to replicate each genotype 16 times, and grown over a period of 92 d. Plants were assessed for growth rates, asexual and sexual reproductive traits, and allocation to above- and below-ground regenerative biomass. Key Results The degree of sexual versus asexual reproductive investment appears to be under genetic control, with three distinct ecotypes found in this study. Protonemal growth rate was positively correlated with asexual reproduction and sexual reproduction, whereas asexual reproduction was negatively correlated (appeared to trade-off) with vegetative growth (shoot production). No sex-specific trade-offs were detected. Female sex-expressing shoots were longer than males, but the sexes did not differ in growth traits, asexual traits, sexual induction times, or above- and below-ground biomass. Males, however, had much higher rates of inflorescence production than females, which translated into a significantly higher (24x) prezygotic investment for males relative to females. Conclusions Evidence for three distinct ecotypes is presented for a bryophyte based on regeneration traits. Prior to zygote production, the sexes of this bryophyte did not differ in vegetative growth traits but significantly differed in reproductive investment, with the latter differences potentially implicated in the strongly biased female sex ratio. The disparity between males and females for prezygotic reproductive investment is the highest known for bryophytes. PMID:21320878

Sensitizing Clostridium difficile Spores with Germinants on Skin and Environmental Surfaces Represents a New Strategy for Reducing Spores via Ambient Mechanisms

PubMed Central

Nerandzic, Michelle M.; Donskey, Curtis J.

2017-01-01

Background Clostridium difficile is a leading cause of healthcare-associated infections worldwide. Prevention of C. difficile transmission is challenging because spores are not killed by alcohol-based hand sanitizers or many commonly used disinfectants. One strategy to control spores is to induce germination, thereby rendering the spores more susceptible to benign disinfection measures and ambient stressors. Methods/Results C. difficile spores germinated on skin after a single application of cholic acid-class bile salts and co-germinants; for 4 C. difficile strains, recovery of viable spores from skin was reduced by ~0.3 log10CFU to 2 log10CFU after 2 hours and ~1 log10CFU to > 2.5 log10CFU after 24 hours. The addition of taurocholic acid to 70% and 30% ethanol significantly enhanced reduction of viable spores on skin and on surfaces. Desiccation, and to a lesser extent the presence of oxygen, were identified as the stressors responsible for reductions of germinated spores on skin and surfaces. Additionally, germinated spores became susceptible to killing by pH 1.5 hydrochloric acid, suggesting that germinated spores that remain viable on skin and surfaces might be killed by gastric acid after ingestion. Antibiotic-treated mice did not become colonized after exposure to germinated spores, whereas 100% of mice became colonized after exposure to the same quantity of dormant spores. Conclusions Germination could provide a new approach to reduce C. difficile spores on skin and in the environment and to render surviving spores less capable of causing infection. Our findings suggest that it may be feasible to develop alcohol-based hand sanitizers containing germinants that reduce spores on hands. PMID:29167835

Fighting Ebola with novel spore decontamination technologies for the military.

PubMed

Doona, Christopher J; Feeherry, Florence E; Kustin, Kenneth; Olinger, Gene G; Setlow, Peter; Malkin, Alexander J; Leighton, Terrance

2015-01-01

Recently, global public health organizations such as Doctors without Borders (MSF), the World Health Organization (WHO), Public Health Canada, National Institutes of Health (NIH), and the U.S. government developed and deployed Field Decontamination Kits (FDKs), a novel, lightweight, compact, reusable decontamination technology to sterilize Ebola-contaminated medical devices at remote clinical sites lacking infra-structure in crisis-stricken regions of West Africa (medical waste materials are placed in bags and burned). The basis for effectuating sterilization with FDKs is chlorine dioxide (ClO2) produced from a patented invention developed by researchers at the US Army Natick Soldier RD&E Center (NSRDEC) and commercialized as a dry mixed-chemical for bacterial spore decontamination. In fact, the NSRDEC research scientists developed an ensemble of ClO2 technologies designed for different applications in decontaminating fresh produce; food contact and handling surfaces; personal protective equipment; textiles used in clothing, uniforms, tents, and shelters; graywater recycling; airplanes; surgical instruments; and hard surfaces in latrines, laundries, and deployable medical facilities. These examples demonstrate the far-reaching impact, adaptability, and versatility of these innovative technologies. We present herein the unique attributes of NSRDEC's novel decontamination technologies and a Case Study of the development of FDKs that were deployed in West Africa by international public health organizations to sterilize Ebola-contaminated medical equipment. FDKs use bacterial spores as indicators of sterility. We review the properties and structures of spores and the mechanisms of bacterial spore inactivation by ClO2. We also review mechanisms of bacterial spore inactivation by novel, emerging, and established non-thermal technologies for food preservation, such as high pressure processing, irradiation, cold plasma, and chemical sanitizers, using an array of Bacillus subtilis mutants to probe mechanisms of spore germination and inactivation. We employ techniques of high-resolution atomic force microscopy and phase contrast microscopy to examine the effects of γ-irradiation on bacterial spores of Bacillus anthracis, Bacillus thuringiensis, and Bacillus atrophaeus spp. and of ClO2 on B. subtilis spores, and present in detail assays using spore bio-indicators to ensure sterility when decontaminating with ClO2.

Fighting Ebola with novel spore decontamination technologies for the military

PubMed Central

Doona, Christopher J.; Feeherry, Florence E.; Kustin, Kenneth; Olinger, Gene G.; Setlow, Peter; Malkin, Alexander J.; Leighton, Terrance

2015-01-01

Recently, global public health organizations such as Doctors without Borders (MSF), the World Health Organization (WHO), Public Health Canada, National Institutes of Health (NIH), and the U.S. government developed and deployed Field Decontamination Kits (FDKs), a novel, lightweight, compact, reusable decontamination technology to sterilize Ebola-contaminated medical devices at remote clinical sites lacking infra-structure in crisis-stricken regions of West Africa (medical waste materials are placed in bags and burned). The basis for effectuating sterilization with FDKs is chlorine dioxide (ClO2) produced from a patented invention developed by researchers at the US Army Natick Soldier RD&E Center (NSRDEC) and commercialized as a dry mixed-chemical for bacterial spore decontamination. In fact, the NSRDEC research scientists developed an ensemble of ClO2 technologies designed for different applications in decontaminating fresh produce; food contact and handling surfaces; personal protective equipment; textiles used in clothing, uniforms, tents, and shelters; graywater recycling; airplanes; surgical instruments; and hard surfaces in latrines, laundries, and deployable medical facilities. These examples demonstrate the far-reaching impact, adaptability, and versatility of these innovative technologies. We present herein the unique attributes of NSRDEC’s novel decontamination technologies and a Case Study of the development of FDKs that were deployed in West Africa by international public health organizations to sterilize Ebola-contaminated medical equipment. FDKs use bacterial spores as indicators of sterility. We review the properties and structures of spores and the mechanisms of bacterial spore inactivation by ClO2. We also review mechanisms of bacterial spore inactivation by novel, emerging, and established non-thermal technologies for food preservation, such as high pressure processing, irradiation, cold plasma, and chemical sanitizers, using an array of Bacillus subtilis mutants to probe mechanisms of spore germination and inactivation. We employ techniques of high-resolution atomic force microscopy and phase contrast microscopy to examine the effects of γ-irradiation on bacterial spores of Bacillus anthracis, Bacillus thuringiensis, and Bacillus atrophaeus spp. and of ClO2 on B. subtilis spores, and present in detail assays using spore bio-indicators to ensure sterility when decontaminating with ClO2. PMID:26322021

Challenges of assessing the clinical efficacy of asexual blood-stage Plasmodium falciparum malaria vaccines.

PubMed

Sheehy, Susanne H; Douglas, Alexander D; Draper, Simon J

2013-09-01

In the absence of any highly effective vaccine candidate against Plasmodium falciparum malaria, it remains imperative for the field to pursue all avenues that may lead to the successful development of such a formulation. The development of a subunit vaccine targeting the asexual blood-stage of Plasmodium falciparum malaria infection has proven particularly challenging with only limited success to date in clinical trials. However, only a fraction of potential blood-stage vaccine antigens have been evaluated as targets, and a number of new promising candidate antigen formulations and delivery platforms are approaching clinical development. It is therefore essential that reliable and sensitive methods of detecting, or ruling out, even modest efficacy of blood-stage vaccines in small clinical trials be established. In this article we evaluate the challenges facing blood-stage vaccine developers, assess the appropriateness and limitations of various in vivo approaches for efficacy assessment and suggest future directions for the field.

Production of ochratoxin A by Aspergillus ochraceus NRRL-3174 before and after exposures to 60Co irradiation.

PubMed Central

Applegate, K L; Chipley, J R

1976-01-01

Spores from the toxigenic organism Aspergillus ochraceus NRRL-3174 were exposed to specific levels of gamma irradiation and then allowed to germinate on selected media. Increases in ochratoxin A production by irradiated, compared to non-irradiated, spores were observed after inoculation of spores onto a cracked red wheat or into a synthetic liquid medium. Variations in daily ochratoxin production were also observed for control and irradiated spore-derived cultures developing on both media, with maximum toxin production varying from 7 to 11 days of incubation. The most notable increases in ochratoxin A production occurred from cultures developing from spores having been irradiated with 10, 25, or 50 krad. Exposures to 400 or 600 krad resulted in complete inhibition of spore germination and, consequently, no ochratoxin production. Of the two substrates used, wheat and synthetic, the quantities of ochratoxin A produced were significantly lower in the synthetic media than on the natural substrate. Higher and more rapid toxin production occurred from spores having been irradiated with 10, 25, 50, and 100 krad than occurred from the non-irradiated control spores when grown on synthetic media. Cultures derived from spores having been exposed to 10, 25, 50, and 100 krad produced significantly higher levels of ochratoxin A after 8 days of incubation on natural substrate than did the controls. Analysis of variance revealed that substrate, length of incubation, as well as irradiation levels all affected the time required to produce maximum levels of ochratoxin A. PMID:938031

Proteomic profile of dormant Trichophyton Rubrum conidia

PubMed Central

Leng, Wenchuan; Liu, Tao; Li, Rui; Yang, Jian; Wei, Candong; Zhang, Wenliang; Jin, Qi

2008-01-01

Background Trichophyton rubrum is the most common dermatophyte causing fungal skin infections in humans. Asexual sporulation is an important means of propagation for T. rubrum, and conidia produced by this way are thought to be the primary cause of human infections. Despite their importance in pathogenesis, the conidia of T. rubrum remain understudied. We intend to intensively investigate the proteome of dormant T. rubrum conidia to characterize its molecular and cellular features and to enhance the development of novel therapeutic strategies. Results The proteome of T. rubrum conidia was analyzed by combining shotgun proteomics with sample prefractionation and multiple enzyme digestion. In total, 1026 proteins were identified. All identified proteins were compared to those in the NCBI non-redundant protein database, the eukaryotic orthologous groups database, and the gene ontology database to obtain functional annotation information. Functional classification revealed that the identified proteins covered nearly all major biological processes. Some proteins were spore specific and related to the survival and dispersal of T. rubrum conidia, and many proteins were important to conidial germination and response to environmental conditions. Conclusion Our results suggest that the proteome of T. rubrum conidia is considerably complex, and that the maintenance of conidial dormancy is an intricate and elaborate process. This data set provides the first global framework for the dormant T. rubrum conidia proteome and is a stepping stone on the way to further study of the molecular mechanisms of T. rubrum conidial germination and the maintenance of conidial dormancy. PMID:18578874

[Sporogenesis, sporoderm and mature spore ornamentation in Lycopodiaceae].

PubMed

Rincon Baron, Edgar Javier; Rolleri, Cristina Hilda; Passarelli, Lilian M; Espinosa Matías, Silvia; Torres, Alba Marina

2014-09-01

Studies on reproductive aspects, spore morphology and ultrastructure of Lycopodiaceae are not very common in the scientific literature, and constitute essential information to support taxonomic and systematic relationships among the group. In order to complete existing information, adding new and broader contributions on these topics, a comparative analysis of the sporogenesis ultrastructure, with emphasis on cytological aspects of the sporocyte coat development, tapetum, monoplastidic and polyplastidic meiosis, sporoderm ontogeny and ornamentation of the mature spores, was carried out in 43 taxa of eight genera of the Lycopodiaceae: Austrolycopodium, Diphasium, Diphasiastrum, Huperzia (including Phlegmariurus), Lycopodium, Lycopodiella, Palhinhaea and Pseudolycopodiella growing in the Andes of Colombia and the Neotropics. For this study, the transmission elec- tron microscopy (TEM) samples were collected in Cauca and Valle del Cauca Departments, while most of the spores for scanning electron microscopy (SEM) analysis were obtained from herbarium samples. We followed standard preparation procedures for spore observation by TEM and SEM. Results showed that the sporocyte coat is largely composed by primary wall components; the sporocyte develop much of their metabolic activity in the production of their coat, which is retained until the spores release; protective functions for the diploid cells undergoing meiosis is postulated here for this layer. The abundance of dictyosomes in the sporocyte cytoplasm was related to the formation and development of the sporocyte coat. Besides microtubule activity, the membrane of sporocyte folds, associated with electrodense material, and would early determine the final patterns of spore ornamentation. Monoplastidic condition is common in Lycopodium s.l., whereas polyplastidic condition was observed in species of Huperzia and Lycopodiella s. l. In monoplastidic species, the tapetum presents abun- dant multivesicular bodies, while in polyplastidic species, the secretory activity of the tapetum is less intense. Sporoderm development is centripetal, exospore is the first formed layer, then the endospore and, if present, perispore is the final deposited layer. Adult spores of the Lycopodiaceae showed two patterns of ornamentation: negative or caviform (foveolate spores) and positive or muriform ornamentation, the latter with two subtypes (rugate and reticulate spores). The spores of Huperzia are characteristically foveolate, the rugate spores were found in a few species of Huperzia and in all of the Lycopodiella s. l. taxa studied, while Lycopodium s.l. spores bear reticulate ornamentation. Numerous ornamentation traits are diagnostic at the specific level. The types of ornamentation found do not support the recent extreme fragmentation of the family in several genera, but could match, a priori, with the idea of three subfamilies. The findings of sporogenesis, extremely similar in all taxa studied, point more to consider fewer genera, more comprehensive, than the recent, marked splitting of the family.

Convergent acquisition of non-embryonic development in styelid ascidians.

PubMed

Alié, Alexandre; Hiebert, Laurel Sky; Simion, Paul; Scelzo, Marta; Prünster, Maria Mandela; Lotito, Sonia; Delsuc, Frédéric; Douzery, Emmanuel J P; Dantec, Christelle; Lemaire, Patrick; Darras, Sébastien; Kawamura, Kazuo; Brown, Federico D; Tiozzo, Stefano

2018-04-11

Asexual propagation and whole body regeneration are forms of non-embryonic development (NED) widespread across animal phyla and central in life history and evolutionary diversification of metazoans. Whereas it is challenging to reconstruct the gains or losses of NED at large phylogenetic scale, comparative studies could benefit from being conducted at more restricted taxonomic scale, in groups for which phylogenetic relationships are well established. The ascidian family of Styelidae encompasses strictly sexually reproducing solitary forms as well as colonial species that combine sexual reproduction with different forms of NED. To date, the phylogenetic relationships between colonial and solitary styelids remain controversial and so is the pattern of NED evolution. In this study, we built an original pipeline to combine 8 genomes with 18 de novo assembled transcriptomes and constructed datasets of unambiguously orthologous genes. Using a phylogenomic super-matrix of 4,908 genes from these 26 tunicates we provided a robust phylogeny of this family of chordates, which supports two convergent acquisitions of NED. This result prompted us to further describe the budding process in the species Polyandrocarpa zorritensis, leading to the discovery of a novel mechanism of asexual development. While the pipeline and the datasets produced can be used for further phylogenetic reconstructions in tunicates, the phylogeny provided here sets an evolutionary framework for future experimental studies on the emergence and disappearance of complex characters such as asexual propagation and whole body regeneration.

Convergent Acquisition of Nonembryonic Development in Styelid Ascidians

PubMed Central

Alié, Alexandre; Hiebert, Laurel Sky; Simion, Paul; Scelzo, Marta; Prünster, Maria Mandela; Lotito, Sonia; Delsuc, Frédéric; Douzery, Emmanuel J P; Dantec, Christelle; Lemaire, Patrick; Darras, Sébastien; Kawamura, Kazuo; Brown, Federico D; Tiozzo, Stefano

2018-01-01

Abstract Asexual propagation and whole body regeneration are forms of nonembryonic development (NED) widespread across animal phyla and central in life history and evolutionary diversification of metazoans. Whereas it is challenging to reconstruct the gains or losses of NED at large phylogenetic scale, comparative studies could benefit from being conducted at more restricted taxonomic scale, in groups for which phylogenetic relationships are well established. The ascidian family of Styelidae encompasses strictly sexually reproducing solitary forms as well as colonial species that combine sexual reproduction with different forms of NED. To date, the phylogenetic relationships between colonial and solitary styelids remain controversial and so is the pattern of NED evolution. In this study, we built an original pipeline to combine eight genomes with 18 de novo assembled transcriptomes and constructed data sets of unambiguously orthologous genes. Using a phylogenomic super-matrix of 4,908 genes from these 26 tunicates we provided a robust phylogeny of this family of chordates, which supports two convergent acquisitions of NED. This result prompted us to further describe the budding process in the species Polyandrocarpa zorritensis, leading to the discovery of a novel mechanism of asexual development. Whereas the pipeline and the data sets produced can be used for further phylogenetic reconstructions in tunicates, the phylogeny provided here sets an evolutionary framework for future experimental studies on the emergence and disappearance of complex characters such as asexual propagation and whole body regeneration. PMID:29660002

DISSECTING COLONY DEVELOPMENT OF NEUROSPORA CRASSA USING mRNA PROFILING AND COMPARTATIVE GENOMICS APPROACHES

USDA-ARS?s Scientific Manuscript database

Colony development, which includes hyphal extension, branching, anastomosis and asexual sporulation are fundamental aspects of the lifecycle of filamentous fungi; genetic mechanisms underlying these phenomena are poorly understood. We conducted transcriptional profiling during colony development of...

Gα proteins Gvm2 and Gvm3 regulate vegetative growth, asexual development, and pathogenicityon apple in Valsa mali.

PubMed

Song, Na; Dai, Qingqing; Zhu, Baitao; Wu, Yuxing; Xu, Ming; Voegele, Ralf Thomas; Gao, Xiaoning; Kang, Zhensheng; Huang, Lili

2017-01-01

In fungi, heterotrimeric guanine-nucleotide binding proteins (G-proteins) are key elements of signal transduction pathways, which control growth, asexual and sexual development, as well as virulence. In this study, we have identified two genes encoding heterotrimeric G protein alpha subunits, named Gvm2 and Gvm3, from Valsa mali, the causal agent of apple Valsa canker. Characterization of Gvm2 and Gvm3 mutants indicates that Gvm3 may be a crucial regulator of vegetative growth. Deletion of the corresponding gene results in a 20% reduction in growth rate. Besides, Gvm2 and Gvm3 seem to be involved in asexual reproduction, and mutants are hypersensitive to oxidative and cell membrane stresses. Interestingly, both G protein alpha subunits were most probably involved in V. mali virulence. In infection assays using Malus domestica cv. 'Fuji' leaves and twigs, the size of lesions caused by deletion mutants △Gvm2, or △Gvm3 are significantly reduced. Furthermore, many genes encoding hydrolytic enzymes-important virulence factors in V. mali-are expressed at a lower level in these deletion mutants. Our results suggest that Gvm2 and Gvm3 play an important role in virulence probably by regulation of expression of cell wall degrading enzymes. △Gvm2, and △Gvm3 mutants were further analyzed with respect to their impact on the transcript levels of genes in the cAMP/PKA pathway. The expression of the genes encoding adenylate cyclase VmAC, protein kinase A (PKA) regulatory subunit VmPKR, and PKA catalytic subunit VmPKA1 are down-regulated in both mutants. Further analyses indicated that intracellular cAMP level and PKA activity are down-regulated in the △Gvm3 mutant, but are basically unchanged in the △Gvm2 mutant. Overall, our findings indicate that both Gvm2 and Gvm3 play diverse roles in the modulation of vegetative growth, asexual development, and virulence in V. mali.

Ultrastructure of developing ascospores in Sordaria brevicollis.

PubMed

Hackett, C J; Chen, K C

1976-05-01

The ultrastructure of ascospore wall formation in the pyrenomycete Sordaria brevicollis was studied in developing asci at progressive time intervals. From early spore delimitation through final stage of maturation, the wall of the ascospore differentiated into four composite layers, the periascosporium the delineation ascosporium, the subascosproium, and the endoascosproium, While ascospores were at the hyaline stage of development,they possessed only the periascosporium and delineation ascosporium as their wall components. At about 7 to 8 days from the initiation of the cross, the spores developed a yellow color, and this coloration was always associated with the elaboration of the subascorsporium just internal to the ascosporium. Asthe spores continued to progressively darken in color, the subascosporium was seen to increase in complexity, electron density, and thickness. Soon after the formation of the subascosporium, the endoascosporium began to develop de novo and was, therefore, the last wall layer formed as the spore approached maturity.

Development of actinomycetes in brown semidesert soil under low water pressure

NASA Astrophysics Data System (ADS)

Zvyagintsev, D. G.; Zenova, G. M.; Sudnitsyn, I. I.; Gracheva, T. A.; Lapygina, E. E.; Napol'skaya, K. R.; Sydnitsyna, A. E.

2012-07-01

Under laboratory conditions, the spores of a xerotolerant Streptomyces odorifera strain germinated in brown semidesert soil even at extremely low soil water pressure ( P = -96.4 MPa, -964 atm, a w 0.50); the plantlets increased in length and formed mycelium, on which a new generation of spores was produced (a complete development cycle of the actinomycetes—from a spore to the formation of new spores—passed). The duration of the first cycles of the actinomycetes' development varied from 13 days at P = -27 atm to 57 days at P = -964 atm and was directly proportional to the absolute value of the soil water pressure ( P). In the first cycles of the actinomycetes' development, the rate of increase of the concentration of the germinated spores and mycelium, as well as the logarithms of the mycelium-to-germinated spore concentration ratios, was inversely proportional to the logarithm of P. These relationships indicated that the energy state of the water determined its availability to soil biota and, hence, the activity of its physiological and biochemical processes.

Recent progress in Bacillus subtilis spore-surface display: concept, progress, and future.

PubMed

Wang, He; Wang, Yunxiang; Yang, Ruijin

2017-02-01

With the increased knowledge on spore structure and advances in biotechnology engineering, the newly developed spore-surface display system confers several inherent advantages over other microbial cell-surface display systems including enhanced stability and high safety. Bacillus subtilis is the most commonly used Bacillus species for spore-surface display. The expression of heterologous antigen or protein on the surface of B. subtilis spores has now been practiced for over a decade with noteworthy success. As an update and supplement to other previous reviews, we comprehensively summarize recent studies in the B. subtilis spore-surface display technique. We focus on its benefits as well as the critical factors affecting its display efficiency and offer suggestions for the future success of this field.

Quantification of Spore-forming Bacteria Carried by Dust Particles

NASA Technical Reports Server (NTRS)

Lin, Ying; Cholakian, Tanya; Gao, Wenming; Osman, Shariff; Barengoltz, Jack

2006-01-01

In order to establish a biological contamination transport model for predicting the cross contamination risk during spacecraft assembly and upon landing on Mars, it is important to understand the relationship between spore-forming bacteria and their carrier particles. We conducted air and surface sampling in indoor, outdoor, and cleanroom environments to determine the ratio of spore forming bacteria to their dust particle carriers of different sizes. The number of spore forming bacteria was determined from various size groups of particles in a given environment. Our data also confirms the existence of multiple spores on a single particle and spore clumps. This study will help in developing a better bio-contamination transport model, which in turn will help in determining forward contamination risks for future missions.

Cortex content of asporogenous mutants of Bacillus subtilis.

PubMed Central

Imae, Y; Strominger, J L

1976-01-01

A method for the measurement of muramic lactam, which is specifically located in the cortical peptidoglycan of bacterial spores, was developed as a quantitative assay method for spore cortex content. During sporulation of Bacillus subtilis 168, muramic lactam (i.e., spore cortex) began to appear at state IV of sporulation and continued to increase over most of the late stages of sporulation. Spore cortex contents of various spo mutants of B. subitils were surveyed. Cortex was not detected in mutants in which sporulation was blocked earlier than stage II sporulation. Spores of spo IV mutant had about 40% of the cortex content of the wild-type spores. One spo III mutant had a low amount of cortex, but four others had none. PMID:1262319

Laser induced disruption of bacterial spores on a microchip.

PubMed

Hofmann, Oliver; Murray, Kirk; Wilkinson, Alan-Shaun; Cox, Timothy; Manz, Andreas

2005-04-01

We report on the development of a laser based spore disruption method. Bacillus globigii spores were mixed with a laser light absorbing matrix and co-crystallized into 200-microm-wide and 20-microm-deep nanovials formed in a polydimethylsiloxane (PDMS) target plate. Surface tension effects were exploited to effect up to 125-fold spore enrichment. When the target zones were illuminated at atmospheric pressure with pulsed UV-laser light at fluences below 20 mJ cm(-2) a change in spore morphology was observed within seconds. Post illumination PCR analysis suggests the release of endogenous DNA indicative of spore disruption. For laser fluences above 20 mJ cm(-2), desorption of spores and fragments was also observed even without a matrix being employed. Desorbed material was collected in a PDMS flowcell attached to the target plate during laser illumination. This opens up a route towards the direct extraction of released DNA in an integrated spore disruption-PCR amplification microchip device.

Nanoscale Structural and Mechanical Analysis of Bacillus anthracis Spores Inactivated with Rapid Dry Heating

PubMed Central

Felker, Daniel L.; Burggraf, Larry W.

2014-01-01

Effective killing of Bacillus anthracis spores is of paramount importance to antibioterrorism, food safety, environmental protection, and the medical device industry. Thus, a deeper understanding of the mechanisms of spore resistance and inactivation is highly desired for developing new strategies or improving the known methods for spore destruction. Previous studies have shown that spore inactivation mechanisms differ considerably depending upon the killing agents, such as heat (wet heat, dry heat), UV, ionizing radiation, and chemicals. It is believed that wet heat kills spores by inactivating critical enzymes, while dry heat kills spores by damaging their DNA. Many studies have focused on the biochemical aspects of spore inactivation by dry heat; few have investigated structural damages and changes in spore mechanical properties. In this study, we have inactivated Bacillus anthracis spores with rapid dry heating and performed nanoscale topographical and mechanical analysis of inactivated spores using atomic force microscopy (AFM). Our results revealed significant changes in spore morphology and nanomechanical properties after heat inactivation. In addition, we also found that these changes were different under different heating conditions that produced similar inactivation probabilities (high temperature for short exposure time versus low temperature for long exposure time). We attributed the differences to the differential thermal and mechanical stresses in the spore. The buildup of internal thermal and mechanical stresses may become prominent only in ultrafast, high-temperature heat inactivation when the experimental timescale is too short for heat-generated vapor to efficiently escape from the spore. Our results thus provide direct, visual evidences of the importance of thermal stresses and heat and mass transfer to spore inactivation by very rapid dry heating. PMID:24375142

Biotechnology

NASA Image and Video Library

2003-01-22

Anthrax spores are inactive forms of Bacillus anthracis. They can survive for decades inside a spore's tough protective coating; they become active when inhaled by humans. A result of NASA- and industry-sponsored research to develop small greenhouses for space research is the unique AiroCide TiO2 system that kills anthrax spores and other pathogens.

Plant Development & the Fern Life Cycle: Using "Ceratopteris richardii."

ERIC Educational Resources Information Center

Renzaglia, Karen S.; And Others

1995-01-01

Presents laboratory activities focusing on the development of sexually mature gametophytes from single-celled spores. Includes techniques for culture and manipulation of gametophyte development from spores that are applicable for hands-on activities for students at all levels. (MKR)

The Conserved Spore Coat Protein SpoVM Is Largely Dispensable in Clostridium difficile Spore Formation.

PubMed

Ribis, John W; Ravichandran, Priyanka; Putnam, Emily E; Pishdadian, Keyan; Shen, Aimee

2017-01-01

The spore-forming bacterial pathogen Clostridium difficile is a leading cause of health care-associated infections in the United States. In order for this obligate anaerobe to transmit infection, it must form metabolically dormant spores prior to exiting the host. A key step during this process is the assembly of a protective, multilayered proteinaceous coat around the spore. Coat assembly depends on coat morphogenetic proteins recruiting distinct subsets of coat proteins to the developing spore. While 10 coat morphogenetic proteins have been identified in Bacillus subtilis , only two of these morphogenetic proteins have homologs in the Clostridia : SpoIVA and SpoVM. C. difficile SpoIVA is critical for proper coat assembly and functional spore formation, but the requirement for SpoVM during this process was unknown. Here, we show that SpoVM is largely dispensable for C. difficile spore formation, in contrast with B. subtilis . Loss of C. difficile SpoVM resulted in modest decreases (~3-fold) in heat- and chloroform-resistant spore formation, while morphological defects such as coat detachment from the forespore and abnormal cortex thickness were observed in ~30% of spoVM mutant cells. Biochemical analyses revealed that C. difficile SpoIVA and SpoVM directly interact, similarly to their B. subtilis counterparts. However, in contrast with B. subtilis , C. difficile SpoVM was not essential for SpoIVA to encase the forespore. Since C. difficile coat morphogenesis requires SpoIVA-interacting protein L (SipL), which is conserved exclusively in the Clostridia , but not the more broadly conserved SpoVM, our results reveal another key difference between C. difficile and B. subtilis spore assembly pathways. IMPORTANCE The spore-forming obligate anaerobe Clostridium difficile is the leading cause of antibiotic-associated diarrheal disease in the United States. When C. difficile spores are ingested by susceptible individuals, they germinate within the gut and transform into vegetative, toxin-secreting cells. During infection, C. difficile must also induce spore formation to survive exit from the host. Since spore formation is essential for transmission, understanding the basic mechanisms underlying sporulation in C. difficile could inform the development of therapeutic strategies targeting spores. In this study, we determine the requirement of the C. difficile homolog of SpoVM, a protein that is essential for spore formation in Bacillus subtilis due to its regulation of coat and cortex formation. We observed that SpoVM plays a minor role in C. difficile spore formation, in contrast with B. subtilis , indicating that this protein would not be a good target for inhibiting spore formation.

The Conserved Spore Coat Protein SpoVM Is Largely Dispensable in Clostridium difficile Spore Formation

PubMed Central

Ribis, John W.; Ravichandran, Priyanka; Putnam, Emily E.; Pishdadian, Keyan

2017-01-01

ABSTRACT The spore-forming bacterial pathogen Clostridium difficile is a leading cause of health care-associated infections in the United States. In order for this obligate anaerobe to transmit infection, it must form metabolically dormant spores prior to exiting the host. A key step during this process is the assembly of a protective, multilayered proteinaceous coat around the spore. Coat assembly depends on coat morphogenetic proteins recruiting distinct subsets of coat proteins to the developing spore. While 10 coat morphogenetic proteins have been identified in Bacillus subtilis, only two of these morphogenetic proteins have homologs in the Clostridia: SpoIVA and SpoVM. C. difficile SpoIVA is critical for proper coat assembly and functional spore formation, but the requirement for SpoVM during this process was unknown. Here, we show that SpoVM is largely dispensable for C. difficile spore formation, in contrast with B. subtilis. Loss of C. difficile SpoVM resulted in modest decreases (~3-fold) in heat- and chloroform-resistant spore formation, while morphological defects such as coat detachment from the forespore and abnormal cortex thickness were observed in ~30% of spoVM mutant cells. Biochemical analyses revealed that C. difficile SpoIVA and SpoVM directly interact, similarly to their B. subtilis counterparts. However, in contrast with B. subtilis, C. difficile SpoVM was not essential for SpoIVA to encase the forespore. Since C. difficile coat morphogenesis requires SpoIVA-interacting protein L (SipL), which is conserved exclusively in the Clostridia, but not the more broadly conserved SpoVM, our results reveal another key difference between C. difficile and B. subtilis spore assembly pathways. IMPORTANCE The spore-forming obligate anaerobe Clostridium difficile is the leading cause of antibiotic-associated diarrheal disease in the United States. When C. difficile spores are ingested by susceptible individuals, they germinate within the gut and transform into vegetative, toxin-secreting cells. During infection, C. difficile must also induce spore formation to survive exit from the host. Since spore formation is essential for transmission, understanding the basic mechanisms underlying sporulation in C. difficile could inform the development of therapeutic strategies targeting spores. In this study, we determine the requirement of the C. difficile homolog of SpoVM, a protein that is essential for spore formation in Bacillus subtilis due to its regulation of coat and cortex formation. We observed that SpoVM plays a minor role in C. difficile spore formation, in contrast with B. subtilis, indicating that this protein would not be a good target for inhibiting spore formation. PMID:28959733

Sexual recombination as a tool for engineering industrial Penicillium chrysogenum strains.

PubMed

Dahlmann, Tim A; Böhm, Julia; Becker, Kordula; Kück, Ulrich

2015-11-01

The recent discovery and functional characterization of opposite mating-type loci in the industrial penicillin producer Penicillium chrysogenum demonstrated their regulatory role in sexual as well as asexual development. Subsequent experiments further showed that a sexual life cycle can be induced in P. chrysogenum that was for long believed to reproduce exclusively by asexual propagation. Finally, crossing of wild type and production strains resulted in the generation of recombinant ascospore isolates. We predict from these recent findings that recombinant progeny for industrial applications can be obtained by sexual crossings and discuss experimental difficulties that occur when parental strains with karyotype heterogeneity are used for mating.

Human antibodies against spores of the genus Bacillus: a model study for detection of and protection against anthrax and the bioterrorist threat.

PubMed

Zhou, Bin; Wirsching, Peter; Janda, Kim D

2002-04-16

A naive, human single-chain Fv (scFv) phage-display library was used in bio-panning against live, native spores of Bacillus subtilis IFO 3336 suspended in solution. A direct in vitro panning and enzyme-linked immunosorbent assay-based selection afforded a panel of nine scFv-phage clones of which two, 5B and 7E, were chosen for further study. These two clones differed in their relative specificity and affinity for spores of B. subtilis IFO 3336 vs. a panel of spores from 11 other Bacillus species/strains. A variety of enzyme-linked immunosorbent assay protocols indicated these scFv-phage clones recognized different spore epitopes. Notably, some spore epitopes markedly changed between the free and microtiter-plate immobilized state as revealed by antibody-phage binding. An additional library selection procedure also was examined by constructing a Fab chain-shuffled sublibrary from the nine positive clones and by using a subtractive panning strategy to remove crossreactivity with B. licheniformis 5A24. The Fab-phage clone 52 was improved compared with 5B and was comparable to 7E in binding B. subtilis IFO 3336 vs. B. licheniformis 5A24, yet showed a distinctive crossreactivity pattern with other spores. We also developed a method to directly detect individual spores by using fluorescently labeled antibody-phage. Finally, a variety of "powders" that might be used in deploying spores of B. anthracis were examined for antibody-phage binding. The strategies described provide a foundation to discover human antibodies specific for native spores of B. anthracis that can be developed as diagnostic and therapeutic reagents.

New insights in the bacterial spore resistance to extreme terrestrial and extraterrestrial factors

NASA Astrophysics Data System (ADS)

Moeller, Ralf; Horneck, Gerda; Reitz, Guenther

Based on their unique resistance to various space parameters, Bacillus endospores are one of the model systems used for astrobiological studies. The extremely high resistance of bacterial endospores to environmental stress factors has intrigued researchers since long time and many characteristic spore features, especially those involved in the protection of spore DNA, have already been uncovered. The disclosure of the complete genomic sequence of Bacillus subtilis 168, one of the often used astrobiological model system, and the rapid development of tran-scriptional microarray techniques have opened new opportunities of gaining further insights in the enigma of spore resistance. Spores of B. subtilis were exposed to various extreme ter-restrial and extraterrestrial stressors to reach a better understanding of the DNA protection and repair strategies, which them to cope with the induced DNA damage. Following physical stress factors of environmental importance -either on Earth or in space -were selected for this thesis: (i) mono-and polychromatic UV radiation, (ii) ionizing radiation, (iii) exposure to ultrahigh vacuum; and (iv) high shock pressures simulating meteorite impacts. To reach a most comprehensive understanding of spore resistance to those harsh terrestrial or simulated extraterrestrial conditions, a standardized experimental protocol of the preparation and ana-lyzing methods was established including the determination of the following spore responses: (i) survival, (ii) induced mutations, (iii) DNA damage, (iv) role of different repair pathways by use of a set of repair deficient mutants, and (v) transcriptional responses during spore germi-nation by use of genome-wide transcriptome analyses and confirmation by RT-PCR. From this comprehensive set of data on spore resistance to a variety of environmental stress parameters a model of a "built-in" transcriptional program of bacterial spores in response to DNA damaging treatments to ensure DNA restoration during germination has been developed.

FvSTR1, a striatin orthologue in Fusarium virguliforme, is required for asexual development and virulence.

PubMed

Islam, Kazi T; Bond, Jason P; Fakhoury, Ahmad M

2017-08-01

The soil-borne fungus Fusarium virguliforme causes sudden death syndrome (SDS), one of the most devastating diseases of soybean in North and South America. Despite the importance of SDS, a clear understanding of the fungal pathogenicity factors that affect the development of this disease is still lacking. We have identified FvSTR1, a F. virguliforme gene, which encodes a protein similar to a family of striatin proteins previously reported to regulate signalling pathways, cell differentiation, conidiation, sexual development, and virulence in filamentous fungi. Striatins are multi-domain proteins that serve as scaffolding units in the striatin-interacting phosphatase and kinase (STRIPAK) complex in fungi and animals. To address the function of a striatin homologue in F. virguliforme, FvSTR1 was disrupted and functionally characterized using a gene knock out strategy. The resulting Fvstr1 mutants were largely impaired in conidiation and pigmentation, and displayed defective conidia and conidiophore morphology compared to the wild-type and ectopic transformants. Greenhouse virulence assays revealed that the disruption of FvSTR1 resulted in complete loss of virulence in F. virguliforme. Microtome studies using fluorescence microscopy showed that the Fvstr1 mutants were defective in their ability to colonize the vascular system. The Fvstr1 mutants also showed a reduced transcript level of genes involved in asexual reproduction and in the production of secondary metabolites. These results suggest that FvSTR1 has a critical role in asexual development and virulence in F. virguliforme.

Regulation of Conidiation by Light in Aspergillus nidulans

PubMed Central

Ruger-Herreros, Carmen; Rodríguez-Romero, Julio; Fernández-Barranco, Raul; Olmedo, María; Fischer, Reinhard; Corrochano, Luis M.; Canovas, David

2011-01-01

Light regulates several aspects of the biology of many organisms, including the balance between asexual and sexual development in some fungi. To understand how light regulates fungal development at the molecular level we have used Aspergillus nidulans as a model. We have performed a genome-wide expression analysis that has allowed us to identify >400 genes upregulated and >100 genes downregulated by light in developmentally competent mycelium. Among the upregulated genes were genes required for the regulation of asexual development, one of the major biological responses to light in A. nidulans, which is a pathway controlled by the master regulatory gene brlA. The expression of brlA, like conidiation, is induced by light. A detailed analysis of brlA light regulation revealed increased expression after short exposures with a maximum after 60 min of light followed by photoadaptation with longer light exposures. In addition to brlA, genes flbA–C and fluG are also light regulated, and flbA–C are required for the correct light-dependent regulation of the upstream regulator fluG. We have found that light induction of brlA required the photoreceptor complex composed of a phytochrome FphA, and the white-collar homologs LreA and LreB, and the fluffy genes flbA–C. We propose that the activation of regulatory genes by light is the key event in the activation of asexual development by light in A. nidulans. PMID:21624998

Impact of two treatments of a formulation of Beauveria bassiana (Deuteromycota: Hyphomycetes) conidia on Varroa mites (Acari: Varroidae) and on honeybee (Hymenoptera: Apidae) colony health.

PubMed

Meikle, William G; Mercadier, Guy; Holst, Niels; Girod, Vincent

2008-12-01

Bee colonies in southern France were treated with conidia (asexual spores) from two strains of Beauveria bassiana, an entomopathogenic fungus. One strain was commercial (GHA) and the other had been isolated from Varroa mites in the region (Bb05002). Objectives were to evaluate treatment effect on colony weight, adult bee mass, capped brood, and on Varroa fall onto sticky boards. Treatments included conidia formulated with either carnauba or candelilla wax powder, candelilla wax powder alone, or control; in two treatment groups formulation was applied a second time after one week. Treatment did not affect colony health. Colonies treated twice with Bb05002 conidia and carnauba wax powder had significantly higher mite fall compared to colonies treated with blank candelilla wax powder. The proportion of fallen mites that were infected in both conidia treatments was higher than controls for 18 days after the second treatment. The number of fungal propagules on the bees themselves remained elevated for about 14 days after the second treatment. These results were compared to published results from previous experiments with regard to infection duration.

Rapid Detection of Bacillus anthracis Spores Using Immunomagnetic Separation and Amperometry

PubMed Central

Waller, David F.; Hew, Brian E.; Holdaway, Charlie; Jen, Michael; Peckham, Gabriel D.

2016-01-01

Portable detection and quantitation methods for Bacillus anthracis (anthrax) spores in pure culture or in environmental samples are lacking. Here, an amperometric immunoassay has been developed utilizing immunomagnetic separation to capture the spores and remove potential interferents from test samples followed by amperometric measurement on a field-portable instrument. Antibody-conjugated magnetic beads and antibody-conjugated glucose oxidase were used in a sandwich format for the capture and detection of target spores. Glucose oxidase activity of spore pellets was measured indirectly via amperometry by applying a bias voltage after incubation with glucose, horseradish peroxidase, and the electron mediator 2,2′-azino-bis (3-ethylbenzthiazoline-6-sulphonic acid). Target capture was mediated by polyclonal antisera, whereas monoclonal antibodies were used for signal generation. This strategy maximized sensitivity (500 target spores, 5000 cfu/mL), while also providing a good specificity for Bacillus anthracis spores. Minimal signal deviation occurs in the presence of environmental interferents including soil and modified pH conditions, demonstrating the strengths of immunomagnetic separation. The simultaneous incubation of capture and detection antibodies and rapid substrate development (5 min) result in short sample-to-signal times (less than an hour). With attributes comparable or exceeding that of ELISA and LFDs, amperometry is a low-cost, low-weight, and practical method for detecting anthrax spores in the field. PMID:27999382

Progressing the global antimalarial portfolio: finding drugs which target multiple Plasmodium life stages.

PubMed

Smith, Paul W; Diagana, Thierry T; Yeung, Bryan K S

2014-01-01

The number of novel antimalarial candidates entering preclinical development has seen an increase over the last several years. Most of these drug candidates were originally identified as hits coming from screening large chemical libraries specifically targeting the asexual blood stages of Plasmodium falciparum. Indeed, a large proportion of the current antimalarial arsenal has mainly targeted the asexual blood stage which is responsible for clinical symptoms of the disease. However, as part of the eradication agenda and to address resistance, any next-generation antimalarial should have additional activity on at least one other parasite life stage, i.e. gametocytocidal and/or tissue schizonticidal activity. We have applied this approach by screening compounds with intrinsic activity on asexual blood stages in assays against sexual and liver stages and identified two new antimalarial chemotypes with activity on multiple parasite life stages. This strategy can be expanded to identify other chemical classes of molecules with similar activity profiles for the next generation antimalarials. The following review summarizes the discovery of the spiroindolones and imidazolopiperazine classes of antimalarials developed by the NGBS consortium (Novartis Institute for Tropical Diseases, Genomic Institute of the Novartis Research Foundation, Biomedical Primate Research Center, and the Swiss Tropical and Public Health Institute) currently in clinical trials.

GmcA Is a Putative Glucose-Methanol-Choline Oxidoreductase Required for the Induction of Asexual Development in Aspergillus nidulans

PubMed Central

Etxebeste, Oier; Herrero-García, Erika; Cortese, Marc S.; Garzia, Aitor; Oiartzabal-Arano, Elixabet; de los Ríos, Vivian; Ugalde, Unai; Espeso, Eduardo A.

2012-01-01

Aspergillus nidulans asexual differentiation is induced by Upstream Developmental Activators (UDAs) that include the bZIP-type Transcription Factor (TF) FlbB. A 2D-PAGE/MS-MS-coupled screen for proteins differentially expressed in the presence and absence of FlbB identified 18 candidates. Most candidates belong to GO term classes involved in osmotic and/or oxidative stress response. Among these, we focused on GmcA, a putative glucose-methanol-choline oxidoreductase which is upregulated in a ΔflbB background. GmcA is not required for growth since no differences were detected in the radial extension upon deletion of gmcA. However, its activity is required to induce conidiation under specific culture conditions. A ΔgmcA strain conidiates profusely under acid conditions but displays a characteristic fluffy aconidial phenotype in alkaline medium. The absence of asexual development in a ΔgmcA strain can be suppressed, on one hand, using high concentrations of non-fermentable carbon sources like glycerol, and on the other hand, when the cMyb-type UDA TF flbD is overexpressed. Overall, the results obtained in this work support a role for GmcA at early stages of conidiophore initiation. PMID:22792266

Evolution of the asexual queen succession system and its underlying mechanisms in termites.

PubMed

Matsuura, Kenji

2017-01-01

One major advantage of sexual reproduction over asexual reproduction is its promotion of genetic variation, although it reduces the genetic contribution to offspring. Queens of social insects double their contribution to the gene pool, while overuse of asexual reproduction may reduce the ability of the colony to adapt to environmental stress because of the loss of genetic diversity. Recent studies have revealed that queens of some termite species can solve this tradeoff by using parthenogenesis to produce the next generation of queens and sexual reproduction to produce other colony members. This reproductive system, known as asexual queen succession (AQS), has been identified in the subterranean termites Reticulitermes speratus, Reticulitermes virginicus and Reticulitermes lucifugus and in the Neotropical higher termites Embiratermes neotenicus and Cavitermes tuberosus The studies presented here have uncovered the unusual modes of reproduction in termites and have aimed to identify their underlying mechanisms. The study of AQS, the mixed use of sexual and asexual reproduction, is of fundamental importance as it may provide a key to solve the evolutionary paradox of sex. © 2017. Published by The Company of Biologists Ltd.

Observations of asexual reproductive strategies in Antarctic hexactinellid sponges from ROV video records

NASA Astrophysics Data System (ADS)

Teixidó, Núria; Gili, Josep-Maria; Uriz, María-J.; Gutt, Julian; Arntz, Wolf E.

2006-04-01

Hexactinellid sponges are one of the structuring taxa of benthic communities on the Weddell Sea shelf (Antarctica). However, little is known about their reproduction patterns (larval development, release, settlement, and recruitment), particularly in relation to sexual and asexual processes in sponge populations. Video stations obtained during several expeditions covering a wide depth range and different areas recorded a high frequency of asexual reproductive strategies (ARS) (bipartition and budding) among hexactinellids. Analysis of seabed video strips between 108 and 256 m depth, representing an area of 1400 m 2, showed that about 28% of these sponges exhibited ARS. The Rossella nuda type dominated most of the video stations and exhibited the highest proportion of budding (35%). This proportion increased with the size class. Size class >20 cm exhibited in all the stations a mean value of 8.3±0.7 (SE) for primary and of 2.5±0.2 (SE) for secondary propagules per sponge, respectively. Results from a shallow station (Stn 059, 117 m depth) showed the highest relative abundance of R. nuda type and budding (>20 cm ˜72%, 10-20 cm ˜60%, 5-10 cm ˜12%, and <5 cm ˜3%). A potential influence of iceberg scouring disturbance on the occurrence of budding and number of propagules also was investigated. We conclude that asexual reproduction in hexactinellid sponges may be more frequent than has been thought before and it may greatly influence the genetic structure of populations.

Rapid inactivation of Penicillium digitatum spores using high-density nonequilibrium atmospheric pressure plasma

NASA Astrophysics Data System (ADS)

Iseki, Sachiko; Ohta, Takayuki; Aomatsu, Akiyoshi; Ito, Masafumi; Kano, Hiroyuki; Higashijima, Yasuhiro; Hori, Masaru

2010-04-01

A promising, environmentally safe method for inactivating fungal spores of Penicillium digitatum, a difficult-to-inactivate food spoilage microorganism, was developed using a high-density nonequilibrium atmospheric pressure plasma (NEAPP). The NEAPP employing Ar gas had a high electron density on the order of 1015 cm-3. The spores were successfully and rapidly inactivated using the NEAPP, with a decimal reduction time in spores (D value) of 1.7 min. The contributions of ozone and UV radiation on the inactivation of the spores were evaluated and concluded to be not dominant, which was fundamentally different from the conventional sterilizations.

Rapid inactivation of Penicillium digitatum spores using high-density nonequilibrium atmospheric pressure plasma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Iseki, Sachiko; Hori, Masaru; Ohta, Takayuki

2010-04-12

A promising, environmentally safe method for inactivating fungal spores of Penicillium digitatum, a difficult-to-inactivate food spoilage microorganism, was developed using a high-density nonequilibrium atmospheric pressure plasma (NEAPP). The NEAPP employing Ar gas had a high electron density on the order of 10{sup 15} cm{sup -3}. The spores were successfully and rapidly inactivated using the NEAPP, with a decimal reduction time in spores (D value) of 1.7 min. The contributions of ozone and UV radiation on the inactivation of the spores were evaluated and concluded to be not dominant, which was fundamentally different from the conventional sterilizations.

Nosema ceranae Can Infect Honey Bee Larvae and Reduces Subsequent Adult Longevity.

PubMed

Eiri, Daren M; Suwannapong, Guntima; Endler, Matthew; Nieh, James C

2015-01-01

Nosema ceranae causes a widespread disease that reduces honey bee health but is only thought to infect adult honey bees, not larvae, a critical life stage. We reared honey bee (Apis mellifera) larvae in vitro and provide the first demonstration that N. ceranae can infect larvae and decrease subsequent adult longevity. We exposed three-day-old larvae to a single dose of 40,000 (40K), 10,000 (10K), zero (control), or 40K autoclaved (control) N. ceranae spores in larval food. Spores developed intracellularly in midgut cells at the pre-pupal stage (8 days after egg hatching) of 41% of bees exposed as larvae. We counted the number of N. ceranae spores in dissected bee midguts of pre-pupae and, in a separate group, upon adult death. Pre-pupae exposed to the 10K or 40K spore treatments as larvae had significantly elevated spore counts as compared to controls. Adults exposed as larvae had significantly elevated spore counts as compared to controls. Larval spore exposure decreased longevity: a 40K treatment decreased the age by which 75% of adult bees died by 28%. Unexpectedly, the low dose (10K) led to significantly greater infection (1.3 fold more spores and 1.5 fold more infected bees) than the high dose (40K) upon adult death. Differential immune activation may be involved if the higher dose triggered a stronger larval immune response that resulted in fewer adult spores but imposed a cost, reducing lifespan. The impact of N. ceranae on honey bee larval development and the larvae of naturally infected colonies therefore deserve further study.

Role of YpeB in Cortex Hydrolysis during Germination of Bacillus anthracis Spores

PubMed Central

Bernhards, Casey B.

2014-01-01

The infectious agent of the disease anthrax is the spore of Bacillus anthracis. Bacterial spores are extremely resistant to environmental stresses, which greatly hinders spore decontamination efforts. The spore cortex, a thick layer of modified peptidoglycan, contributes to spore dormancy and resistance by maintaining the low water content of the spore core. The cortex is degraded by germination-specific lytic enzymes (GSLEs) during spore germination, rendering the cells vulnerable to common disinfection techniques. This study investigates the relationship between SleB, a GSLE in B. anthracis, and YpeB, a protein necessary for SleB stability and function. The results indicate that ΔsleB and ΔypeB spores exhibit similar germination phenotypes and that the two proteins have a strict codependency for their incorporation into the dormant spore. In the absence of its partner protein, SleB or YpeB is proteolytically degraded soon after expression during sporulation, rather than escaping the developing spore. The three PepSY domains of YpeB were examined for their roles in the interaction with SleB. YpeB truncation mutants illustrate the necessity of a region beyond the first PepSY domain for SleB stability. Furthermore, site-directed mutagenesis of highly conserved residues within the PepSY domains resulted in germination defects corresponding to reduced levels of both SleB and YpeB in the mutant spores. These results identify residues involved in the stability of both proteins and reiterate their codependent relationship. It is hoped that the study of GSLEs and interacting proteins will lead to the use of GSLEs as targets for efficient activation of spore germination and facilitation of spore cleanup. PMID:25022853

Administration of Brevibacillus laterosporus spores as a poultry feed additive to inhibit house fly development in feces: a new eco-sustainable concept.

PubMed

Ruiu, L; Satta, A; Floris, I

2014-03-01

The success of a microbial pesticide application against house flies developing in manure should accomplish the uniform mixing of active ingredients with this breeding medium, thus enhancing residual effects. The oral administration of the entomopathogenic bacterium Brevibacillus laterosporus to caged poultry species allows the homogeneous incorporation of its active ingredients with fly breeding media. Feces from treated broilers or hens show toxicity against exposed fly adults and larvae. Insecticidal effects are concentration-dependent with a lethal median concentration (LC50) value of 1.34 × 10(8) and 0.61 × 10(8) spores/g of feces for adults and larvae, respectively. Manure toxicity against flies was maintained as long as chickens were fed a diet containing adequate concentrations of B. laterosporus spores. Toxicity significantly decreased after spore administration to birds was interrupted. When poultry diet contained 10(10) spores/g, mortality of flies reared on feces exceeded 80%. The use of B. lateroporus spores as a feed additive in poultry production systems fostering a more integrated approach to farming is discussed.

The development and structure of thick-walled, multicellular, aerial spores in Diheterospora chlamydosporia (=Verticillium chlamydosporium).

PubMed

Cambell, W P; Griffiths, D A

1975-07-01

The aerial, thick-walled spores in Diheterospara chlamydosporia arose as terminal swellings on erect hyphae. Repeated septation of the continuously swelling spore resulted in a multicellular structure. Immediately after the onset of septation secondary wall material was laid down between the two-layered primary wall and the plasmalemma. The presence of secondary wall material indicates that the multicellular spore is a dictyochlamydospore and not an aleuriospore. The relationship between chlamydospores and aleuriospores in other fungi is discussed.

Basal body structure and composition in the apicomplexans Toxoplasma and Plasmodium.

PubMed

Francia, Maria E; Dubremetz, Jean-Francois; Morrissette, Naomi S

2015-01-01

The phylum Apicomplexa encompasses numerous important human and animal disease-causing parasites, including the Plasmodium species, and Toxoplasma gondii, causative agents of malaria and toxoplasmosis, respectively. Apicomplexans proliferate by asexual replication and can also undergo sexual recombination. Most life cycle stages of the parasite lack flagella; these structures only appear on male gametes. Although male gametes (microgametes) assemble a typical 9+2 axoneme, the structure of the templating basal body is poorly defined. Moreover, the relationship between asexual stage centrioles and microgamete basal bodies remains unclear. While asexual stages of Plasmodium lack defined centriole structures, the asexual stages of Toxoplasma and closely related coccidian apicomplexans contain centrioles that consist of nine singlet microtubules and a central tubule. There are relatively few ultra-structural images of Toxoplasma microgametes, which only develop in cat intestinal epithelium. Only a subset of these include sections through the basal body: to date, none have unambiguously captured organization of the basal body structure. Moreover, it is unclear whether this basal body is derived from pre-existing asexual stage centrioles or is synthesized de novo. Basal bodies in Plasmodium microgametes are thought to be synthesized de novo, and their assembly remains ill-defined. Apicomplexan genomes harbor genes encoding δ- and ε-tubulin homologs, potentially enabling these parasites to assemble a typical triplet basal body structure. Moreover, the UNIMOD components (SAS6, SAS4/CPAP, and BLD10/CEP135) are conserved in these organisms. However, other widely conserved basal body and flagellar biogenesis elements are missing from apicomplexan genomes. These differences may indicate variations in flagellar biogenesis pathways and in basal body arrangement within the phylum. As apicomplexan basal bodies are distinct from their metazoan counterparts, it may be possible to selectively target parasite structures in order to inhibit microgamete motility which drives generation of genetic diversity in Toxoplasma and transmission for Plasmodium.

Investigating the Detrimental Effects of Low Pressure Plasma Sterilization on the Survival of Bacillus subtilis Spores Using Live Cell Microscopy.

PubMed

Fuchs, Felix M; Raguse, Marina; Fiebrandt, Marcel; Madela, Kazimierz; Awakowicz, Peter; Laue, Michael; Stapelmann, Katharina; Moeller, Ralf

2017-11-30

Plasma sterilization is a promising alternative to conventional sterilization methods for industrial, clinical, and spaceflight purposes. Low pressure plasma (LPP) discharges contain a broad spectrum of active species, which lead to rapid microbial inactivation. To study the efficiency and mechanisms of sterilization by LPP, we use spores of the test organism Bacillus subtilis because of their extraordinary resistance against conventional sterilization procedures. We describe the production of B. subtilis spore monolayers, the sterilization process by low pressure plasma in a double inductively coupled plasma reactor, the characterization of spore morphology using scanning electron microscopy (SEM), and the analysis of germination and outgrowth of spores by live cell microscopy. A major target of plasma species is genomic material (DNA) and repair of plasma-induced DNA lesions upon spore revival is crucial for survival of the organism. Here, we study the germination capacity of spores and the role of DNA repair during spore germination and outgrowth after treatment with LPP by tracking fluorescently-labelled DNA repair proteins (RecA) with time-resolved confocal fluorescence microscopy. Treated and untreated spore monolayers are activated for germination and visualized with an inverted confocal live cell microscope over time to follow the reaction of individual spores. Our observations reveal that the fraction of germinating and outgrowing spores is dependent on the duration of LPP-treatment reaching a minimum after 120 s. RecA-YFP (yellow fluorescence protein) fluorescence was detected only in few spores and developed in all outgrowing cells with a slight elevation in LPP-treated spores. Moreover, some of the vegetative bacteria derived from LPP-treated spores showed an increase in cytoplasm and tended to lyse. The described methods for analysis of individual spores could be exemplary for the study of other aspects of spore germination and outgrowth.

A Novel Protocol for Decoating and Permeabilizing Bacterial Spores for Epifluorescent Microscopy

NASA Technical Reports Server (NTRS)

LaDuc, Myron T.; Mohapatra, Bidyut

2014-01-01

Based on previously reported procedures for permeabilizing vegetative bacterial cells, and numerous trial-and-error attempts with bacterial endospores, a protocol was developed for effectively permeabilizing bacterial spores, which facilitated the applicability of fluorescent in situ hybridization (FISH) microscopy. Bacterial endospores were first purified from overgrown, sporulated suspensions of B. pumilus SAFR-032. Purified spores at a concentration of approx equals 10 million spores/mL then underwent proteinase-K treatment, in a solution of 468.5 µL of 100 mM Tris-HCl, 30 µL of 10% SDS, and 1.5 microL of 20 mg/mL proteinase-K for ten minutes at 35 ºC. Spores were then harvested by centrifugation (15,000 g for 15 minutes) and washed twice with sterile phosphate-buffered saline (PBS) solution. This washing process consisted of resuspending the spore pellets in 0.5 mL of PBS, vortexing momentarily, and harvesting again by centrifugation. Treated and washed spore pellets were then resuspended in 0.5 mL of decoating solution, which consisted of 4.8 g urea, 3 mL Milli-Q water, 1 mL 0.5M Tris, 1 mL 1M dithiothreitol (DTT), and 2 mL 10% sodium-dodecylsulfate (SDS), and were incubated at 65 ºC for 15 minutes while being shaken at 165 rpm. Decoated spores were then, once again, washed twice with sterile PBS, and subjected to lysozyme/mutanolysin treatment (7 mg/mL lysozyme and 7U mutanolysin) for 15 minutes at 35 C. Spores were again washed twice with sterile PBS, and spore pellets were resuspended in 1-mL of 2% SDS. This treatment, facilitating inner membrane permeabilization, lasted for ten minutes at room temperature. Permeabilized spores were washed two final times with PBS, and were resuspended in 200 mkcroL of sterile PBS. At this point, the spores were permeable and ready for downstream processing, such as oligonucleotideprobe infiltration, hybridization, and microscopic evaluation. FISH-microscopic imagery confirmed the effective and efficient (˜50% successful permeabilization and recovery) permeabilization of numerous spore preparations. The novelty of the technology developed here is in its applicability to bacterial endospores. While protocols abound for the effective permeabilization of bacterial, archaeal, and eukaryotic vegetative cells, there are no such reliable methods for decoating and permeabilizing bacterial endospores in a manner that is amenable to downstream FISH microscopic analyses. This innovation enables the direct visualization and enumeration of spores via FISH-based microscopic techniques, circumventing the complications that accompany previously required germination regimes. The synergistic enzymatic weakening of the many spore layers facilitates a structural compromise that is just enough to render the spores permeable without degrading the spore to a level, which precludes it from recognition.

The sps Gene Products Affect the Germination, Hydrophobicity, and Protein Adsorption of Bacillus subtilis Spores

PubMed Central

Cangiano, Giuseppina; Sirec, Teja; Panarella, Cristina; Isticato, Rachele; Baccigalupi, Loredana; De Felice, Maurilio

2014-01-01

The multilayered surface of the Bacillus subtilis spore is composed of proteins and glycans. While over 70 different proteins have been identified as surface components, carbohydrates associated with the spore surface have not been characterized in detail yet. Bioinformatic data suggest that the 11 products of the sps operon are involved in the synthesis of polysaccharides present on the spore surface, but an experimental validation is available only for the four distal genes of the operon. Here, we report a transcriptional analysis of the sps operon and a functional study performed by constructing and analyzing two null mutants lacking either all or only the promoter-proximal gene of the operon. Our results show that both sps mutant spores apparently have normal coat and crust but have a small germination defect and are more hydrophobic than wild-type spores. We also show that spores lacking all Sps proteins are highly adhesive and form extensive clumps. In addition, sps mutant spores have an increased efficiency in adsorbing a heterologous enzyme, suggesting that hydrophobic force is a major determinant of spore adsorption and indicating that a deep understanding of the surface properties of the spore is essential for its full development as a surface display platform. PMID:25239894

Flavonoids released naturally from alfalfa promote development of symbiotic glomus spores in vitro.

PubMed

Tsai, S M; Phillips, D A

1991-05-01

Because flavonoids from legumes induce transcription of nodulation genes in symbiotic rhizobial bacteria, it is reasonable to test whether these compounds alter the development of vesicular-arbuscular mycorrhizal (VAM) fungi that infect those plants. Quercetin-3-O-galactoside, the dominant flavonoid released naturally from alfalfa (Medicago sativa L.) seeds, promoted spore germination of Glomus etunicatum and Glomus macrocarpum in vitro. Quercetin produced the maximum increases in spore germination, hyphal elongation, and hyphal branching in G. etunicatum at 1 to 2.5 muM concentrations. Two flavonoids exuded from alfalfa roots, 4',7-dihydroxyflavone and 4',7-dihydroxyflavanone, also enhanced spore germination of this fungal species. Formononetin, an isoflavone that is released from stressed alfalfa roots, inhibited germination of both Glomus species. These in vitro results suggest that plant flavonoids may facilitate or regulate the development of VAM symbioses and offer new hope for developing pure, plant-free cultures of VAM fungi.

Development of a heat-stable and orally delivered recombinant M2e-expressing B. subtilis spore-based influenza vaccine.

PubMed

Zhao, Guangyu; Miao, Yu; Guo, Yan; Qiu, Hongjie; Sun, Shihui; Kou, Zhihua; Yu, Hong; Li, Junfeng; Chen, Yue; Jiang, Shibo; Du, Lanying; Zhou, Yusen

2014-01-01

Highly conserved ectodomain of influenza virus M2 protein (M2e) is an important target for the development of universal influenza vaccines. Today, the use of chemical or genetic fusion constructs have been undertaken to overcome the low immunogenicity of M2e in vaccine formulation. However, current M2e vaccines are neither orally delivered nor heat-stable. In this study, we evaluated the immune efficacy of an orally delivered recombinant M2e vaccine containing 3 molcules of M2e consensus sequence of influenza A viruses, termed RSM2e3. To accomplish this, CotB, a spore coat of Bacillus subtilis (B. subtilis), was used as a fusion partner, and heat-stable nonpathogenic B. subtilis spores were used as the carrier. Our results showed that CotB-M2e3 fusion had no effect on spore structure or function in the resultant recombinant RSM2e3 strain and that heterologous influenza virus M2e protein was successfully displayed on the surface of the recombinant RSM2e3 spore. Importantly, recombinant RSM2e3 spores elicited strong and long-term M2e-specific systemic and mucosal immune responses, completely protecting immunized mice from lethal challenge of A/PR/8/34(H1N1) influenza virus. Taken together, our study forms a solid basis for the development of a novel orally delivered and heat-stable influenza vaccine based on B. subtilis spore surface display.

Temporal and non-permanent division of labor during sorocarp formation in the social amoeba Acytostelium subglobosum.

PubMed

Mohri, Kurato; Kiyota, Yu; Kuwayama, Hidekazu; Urushihara, Hideko

2013-03-15

Somatic cell differentiation is crucial for the development of multicellular organisms. While the development of a fruiting body in Dictyostelium discoideum represents a simple model of this process with separation of stalk cells from the spore lineage, that of Acytostelium subglobosum is not accompanied by cell type separation. This species produces acellular stalks and, seemingly, all aggregated amoebae become spores; however, it possesses homologs for the stalk-cell marker genes of D. discoideum. In this study, we analyzed the spatio-temporal expression of A. subglobosum orthologs for D. discoideum stalk- or spore-lineage markers to clarify the developmental process of A. subglobosum. We first found that the prespore vesicles, which contained spore coat proteins, started to accumulate in the tip region and were observed in the entire sorogen throughout later development, confirming that all A. subglobosum cells became spores. The expression of a stalk-lineage gene ortholog, As-ecmA, started at the mound stage and was prominent in the protruding sorogen. Although two spore-lineage gene orthologs, As-cotD1 and -cotD2, were likewise detected shortly after cell aggregation and increased in intensity until tip formation, their expression diminished in the protruding sorogen. Double-fluorescence staining of these prestalk and prespore marker genes revealed that the expression of these marker genes was mutually exclusive and that expression switching occurred in the early tip. Our results indicate that A. subglobosum cells become committed to the spore lineage first, and then, while keeping this commitment intact, participate in stalk formation. Instead of the permanent division of labor observed in D. discoideum, A. subglobosum produces fruiting bodies by all cells contributing to the formation of the stalk as well as forming spores. Copyright © 2013 Elsevier Inc. All rights reserved.

Continuous development precludes radioprotection in a colonial ascidian.

PubMed

Laird, Diana J; Weissman, Irving L

2004-03-01

Colonial organisms provide a unique experimental system for stem cell biology. The colonial Urochordate Botryllus schlosseri reproduces sexually as well as by continuous asexual budding. Adjacent colonies with a shared histocompatibility allele undergo vascular fusion and establish a common blood circulation, performing natural transplantation. Fused colonies become chimeras, often with complete somatic replacement of the host cell genotype by the fused parabiont. We attempted to establish a radioprotection assay for the somatic stem cells that induce long-term chimerism in Botryllus. We demonstrate over a range of radiation doses that neither autologous nor allogeneic cell transplantation enhances survival of host colonies. This suggests that high mitotic index associated with continuous asexual development leads to radiosensitivity of organs and structures essential to survival during engraftment. We observe that radiation induces uncontrolled epithelial cell proliferation in abnormally terminated buds, suggesting that stem cells are not required for the initial stages of bud development.

Binding Affinity of Glycoconjugates to BACILLUS Spores and Toxins

NASA Astrophysics Data System (ADS)

Rasol, Aveen; Eassa, Souzan; Tarasenko, Olga

2010-04-01

Early recognition of Bacillus cereus group species is important since they can cause food-borne illnesses and deadly diseases in humans. Glycoconjugates (GCs) are carbohydrates covalently linked to non-sugar moieties including lipids, proteins or other entities. GCs are involved in recognition and signaling processes intrinsic to biochemical functions in cells. They also stimulate cell-cell adhesion and subsequent recognition and activation of receptors. We have demonstrated that GCs are involved in Bacillus cereus spore recognition. In the present study, we have investigated whether GCs possess the ability to bind and recognize B. cereus spores and Bacillus anthracis recombinant single toxins (sTX) and complex toxins (cTX). The affinity of GCs to spores + sTX and spores + cTX toxins was studied in the binding essay. Our results demonstrated that GC9 and GC10 were able to selectively bind to B. cereus spores and B. anthracis toxins. Different binding affinities for GCs were found toward Bacillus cereus spores + sTX and spores + cTX. Dilution of GCs does not impede the recognition and binding. Developed method provides a tool for simultaneous recognition and targeting of spores, bacteria toxins, and/or other entities.

Rapid detection of Bacillus anthracis spores using a super-paramagnetic lateral-flow immunological detection system.

PubMed

Wang, Dian-Bing; Tian, Bo; Zhang, Zhi-Ping; Deng, Jiao-Yu; Cui, Zong-Qiang; Yang, Rui-Fu; Wang, Xu-Ying; Wei, Hong-Ping; Zhang, Xian-En

2013-04-15

There is an urgent need for convenient, sensitive, and specific methods to detect the spores of Bacillus anthracis, the causative agent of anthrax, because of the bioterrorism threat posed by this bacterium. In this study, we firstly develop a super-paramagnetic lateral-flow immunological detection system for B. anthracis spores. This system involves the use of a portable magnetic assay reader, super-paramagnetic iron oxide particles, lateral-flow strips and two different monoclonal antibodies directed against B. anthracis spores. This detection system specifically recognises as few as 400 pure B. anthracis spores in 30 min. This system has a linear range of 4×10³-10⁶ CFU ml⁻¹ and reproducible detection limits of 200 spores mg⁻¹ milk powder and 130 spores mg⁻¹ soil for simulated samples. In addition, this approach shows no obvious cross-reaction with other related Bacillus spores, even at high concentrations, and has no significant dependence on the duration of the storage of the immunological strips. Therefore, this super-paramagnetic lateral-flow immunological detection system is a promising tool for the rapid and sensitive detection of Bacillus anthracis spores under field conditions. Copyright © 2012 Elsevier B.V. All rights reserved.

Seasonal variation of Ganoderma spore concentrations in urban and suburban districts of the city of Szczecin, Poland.

PubMed

Grinn-Gofroń, Agnieszka; Strzelczak, Agnieszka; Przestrzelska, Katarzyna

2015-01-01

According to recent studies, Ganoderma may be the third genus, after Alternaria and Cladosporium, the spores of which cause symptoms of allergy, and concentration is related to meteorological factors. The aerobiology of Ganoderma spores in Szczecin in urban and suburban districts was examined using Lanzoni Volumetric Spore Traps in 2008-2010. Ganoderma spores were present in the atmosphere on more than 90% of the days from June through September with peak concentrations in June, July and September. The number of days with spores was lower in the suburban district, while the total number of spores collected was higher there than in the urban district. Correlation and multiple regression analyses revealed weak relationships between Ganoderma and meteorological conditions, while testing the significance of differences between the districts showed that urban development did not have a clear impact on the values of meteorological parameters. A significantly higher abundance of spores in the suburbs of Szczecin seemed to be conditioned by the closeness of potential area sources. This study indicates that a single measuring site in the city centre insufficiently reflected the dynamics and level of Ganoderma spore concentration in peripheral districts.

Near-infrared surface-enhanced-Raman-scattering (SERS) mediated detection of single optically trapped bacterial spores

NASA Astrophysics Data System (ADS)

Alexander, Troy A.; Pellegrino, Paul M.; Gillespie, James B.

2003-08-01

A novel methodology has been developed for the investigation of bacterial spores. Specifically, this method has been used to probe the spore coat composition of two different Bacillus stearothermophilus variants. This technique may be useful in many applications; most notably, development of novel detection schemes toward potentially harmful bacteria. This method would also be useful as an ancillary environmental monitoring system where sterility is of importance (i.e., food preparation areas as well as invasive and minimally invasive medical applications). This unique detection scheme is based on the near-infrared (NIR) Surface-Enhanced-Raman-Scattering (SERS) from single, optically trapped, bacterial spores. The SERS spectra of bacterial spores in aqueous media have been measured using SERS substrates based on ~60-nm diameter gold colloids bound to 3-Aminopropyltriethoxysilane derivatized glass. The light from a 787-nm laser diode was used to trap/manipulate as well as simultaneously excite the SERS of an individual bacterial spore. The collected SERS spectra were examined for uniqueness and the applicability of this technique for the strain discrimination of Bacillus stearothermophilus spores. Comparison of normal Raman and SERS spectra reveal not only an enhancement of the normal Raman spectral features but also the appearance of spectral features absent in the normal Raman spectrum.

Near-infrared Surface-Enhanced-Raman-Scattering (SERS) mediated discrimination of single optically trapped bacterial spores

NASA Astrophysics Data System (ADS)

Alexander, Troy A.; Pellegrino, Paul M.; Gillespie, James B.

2004-03-01

A novel methodology has been developed for the investigation of bacterial spores. Specifically, this method has been used to probe the spore coat composition of two different Bacillus stearothermophilus variants. This technique may be useful in many applications; most notably, development of novel detection schemes toward potentially harmful bacteria. This method would also be useful as an ancillary environmental monitoring system where sterility is of importance (i.e., food preparation areas as well as invasive and minimally invasive medical applications). This unique detection scheme is based on the near-infrared (NIR) Surface-Enhanced-Raman- Scattering (SERS) from single, optically trapped, bacterial spores. The SERS spectra of bacterial spores in aqueous media have been measured using SERS substrates based on ~60-nm diameter gold colloids bound to 3-Aminopropyltriethoxysilane derivatized glass. The light from a 787-nm laser diode was used to trap/manipulate as well as simultaneously excite the SERS of an individual bacterial spore. The collected SERS spectra were examined for uniqueness and the applicability of this technique for the strain discrimination of Bacillus stearothermophilus spores. Comparison of normal Raman and SERS spectra reveal not only an enhancement of the normal Raman spectral features but also the appearance of spectral features absent in the normal Raman spectrum.

Small acid soluble proteins for rapid spore identification.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Branda, Steven S.; Lane, Todd W.; VanderNoot, Victoria A.

2006-12-01

This one year LDRD addressed the problem of rapid characterization of bacterial spores such as those from the genus Bacillus, the group that contains pathogenic spores such as B. anthracis. In this effort we addressed the feasibility of using a proteomics based approach to spore characterization using a subset of conserved spore proteins known as the small acid soluble proteins or SASPs. We proposed developing techniques that built on our previous expertise in microseparations to rapidly characterize or identify spores. An alternative SASP extraction method was developed that was amenable to both the subsequent fluorescent labeling required for laser-induced fluorescencemore » detection and the low ionic strength requirements for isoelectric focusing. For the microseparations, both capillary isoelectric focusing and chip gel electrophoresis were employed. A variety of methods were evaluated to improve the molecular weight resolution for the SASPs, which are in a molecular weight range that is not well resolved by the current methods. Isoelectric focusing was optimized and employed to resolve the SASPs using UV absorbance detection. Proteomic signatures of native wild type Bacillus spores and clones genetically engineered to produce altered SASP patterns were assessed by slab gel electrophoresis, capillary isoelectric focusing with absorbance detection as well as microchip based gel electrophoresis employing sensitive laser-induced fluorescence detection.« less

Risk factors for Plasmodium falciparum and Plasmodium vivax gametocyte carriage in Papua New Guinean children with uncomplicated malaria.

PubMed

Karl, Stephan; Laman, Moses; Moore, Brioni R; Benjamin, John M; Salib, Mary; Lorry, Lina; Maripal, Samuel; Siba, Peter; Robinson, Leanne J; Mueller, Ivo; Davis, Timothy M E

2016-08-01

There are limited data on gametocytaemia risk factors before/after treatment with artemisinin combination therapy in children from areas with transmission of multiple Plasmodium species. We utilised data from a randomised trial comparing artemether-lumefantrine (AL) and artemisinin-naphthoquine (AN) in 230 Papua New Guinean children aged 0.5-5 years with uncomplicated malaria in whom determinants of gametocytaemia by light microscopy were assessed at baseline using logistic regression and during follow-up using multilevel mixed effects modelling. Seventy-four (32%) and 18 (8%) children presented with P. falciparum and P. vivax gametocytaemia, respectively. Baseline P. falciparum gametocytaemia was associated with Hackett spleen grade 1 (odds ratio (95% CI) 4.01 (1.60-10.05) vs grade 0; P<0.001) and haemoglobin (0.95 (0.92-0.97) per 1g/L increase; P<0.001), and P. falciparum asexual parasitaemia in slide-positive cases (0.36 (0.19-0.68) for a 10-fold increase; P=0.002). Baseline P. vivax gametocytaemia was associated with Hackett grade 2 (12.66 (1.31-122.56); P=0.028), mixed P. falciparum/vivax infection (0.16 (0.03-1.00); P=0.050), P. vivax asexual parasitaemia (5.68 (0.98-33.04); P=0.053) and haemoglobin (0.94 (0.88-1.00); P=0.056). For post-treatment P. falciparum gametocytaemia, independent predictors were AN vs AL treatment (4.09 (1.43-11.65)), haemoglobin (0.95 (0.93-0.97)), presence/absence of P. falciparum asexual forms (3.40 (1.66-0.68)) and day post-treatment (0.086 (0.82-0.90)) (P<0.001). Post-treatment P. vivax gametocytaemia was predicted by presence of P. vivax asexual forms (596 (12-28,433); P<0.001). Consistent with slow P. falciparum gametocyte maturation, low haemoglobin, low asexual parasite density and higher spleen grading, markers of increased prior infection exposure/immunity, were strong associates of pre-treatment gametocyte positivity. The persistent inverse association between P. falciparum gametocytaemia and haemoglobin during follow-up suggests an important role for bone marrow modulation of gametocytogenesis. In P. vivax infections, baseline and post-treatment gametocyte carriage was positively related to the acute parasite burden, reflecting the close association between the development of asexual and sexual forms. Copyright © 2016 Elsevier B.V. All rights reserved.

Deposition of Bacillus subtilis spores using an airbrush-spray or spots to study surface decontamination by pulsed light.

PubMed

Levy, Caroline; Bornard, Isabelle; Carlin, Frédéric

2011-02-01

Microbial contamination on surfaces of food processing equipment is a major concern in industries. A new method to inoculate a single-cell layer (monolayer) of microorganisms onto polystyrene was developed, using a deposition with an airbrush. A homogeneous dispersion of Bacillus subtilis DSM 402 spores sprayed on the surface was observed using both plate count and scanning electron microscopy. No clusters were found, even with high spore concentrations (10(7) spores/inoculated surface). A monolayer of microorganisms was also obtained after deposition of 10 μL droplets containing 3×10(4) spores/spot on polystyrene disks, but not with a higher spore concentration. Pulsed light (PL) applied to monolayers of B. subtilis spores allowed log reductions higher than 6. As a consequence of clusters formation in spots of 10 μL containing more than 3×10(5) spores, log reductions obtained by PL were significantly lower. The comparative advantages of spot and spray depositions were discussed. Copyright © 2010 Elsevier B.V. All rights reserved.

Measurement of 100 B. anthracis Ames spores within 15 minutes by SERS at the US Army Edgewood Chemical Biological Ctr.

NASA Astrophysics Data System (ADS)

Farquharson, Stuart; Shende, Chetan; Smith, Wayne; Huang, Hermes; Sperry, Jay; Sickler, Todd; Prugh, Amber; Guicheteau, Jason

2014-05-01

Since the distribution of Bacillus anthracis-Ames spores through the US Postal System, there has been a persistent fear that biological warfare agents will be used by terrorists against our military abroad and our civilians at home. While there has been substantial effort since the anthrax attack of 2001 to develop analyzers to detect this and other biological warfare agents, the analyzers remain either too slow, lack sensitivity, produce high false-positive rates, or cannot be fielded. In an effort to overcome these limitations we have been developing a surface-enhanced Raman spectroscopy system. Here we describe the use of silver nanoparticles functionalized with a short peptide to selectively capture Bacillus anthracis spores and produce SER scattering. Specifically, measurements of 100 B. anthracis-Ames spores/mL in ~25 minutes performed at the US Army's Edgewood Chemical Biological Center are presented. The measurements provide a basis for the development of systems that can detect spores collected from the air or water supplies with the potential of saving lives during a biological warfare attack.

Dielectric Study of the Physical State of Electrolytes and Water Within Bacillus cereus Spores

PubMed Central

Carstensen, Edwin L.; Marquis, Robert E.; Gerhardt, Philipp

1971-01-01

Dielectric measurements revealed that dormant spores of Bacillus cereus have extremely low conductivities at high frequencies (50 MHz) and so must contain remarkably low concentrations of mobile ions both within the core and in the surrounding integuments. Activation, germination, and outgrowth were all accompanied by increases in conductivity of the cells and their suspending medium, and this result indicated that intracellular electrolytes had become ionized and leaked from the spores. High-frequency dielectric constants of spores were consistent with normal states for cell water. These values increased during successive stages of development from dormant spore to vegetative bacillus, and they could be directly related to increases in cell water content. In all, the results refuted a model of the dormant spore involving freely mobile, ionized electrolytes and supported a model involving electrostatically bound electrolytes. PMID:4998245

Inactivation of Clostridium perfringens spores adhered onto stainless steel surface by agents used in a clean-in-place procedure.

PubMed

Alzubeidi, Yasmeen S; Udompijitkul, Pathima; Talukdar, Prabhat K; Sarker, Mahfuzur R

2018-07-20

Enterotoxigenic Clostridium perfringens, a leading foodborne pathogen can be cross-contaminated from food processing stainless steel (SS) surfaces to the finished food products. This is mostly due to the high resistance of C. perfringens spores adhered onto SS surfaces to various disinfectants commonly used in food industries. In this study, we aimed to investigate the survivability and adherence of C. perfringens spores onto SS surfaces and then validate the effectiveness of a simulated Clean-in-Place (CIP) regime on inactivation of spores adhered onto SS surfaces. Our results demonstrated that, 1) C. perfringens spores adhered firmly onto SS surfaces and survived for at-least 48 h, unlike their vegetative cells who died within 30 min, after aerobic incubation at refrigerated and ambient temperatures; 2) Spores exhibited higher levels of hydrophobicity than vegetative cells, suggesting a correlation between cell surface hydrophobicity and adhesion to solid surfaces; 3) Intact spores were more hydrophobic than the decoated spores, suggesting a positive role of spore coat components on spores' hydrophobicity and thus adhesion onto SS surfaces; and finally 4) The CIP regime (NaOH + HNO 3 ) successfully inactivated C. perfringens spores adhered onto SS surfaces, and most of the effect of CIP regime appeared to be due to the NaOH. Collectively, our current findings may well contribute towards developing a strategy to control cross-contamination of C. perfringens spores into food products, which should help reducing the risk of C. perfringens-associated food poisoning outbreaks. Copyright © 2018 Elsevier B.V. All rights reserved.

Spore populations among bulk tank raw milk and dairy powders are significantly different.

PubMed

Miller, Rachel A; Kent, David J; Watterson, Matthew J; Boor, Kathryn J; Martin, Nicole H; Wiedmann, Martin

2015-12-01

To accommodate stringent spore limits mandated for the export of dairy powders, a more thorough understanding of the spore species present will be necessary to develop prospective strategies to identify and reduce sources (i.e., raw materials or in-plant) of contamination. We characterized 1,523 spore isolates obtained from bulk tank raw milk (n=33 farms) and samples collected from 4 different dairy powder-processing plants producing acid whey, nonfat dry milk, sweet whey, or whey protein concentrate 80. The spores isolated comprised 12 genera, at least 44 species, and 216 rpoB allelic types. Bacillus and Geobacillus represented the most commonly isolated spore genera (approximately 68.9 and 12.1%, respectively, of all spore isolates). Whereas Bacillus licheniformis was isolated from samples collected from all plants and farms, Geobacillus spp. were isolated from samples from 3 out of 4 plants and just 1 out of 33 farms. We found significant differences between the spore population isolated from bulk tank raw milk and those isolated from dairy powder plant samples, except samples from the plant producing acid whey. A comparison of spore species isolated from raw materials and finished powders showed that although certain species, such as B. licheniformis, were found in both raw and finished product samples, other species, such as Geobacillus spp. and Anoxybacillus spp., were more frequently isolated from finished powders. Importantly, we found that 8 out of 12 genera were isolated from at least 2 different spore count methods, suggesting that some spore count methods may provide redundant information if used in parallel. Together, our results suggest that (1) Bacillus and Geobacillus are the predominant spore contaminants in a variety of dairy powders, implying that future research efforts targeted at elucidating approaches to reduce levels of spores in dairy powders should focus on controlling levels of spore isolates from these genera; and (2) the spore populations isolated from bulk tank raw milk and some dairy powder products are significantly different, suggesting that targeting in-plant sources of contamination may be important for achieving low spore counts in the finished product. These data provide important insight regarding the diversity of spore populations isolated from dairy powders and bulk tank raw milk, and demonstrate that several spore genera are detected by multiple spore count methods. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Fern Spore Longevity in Saline Water: Can Sea Bottom Sediments Maintain a Viable Spore Bank?

PubMed Central

de Groot, G. Arjen; During, Heinjo

2013-01-01

Freshwater and marine sediments often harbor reservoirs of plant diaspores, from which germination and establishment may occur whenever the sediment falls dry. Therewith, they form valuable records of historical inter- and intraspecific diversity, and are increasingly exploited to facilitate diversity establishment in new or restored nature areas. Yet, while ferns may constitute a considerable part of a vegetation’s diversity and sediments are known to contain fern spores, little is known about their longevity, which may suffer from inundation and - in sea bottoms - salt stress. We tested the potential of ferns to establish from a sea or lake bottom, using experimental studies on spore survival and gametophyte formation, as well as a spore bank analysis on sediments from a former Dutch inland sea. Our experimental results revealed clear differences among species. For Asplenium scolopendrium and Gymnocarpium dryopteris, spore germination was not affected by inundated storage alone, but decreased with rising salt concentrations. In contrast, for Asplenium trichomanes subsp. quadrivalens germination decreased following inundation, but not in response to salt. Germination rates decreased with time of storage in saline water. Smaller and less viable gametophytes were produced when saline storage lasted for a year. Effects on germination and gametophyte development clearly differed among genotypes of A. scolopendrium. Spore bank analyses detected no viable spores in marine sediment layers. Only two very small gametophytes (identified as Thelypteris palustris via DNA barcoding) emerged from freshwater sediments. Both died before maturation. We conclude that marine, and likely even freshwater sediments, will generally be of little value for long-term storage of fern diversity. The development of any fern vegetation on a former sea floor will depend heavily on the deposition of spores onto the drained land by natural or artificial means of dispersal. PMID:24223951

The Spore Coat Protein CotE Facilitates Host Colonization by Clostridium difficile

PubMed Central

Hong, Huynh A; Ferreira, William T; Hosseini, Siamand; Anwar, Saba; Hitri, Krisztina; Wilkinson, Anthony J; Vahjen, Wilfried; Zentek, Jürgen; Soloviev, Mikhail; Cutting, Simon M

2017-01-01

Abstract Clostridium difficile infection (CDI) is an important hospital-acquired infection resulting from the germination of spores in the intestine as a consequence of antibiotic-mediated dysbiosis of the gut microbiota. Key to this is CotE, a protein displayed on the spore surface and carrying 2 functional elements, an N-terminal peroxiredoxin and a C-terminal chitinase domain. Using isogenic mutants, we show in vitro and ex vivo that CotE enables binding of spores to mucus by direct interaction with mucin and contributes to its degradation. In animal models of CDI, we show that when CotE is absent, both colonization and virulence were markedly reduced. We demonstrate here that the attachment of spores to the intestine is essential in the development of CDI. Spores are usually regarded as biochemically dormant, but our findings demonstrate that rather than being simply agents of transmission and dissemination, spores directly contribute to the establishment and promotion of disease. PMID:28968845

Detection of Gene Flow from Sexual to Asexual Lineages in Thrips tabaci (Thysanoptera: Thripidae)

PubMed Central

Li, Xiao-Wei; Wang, Ping; Fail, Jozsef; Shelton, Anthony M.

2015-01-01

Populations of Thrips tabaci are known to have two sympatric but genetically isolated reproductive modes, arrhenotoky (sexual reproduction) and thelytoky (asexual reproduction). Herein, we report behavioral, ecological and genetic studies to determine whether there is gene flow between arrhenotokous and thelytokous T. tabaci. We did not detect significant preference by arrhenotokous males to mate with females of a particular reproductive mode, nor did we detect significant behavioral differences between arrhenotokous males mated with arrhenotokous or thelytokous females in their pre-copulation, copulation duration and mating frequency. Productive gene transfer resulting from the mating between the two modes was experimentally confirmed. Gene transfer from arrhenotokous T. tabaci to thelytokous T. tabaci was further validated by confirmation of the passage of the arrhenotokous male-originated nuclear gene (histone H3 gene) allele to the F2 generation. These behavioral, ecological and genetic studies confirmed gene transfer from the sexual arrhenotokous mode to the asexual thelytokous mode of T. tabaci in the laboratory. These results demonstrate that asexual T. tabaci populations may acquire genetic variability from sexual populations, which could offset the long-term disadvantage of asexual reproduction. PMID:26375283

Contribution of Spores to the Ability of Clostridium difficile To Adhere to Surfaces

PubMed Central

Joshi, Lovleen Tina; Phillips, Daniel S.; Williams, Catrin F.; Alyousef, Abdullah

2012-01-01

Clostridium difficile is the commonest cause of hospital-acquired infection in the United Kingdom. We characterized the abilities of 21 clinical isolates to form spores; to adhere to inorganic and organic surfaces, including stainless steel and human adenocarcinoma cells; and to germinate. The composition of culture media had a significant effect on spore formation, as significantly more spores were produced in brain heart infusion broth (Student's t test; P = 0.018). The spore surface relative hydrophobicity (RH) varied markedly (14 to 77%) and was correlated with the ability to adhere to stainless steel. We observed no correlation between the ribotype and the ability to adhere to steel. When the binding of hydrophobic (DS1813; ribotype 027; RH, 77%) and hydrophilic (DS1748; ribotype 002; RH, 14%) spores to human gut epithelial cells at different stages of cell development was examined, DS1813 spores adhered more strongly, suggesting the presence of surface properties that aid attachment to human cells. Electron microscopy studies revealed the presence of an exosporium surrounding DS1813 spores that was absent from spores of DS1748. Finally, the ability of spores to germinate was found to be strain and medium dependent. While the significance of these findings to the disease process has yet to be determined, this study has highlighted the importance of analyzing multiple isolates when attempting to characterize the behavior of a bacterial species. PMID:22923404

The Dr-nanos gene is essential for germ cell specification in the planarian Dugesia ryukyuensis.

PubMed

Nakagawa, Haruka; Ishizu, Hirotsugu; Chinone, Ayako; Kobayashi, Kazuya; Matsumoto, Midori

2012-01-01

Homologs of nanos are required for the formation and maintenance of germline stem cell (GSC) systems and for gametogenesis in many metazoans. Planarians can change their reproductive mode seasonally, alternating between asexual and sexual reproduction; they develop and maintain their somatic stem cells (SSCs) and GCSs from pluripotent stem cells known as neoblasts. We isolated a nanos homolog, Dr-nanos, from the expressed sequence tags (ESTs) of the sexualized form of Dugesia ryukyuensis. We examined the expression of Dr-nanos in asexual and sexualized planarians by in situ hybridization and analyzed its function using RNA interference (RNAi) together with a planarian sexualization assay. A nanos homolog, Dr-nanos, was identified in the planarian D. ryukyuensis. Dr-nanos expression was observed in the ovarian primordial cells of the asexual worms. This expression increased in proportion to sexualization and was localized in the early germline cells of the ovaries and testes. In X-ray-irradiated worms, the expression of Dr-nanos decreased to a large extent, indicating that Dr-nanos is expressed in some subpopulations of stem cells, especially in GSCs. During the sexualization process, worms in which Dr-nanos was knocked down by RNAi exhibited decreased numbers of oogonia in the ovaries and failed to develop testes, whereas the somatic sexual organs were not affected. We conclude that Dr-nanos is essential for the development of germ cells in the ovaries and testes and may have a function in the early stages of germ cell specification, but not in the development of somatic sexual organs.

Redeployment of germ layers related TFs shows regionalized expression during two non-embryonic developments.

PubMed

Ricci, Lorenzo; Cabrera, Fabien; Lotito, Sonia; Tiozzo, Stefano

2016-08-01

In all non-vertebrate metazoan phyla, species that evolved non-embryonic developmental pathways as means of propagation or regeneration can be found. In this context, new bodies arise through asexual reproduction processes (such as budding) or whole body regeneration, that lack the familiar temporal and spatial cues classically associated with embryogenesis, like maternal determinants, or gastrulation. The molecular mechanisms underlying those non-embryonic developments (i.e., regeneration and asexual reproduction), and their relationship to those deployed during embryogenesis are poorly understood. We have addressed this question in the colonial ascidian Botryllus schlosseri, which undergoes an asexual reproductive process via palleal budding (PB), as well as a whole body regeneration by vascular budding (VB). We identified early regenerative structures during VB and then followed the fate of differentiating tissues during both non-embryonic developments (PB and VB) by monitoring the expression of genes known to play key functions in germ layer specification with well conserved expression patterns in solitary ascidian embryogenesis. The expression patterns of FoxA1, GATAa, GATAb, Otx, Bra, Gsc and Tbx2/3 were analysed during both PB and VB. We found that the majority of these transcription factors were expressed during both non-embryonic developmental processes, revealing a regionalization of the palleal and vascular buds. Knockdown of GATAa by siRNA in palleal buds confirmed that preventing the correct development of one of these regions blocks further tissue specification. Our results indicate that during both normal and injury-induced budding, a similar alternative developmental program operates via early commitment of epithelial regions. Copyright © 2016. Published by Elsevier Inc.

Macroalgal spore dysfunction: ocean acidification delays and weakens adhesion.

PubMed

Guenther, Rebecca; Miklasz, Kevin; Carrington, Emily; Martone, Patrick T

2018-04-01

Early life stages of marine organisms are predicted to be vulnerable to ocean acidification. For macroalgae, reproduction and population persistence rely on spores to settle, adhere and continue the algal life cycle, yet the effect of ocean acidification on this critical life stage has been largely overlooked. We explicitly tested the biomechanical impact of reduced pH on early spore adhesion. We developed a shear flume to examine the effect of reduced pH on spore attachment time and strength in two intertidal rhodophyte macroalgae, one calcified (Corallina vancouveriensis) and one noncalcified (Polyostea robusta). Reduced pH delayed spore attachment of both species by 40%-52% and weakened attachment strength in C. vancouveriensis, causing spores to dislodge at lower flow-induced shear forces, but had no effect on the attachment strength of P. robusta. Results are consistent with our prediction that reduced pH disrupts proper curing and gel formation of spore adhesives (anionic polysaccharides and glycoproteins) via protonation and cation displacement, although experimental verification is needed. Our results demonstrate that ocean acidification negatively, and differentially, impacts spore adhesion in two macroalgae. If results hold in field conditions, reduced ocean pH has the potential to impact macroalgal communities via spore dysfunction, regardless of the physiological tolerance of mature thalli. © 2017 Phycological Society of America.

Effects of nifedipine on gravi-dependent germination of moss spores

NASA Astrophysics Data System (ADS)

Khorkavtsiv, O. Y.; Demkiv, O. T.

Influence of gravity on germination of spores and dependence of the generation of a polar axis on a Ca2+ influx were investigated. The germination of spores does not depend on gravity but outgrowth polarity is controlled by light and gravity (Sytnik et al., 1989; Pundiak et al., 2001). We have shown that gravity determines the polarity of germination of spores and development of rhizoid and chloronemal outgrowths in both moss species -- Ceratodon purpureus and Pohlia nutans, the alignment of polar of germinating spores in C. purpureus, however, is less dependent on gravistimulus than in P. nutans. In 48 h after sowing onto culture medium+0,2% glucose in vertically oriented petri dishes in darkness spores of P. nutans germinated positively gravitropic rhizoid at the lower spore side and negatively gravitropic chloronema at the opposite one. The germination of C. purpureus spores is similar but the outgrowths show the lower level of alignment to the gravity vector than that of P. nutans, the dispersion of angles being 8,9 vs. 1,2 respectively. The cellular mechanism by which gravity acts remains unknown. The intracellular signaling Ca2+ ions play a crucial role in gravity perception and ability of a single cell to respond to gravity. We determined relative intensity of Ca2+ luminescence in the spores before their germination and at the early stages of outgrowth formation after treatment with the nifedipine and in a dependence on gravity vector. Gravity determined the position of outgrowth initiation zone and later on the growth direction of spore filaments. Treatment with nifedipine suppressed the gravity-directed calcium channel influx and distrupted polar growth of outgrowths. In experiments with calcium channel blocker sterilized spores were pregerminated on normal Knop's agar one day after were transferred to 50 μ M nifedipine just before emergence of the germ tube. After 48 h on nifedipine treatment, 50% spores did not germinate, 35% grew apolarily and in 15% of spores cell filaments oriented parallely with respect to the gravity vector. Results shown suggest that the endogenic competency of a single-cell spore is necessary condition of gravi- induced initiation of polar axis the competency being realized with Ca2+ movement. The highest level of Ca2+ luminescence was at the bottom of spores. In other sites of the spores the Ca2+ luminescence was about 20-fold lower than at the site of Ca2+ influx. In the 24 h after formation of first outgrowth the new site of Ca2+ influx appeared at the opposite site of spore and the second outgrowth arised. Consequently during the period of gravi-dependent spore development the newly top Ca2+ influx was repeatedly established. The direction of the Ca2+ ions influx correlated with re-orientation of spores with respect to the gravity vector. It is known that the nifedipine partially inhibits polar axis formation (Chatterjee et al., 2000) the latter being formed under the influence Ca2+ gradient (Cove, 2000). Thus, our results confirm that the fast change of Ca2+ influx probably is one of the earliest cell-level responses induced by gravity and it plays a key role in guiding polar events of germinating spores. This research was supported by NASA grant NN-09 (R).

Strategy to inactivate Clostridium perfringens spores in meat products.

PubMed

Akhtar, Saeed; Paredes-Sabja, Daniel; Torres, J Antonio; Sarker, Mahfuzur R

2009-05-01

The current study aimed to develop an inactivation strategy for Clostridium perfringens spores in meat through a combination of spore activation at low pressure (100-200 MPa, 7 min) and elevated temperature (80 degrees C, 10 min); spore germination at high temperatures (55, 60 or 65 degrees C); and inactivation of germinated spores with elevated temperatures (80 and 90 degrees C, 10 and 20 min) and high pressure (586 MPa, at 23 and 73 degrees C, 10 min). Low pressures (100-200 MPa) were insufficient to efficiently activate C. perfringens spores for germination. However, C. perfringens spores were efficiently activated with elevated temperature (80 degrees C, 10 min), and germinated at temperatures lethal for vegetative cells (>or= 55 degrees C) when incubated for 60 min with a mixture of L-asparagine and KCl (AK) in phosphate buffer (pH 7) and in poultry meat. Inactivation of spores (approximately 4 decimal reduction) in meat by elevated temperatures (80-90 degrees C for 20 min) required a long germination period (55 degrees C for 60 min). However, similar inactivation level was reached with shorter germination period (55 degrees C for 15 min) when spore contaminated-meat was treated with pressure-assisted thermal processing (568 MPa, 73 degrees C, 10 min). Therefore, the most efficient strategy to inactivate C. perfringens spores in poultry meat containing 50 mM AK consisted: (i) a primary heat treatment (80 degrees C, 10 min) to pasteurize and denature the meat proteins and to activate C. perfringens spores for germination; (ii) cooling of the product to 55 degrees C in about 20 min and further incubation at 55 degrees C for about 15 min for spore germination; and (iii) inactivation of germinated spores by pressure-assisted thermal processing (586 MPa at 73 degrees C for 10 min). Collectively, this study demonstrates the feasibility of an alternative and novel strategy to inactivate C. perfringens spores in meat products formulated with germinants specific for C. perfringens.

Multigeneration Cross Contamination of Mail with Bacillus Species Spores by Tumbling ▿

PubMed Central

Edmonds, Jason; Clark, Paul; Williams, Leslie; Lindquist, H. D. Alan; Martinez, Kenneth; Gardner, Warren; Shadomy, Sean; Hornsby-Myers, Jennifer

2010-01-01

In 2001, envelopes loaded with Bacillus anthracis spores were mailed to Senators Daschle and Leahy as well as to the New York Post and NBC News buildings. Additional letters may have been mailed to other news agencies because there was confirmed anthrax infection of employees at these locations. These events heightened the awareness of the lack of understanding of the mechanism(s) by which objects contaminated with a biological agent might spread disease. This understanding is crucial for the estimation of the potential for exposure to ensure the appropriate response in the event of future attacks. In this study, equipment to simulate interactions between envelopes and procedures to analyze the spread of spores from a “payload” envelope (i.e., loaded internally with a powdered spore preparation) onto neighboring envelopes were developed. Another process to determine whether an aerosol could be generated by opening contaminated envelopes was developed. Subsequent generations of contaminated envelopes originating from a single payload envelope showed a consistent two-log decrease in the number of spores transferred from one generation to the next. Opening a tertiary contaminated envelope resulted in an aerosol containing 103 B. anthracis spores. We developed a procedure for sampling contaminated letters by a nondestructive method aimed at providing information useful for consequence management while preserving the integrity of objects contaminated during the incident and preserving evidence for law enforcement agencies. PMID:20511424

Sporangiospore size dimorphism is linked to virulence of Mucor circinelloides.

PubMed

Li, Charles H; Cervantes, Maria; Springer, Deborah J; Boekhout, Teun; Ruiz-Vazquez, Rosa M; Torres-Martinez, Santiago R; Heitman, Joseph; Lee, Soo Chan

2011-06-01

Mucor circinelloides is a zygomycete fungus and an emerging opportunistic pathogen in immunocompromised patients, especially transplant recipients and in some cases otherwise healthy individuals. We have discovered a novel example of size dimorphism linked to virulence. M. circinelloides is a heterothallic fungus: (+) sex allele encodes SexP and (-) sex allele SexM, both of which are HMG domain protein sex determinants. M. circinelloides f. lusitanicus (Mcl) (-) mating type isolates produce larger asexual sporangiospores that are more virulent in the wax moth host compared to (+) isolates that produce smaller less virulent sporangiospores. The larger sporangiospores germinate inside and lyse macrophages, whereas the smaller sporangiospores do not. sexMΔ mutants are sterile and still produce larger virulent sporangiospores, suggesting that either the sex locus is not involved in virulence/spore size or the sexP allele plays an inhibitory role. Phylogenetic analysis supports that at least three extant subspecies populate the M. circinelloides complex in nature: Mcl, M. circinelloides f. griseocyanus, and M. circinelloides f. circinelloides (Mcc). Mcc was found to be more prevalent among clinical Mucor isolates, and more virulent than Mcl in a diabetic murine model in contrast to the wax moth host. The M. circinelloides sex locus encodes an HMG domain protein (SexP for plus and SexM for minus mating types) flanked by genes encoding triose phosphate transporter (TPT) and RNA helicase homologs. The borders of the sex locus between the three subspecies differ: the Mcg sex locus includes the promoters of both the TPT and the RNA helicase genes, whereas the Mcl and Mcc sex locus includes only the TPT gene promoter. Mating between subspecies was restricted compared to mating within subspecies. These findings demonstrate that spore size dimorphism is linked to virulence of M. circinelloides species and that plasticity of the sex locus and adaptations in pathogenicity have occurred during speciation of the M. circinelloides complex.

Sporangiospore Size Dimorphism Is Linked to Virulence of Mucor circinelloides

PubMed Central

Li, Charles H.; Cervantes, Maria; Springer, Deborah J.; Boekhout, Teun; Ruiz-Vazquez, Rosa M.; Torres-Martinez, Santiago R.; Heitman, Joseph; Lee, Soo Chan

2011-01-01

Mucor circinelloides is a zygomycete fungus and an emerging opportunistic pathogen in immunocompromised patients, especially transplant recipients and in some cases otherwise healthy individuals. We have discovered a novel example of size dimorphism linked to virulence. M. circinelloides is a heterothallic fungus: (+) sex allele encodes SexP and (−) sex allele SexM, both of which are HMG domain protein sex determinants. M. circinelloides f. lusitanicus (Mcl) (−) mating type isolates produce larger asexual sporangiospores that are more virulent in the wax moth host compared to (+) isolates that produce smaller less virulent sporangiospores. The larger sporangiospores germinate inside and lyse macrophages, whereas the smaller sporangiospores do not. sexMΔ mutants are sterile and still produce larger virulent sporangiospores, suggesting that either the sex locus is not involved in virulence/spore size or the sexP allele plays an inhibitory role. Phylogenetic analysis supports that at least three extant subspecies populate the M. circinelloides complex in nature: Mcl, M. circinelloides f. griseocyanus, and M. circinelloides f. circinelloides (Mcc). Mcc was found to be more prevalent among clinical Mucor isolates, and more virulent than Mcl in a diabetic murine model in contrast to the wax moth host. The M. circinelloides sex locus encodes an HMG domain protein (SexP for plus and SexM for minus mating types) flanked by genes encoding triose phosphate transporter (TPT) and RNA helicase homologs. The borders of the sex locus between the three subspecies differ: the Mcg sex locus includes the promoters of both the TPT and the RNA helicase genes, whereas the Mcl and Mcc sex locus includes only the TPT gene promoter. Mating between subspecies was restricted compared to mating within subspecies. These findings demonstrate that spore size dimorphism is linked to virulence of M. circinelloides species and that plasticity of the sex locus and adaptations in pathogenicity have occurred during speciation of the M. circinelloides complex. PMID:21698218

Biodiversity and evolution of the Myxozoa.

PubMed

Canning, Elizabeth U; Okamura, Beth

2004-01-01

Myxozoans (phylum Myxozoa) are metazoan parasites utilizing invertebrate and (mainly) aquatic vertebrate hosts. They have in common with cnidarians the possession of virtually identical, highly complex organelles, namely the polar capsules in myxozoan spores, serving for attachment to new hosts and the nematocysts in surface epithelia of cnidarians, serving for food capture. Although myxozoan spores are multicellular, the simple trophic body forms of almost all species, reduced to syncytial plasmodia or single cells, reveal no clues to myxozoan ancestry or phylogenetic relationships. The myxozoan genus Buddenbrockia is one of only two known genera belonging to a clade which diverged early in the evolution of the Myxozoa. Today the Myxozoa are represented by two classes, the Myxosporea, containing all the better-known genera, which alternate between fish and annelids, and the Malacosporea, containing Buddenbrockia and Tetracapsuloides, parasitising bryozoans. The latter genus also infects salmonid fish, causing proliferative kidney disease (PKD). The enigmatic Buddenbrockia has retained some of its ancestral features in a body wall of two cell layers and a worm-like shape, maintained by four longitudinally-running muscle blocks, similar to a gutless nematode and suggestive of a bilaterian ancestry. Although some analyses of 18S rDNA sequences tend towards a cnidarian (diploblast) affinity for myxozoans, the majority of these studies place them within, or sister to, the Bilateria. The latter view is supported by their possession of central class Hox genes, so far considered to be synapomorphic for Bilateria. The simple body form is, therefore, an extreme example of simplification due to parasitism. Various hypotheses for the occurrence of identical complex organelles (nematocysts and polar capsules) in diploblast and triploblast phyla are evaluated: common ancestry, convergent evolution, gene transfer and, especially, endosymbiosis. A theory of the evolution of their digenetic life cycles is proposed, with the invertebrate as primary host and secondary acquisition of the vertebrate host serving for asexual population increase.

Rgs1 regulates multiple Gα subunits in Magnaporthe pathogenesis, asexual growth and thigmotropism

PubMed Central

Liu, Hao; Suresh, Angayarkanni; Willard, Francis S; Siderovski, David P; Lu, Shen; Naqvi, Naweed I

2007-01-01

Regulators of G-protein signaling (RGS proteins) negatively regulate heterotrimeric G-protein cascades that enable eukaryotic cells to perceive and respond to external stimuli. The rice-blast fungus Magnaporthe grisea forms specialized infection structures called appressoria in response to inductive surface cues. We isolated Magnaporthe RGS1 in a screen for mutants that form precocious appressoria on non-inductive surfaces. We report that a thigmotropic cue is necessary for initiating appressoria and for accumulating cAMP. Similar to an RGS1-deletion strain, magAG187S (RGS-insensitive Gαs) and magAQ208L (GTPase-dead) mutants accumulated excessive cAMP and elaborated appressoria on non-inductive surfaces, suggesting that Rgs1 regulates MagA during pathogenesis. Rgs1 was also found to negatively regulate the Gαi subunit MagB during asexual development. Deficiency of MAGB suppressed the hyper-conidiation defect in RGS1-deletion strain, whereas magBG183S and magBQ204L mutants produced more conidia, similar to the RGS1-deletion strain. Rgs1 physically interacted with GDP·AlF4−-activated forms of MagA, MagB and MagC (a GαII subunit). Thus, Rgs1 serves as a negative regulator of all Gα subunits in Magnaporthe and controls important developmental events during asexual and pathogenic development. PMID:17255942

Bone Marrow Is a Major Parasite Reservoir in Plasmodium vivax Infection

PubMed Central

2018-01-01

ABSTRACT Plasmodium vivax causes heavy burdens of disease across malarious regions worldwide. Mature P. vivax asexual and transmissive gametocyte stages occur in the blood circulation, and it is often assumed that accumulation/sequestration in tissues is not an important phase in their development. Here, we present a systematic study of P. vivax stage distributions in infected tissues of nonhuman primate (NHP) malaria models as well as in blood from human infections. In a comparative analysis of the transcriptomes of P. vivax and Plasmodium falciparum blood-stage parasites, we found a conserved cascade of stage-specific gene expression despite the greatly different gametocyte maturity times of these two species. Using this knowledge, we validated a set of conserved asexual- and gametocyte-stage markers both by quantitative real-time PCR and by antibody assays of peripheral blood samples from infected patients and NHP (Aotus sp.). Histological analyses of P. vivax parasites in organs of 13 infected NHP (Aotus and Saimiri species) demonstrated a major fraction of immature gametocytes in the parenchyma of the bone marrow, while asexual schizont forms were enriched to a somewhat lesser extent in this region of the bone marrow as well as in sinusoids of the liver. These findings suggest that the bone marrow is an important reservoir for gametocyte development and proliferation of malaria parasites. PMID:29739900

Effects of culture conditions on the size, morphology and wet density of spores of Bacillus cereus 569 and Bacillus megaterium QM B1551.

PubMed

Xu Zhou, K; Wisnivesky, F; Wilson, D I; Christie, G

2017-07-01

The influence of variable culture conditions on the size and wet density of spores of Bacillus cereus and Bacillus megaterium were examined in this work. Culture temperature and initial pH was shown to have a significant impact on the size of both species, with increasingly alkaline culture media and elevated culture temperatures resulting in spores that were, on average, up to 25% reduced in volume. Increasing concentrations of inorganic salts in sporulation media exerted differing effects on each species; whereas a fivefold increase in the concentration of all salts resulted in only minor differences to the dimensions of B. cereus spores, B. megaterium spores became more elongated, displaying an average increase in volume of almost 30%. Similarly, as the spore elongated to yield aspect ratios larger than 1·4, their shape changed from typical prolate spheroids to cylinders with hemispherical ends. In contrast with previous studies, culture conditions employed in this study exerted no discernible impact on the wet density of B. cereus or B. megaterium spores. Bacterial spores of the genera Bacillus and Clostridium represent nature's most durable cells in terms of their extreme resistance to a variety of deleterious environments. As a result, they are of concern in the food processing, healthcare and other sectors, and are of increasing biotechnological interest. Improved understanding of variance in spore size, morphology and density may aid the development of certain spore-associated applications (e.g. spore surface display) while contributing to active areas of research such as spore adhesion and resistance to heat. © 2017 The Authors. Letters in Applied Microbiology published by John Wiley & Sons Ltd on behalf of Society for Applied Microbiology.

Evolutionary clade affects resistance of Clostridium difficile spores to Cold Atmospheric Plasma

NASA Astrophysics Data System (ADS)

Connor, Mairéad; Flynn, Padrig B.; Fairley, Derek J.; Marks, Nikki; Manesiotis, Panagiotis; Graham, William G.; Gilmore, Brendan F.; McGrath, John W.

2017-02-01

Clostridium difficile is a spore forming bacterium and the leading cause of colitis and antibiotic associated diarrhoea in the developed world. Spores produced by C. difficile are robust and can remain viable for months, leading to prolonged healthcare-associated outbreaks with high mortality. Exposure of C. difficile spores to a novel, non-thermal atmospheric pressure gas plasma was assessed. Factors affecting sporicidal efficacy, including percentage of oxygen in the helium carrier gas admixture, and the effect on spores from different strains representing the five evolutionary C. difficile clades was investigated. Strains from different clades displayed varying resistance to cold plasma. Strain R20291, representing the globally epidemic ribotype 027 type, was the most resistant. However all tested strains displayed a ~3 log reduction in viable spore counts after plasma treatment for 5 minutes. Inactivation of a ribotype 078 strain, the most prevalent clinical type seen in Northern Ireland, was further assessed with respect to surface decontamination, pH, and hydrogen peroxide concentration. Environmental factors affected plasma activity, with dry spores without the presence of organic matter being most susceptible. This study demonstrates that cold atmospheric plasma can effectively inactivate C. difficile spores, and highlights factors that can affect sporicidal activity.

Inactivation of Aerosolized Biological Agents using Filled Nanocomposite Materials

DTIC Science & Technology

2013-02-01

developed and optimized. The dry -heat inactivation of aerosolized spores was quantified separately from chemical effects and linked to DNA repair...Bacillus spores exposed to dry heat 67 - 79 Chapter 5. Mechanically alloyed Al-I composite materials 80 - 98 Chapter 6. Iodine release...and optimized. The dry -heat inactivation of aerosolized spores was quantified separately from chemical effects and linked to DNA repair mechanisms

Development of Metarhizium anisopliae and Beauveria bassiana formulations for control of malaria mosquito larvae

PubMed Central

2011-01-01

Background The entomopathogenic fungi Metarhizium anisopliae and Beauveria bassiana have demonstrated effectiveness against anopheline larvae in the laboratory. However, utilising these fungi for the control of anopheline larvae under field conditions, relies on development of effective means of application as well as reducing their sensitivity to UV radiation, high temperatures and the inevitable contact with water. This study was conducted to develop formulations that facilitate the application of Metarhizium anisopliae and Beauveria bassiana spores for the control of anopheline larvae, and also improve their persistence under field conditions. Methods Laboratory bioassays were conducted to test the ability of aqueous (0.1% Tween 80), dry (organic and inorganic) and oil (mineral and synthetic) formulations to facilitate the spread of fungal spores over the water surface and improve the efficacy of formulated spores against anopheline larvae as well as improve spore survival after application. Field bioassays were then carried out to test the efficacy of the most promising formulation under field conditions in western Kenya. Results When formulated in a synthetic oil (ShellSol T), fungal spores of both Metarhizium anisopliae and Beauveria bassiana were easy to mix and apply to the water surface. This formulation was more effective against anopheline larvae than 0.1% Tween 80, dry powders or mineral oil formulations. ShellSol T also improved the persistence of fungal spores after application to the water. Under field conditions in Kenya, the percentage pupation of An. gambiae was significantly reduced by 39 - 50% by the ShellSol T-formulated Metarhizium anisopliae and Beauveria bassiana spores as compared to the effects of the application of unformulated spores. Conclusions ShellSol T is an effective carrier for fungal spores when targeting anopheline larvae under both laboratory and field conditions. Entomopathogenic fungi formulated with a suitable carrier are a promising tool for control of larval populations of malaria mosquitoes. Additional studies are required to identify the best delivery method (where, when and how) to make use of the entomopathogenic potential of these fungi against anopheline larvae. PMID:21342492

One-step purification of Enterocytozoon bieneusi spores from human stools by immunoaffinity expanded-bed adsorption.

PubMed

Accoceberry, I; Thellier, M; Datry, A; Desportes-Livage, I; Biligui, S; Danis, M; Santarelli, X

2001-05-01

An original, reliable, and reproducible method for the purification of Enterocytozoon bieneusi spores from human stools is described. We recently reported the production of a species-specific monoclonal antibody (MAb) 6E52D9 immunoglobulin G2a (IgG2a) raised against the exospore of E. bieneusi spore walls. The MAb was used as a ligand to develop an immunoaffinity matrix. The mouse IgG2a MAb was bound directly to a Streamline rProtein A adsorbent, used for expanded-bed adsorption of immunoglobulins, for optimal spatial orientation of the antibody and maximum binding efficiency of the antigen. The complex was then cross-linked covalently using dimethyl pimelimidate dihydrochloride. After incubation of the immunoaffinity matrix with filtered stool samples containing numerous E. bieneusi spores and before elution with 6 M guanidine HCl, the expansion of the adsorbent bed eliminated all the fecal contaminants. The presence of spores in the elution fractions was determined by an indirect immunofluorescence antibody test (IFAT). E. bieneusi spores were found in the elution fraction in all four experiments and were still highly antigenic as indicated by IFAT. Smears examined by light microscopy contained very clean spores with no fecal debris or background bacterial and fungal contaminants. However, spore recovery rates were relatively low: an average of 10(7) spores were purified per run. This technique for isolating E. bieneusi spores directly from human stool samples with a high degree of purity opens up new approaches for studying this parasite.

Artemisone and artemiside - potent pan-reactive antimalarial agents that also synergize redox imbalance in P. falciparum transmissible gametocyte stages.

PubMed

Coertzen, Dina; Reader, Janette; van der Watt, Mariëtte; Nondaba, Sindisiwe H; Gibhard, Liezl; Wiesner, Lubbe; Smith, Peter; D'Alessandro, Sarah; Taramelli, Donatella; Ning Wong, Ho; du Preez, Jan L; Wu, Ronald Wai Keung; Birkholtz, Lyn-Marie; Haynes, Richard K

2018-06-04

The emergence of resistance towards artemisinin combination therapies (ACTs) by the malaria parasite Plasmodium falciparum has the potential to severely compromise malaria control. Therefore, development of new artemisinins in combination with new drugs that impart activities towards both intraerythrocytic proliferative asexual and transmissible gametocyte stages, in particular those of resistant parasites, are urgently required. We define artemisinins as oxidant drugs through their ability to oxidize reduced flavin cofactors of flavin disulfide reductases critical for maintaining redox-homeostasis in the malaria parasite. Here we compare the activities of 10-amino artemisinin derivatives towards the asexual and gametocyte stages of P. falciparum parasites. Of these, artemisone and artemiside inhibited asexual and gametocyte stages, particularly stage V gametocytes in the low nM range. Further, treatment of both early and late gametocyte stages with artemisone or artemiside combined with the pro-oxidant redox partner methylene blue displays notable synergism. These data suggest that modulation of redox-homeostasis likely is an important druggable process, particularly in gametocytes, and thereby enhances the prospect of using combinations of oxidant and redox drugs for malaria control. Copyright © 2018 American Society for Microbiology.

CRISPR-Cas9-modified pfmdr1 protects Plasmodium falciparum asexual blood stages and gametocytes against a class of piperazine-containing compounds but potentiates artemisinin-based combination therapy partner drugs.

PubMed

Ng, Caroline L; Siciliano, Giulia; Lee, Marcus C S; de Almeida, Mariana J; Corey, Victoria C; Bopp, Selina E; Bertuccini, Lucia; Wittlin, Sergio; Kasdin, Rachel G; Le Bihan, Amélie; Clozel, Martine; Winzeler, Elizabeth A; Alano, Pietro; Fidock, David A

2016-08-01

Emerging resistance to first-line antimalarial combination therapies threatens malaria treatment and the global elimination campaign. Improved therapeutic strategies are required to protect existing drugs and enhance treatment efficacy. We report that the piperazine-containing compound ACT-451840 exhibits single-digit nanomolar inhibition of the Plasmodium falciparum asexual blood stages and transmissible gametocyte forms. Genome sequence analyses of in vitro-derived ACT-451840-resistant parasites revealed single nucleotide polymorphisms in pfmdr1, which encodes a digestive vacuole membrane-bound ATP-binding cassette transporter known to alter P. falciparum susceptibility to multiple first-line antimalarials. CRISPR-Cas9 based gene editing confirmed that PfMDR1 point mutations mediated ACT-451840 resistance. Resistant parasites demonstrated increased susceptibility to the clinical drugs lumefantrine, mefloquine, quinine and amodiaquine. Stage V gametocytes harboring Cas9-introduced pfmdr1 mutations also acquired ACT-451840 resistance. These findings reveal that PfMDR1 mutations can impart resistance to compounds active against asexual blood stages and mature gametocytes. Exploiting PfMDR1 resistance mechanisms provides new opportunities for developing disease-relieving and transmission-blocking antimalarials. © 2016 John Wiley & Sons Ltd.

Cell-Type–Specific Transcriptional Profiles of the Dimorphic Pathogen Penicillium marneffei Reflect Distinct Reproductive, Morphological, and Environmental Demands

PubMed Central

Pasricha, Shivani; Payne, Michael; Canovas, David; Pase, Luke; Ngaosuwankul, Nathamon; Beard, Sally; Oshlack, Alicia; Smyth, Gordon K.; Chaiyaroj, Sansanee C.; Boyce, Kylie J.; Andrianopoulos, Alex

2013-01-01

Penicillium marneffei is an opportunistic human pathogen endemic to Southeast Asia. At 25° P. marneffei grows in a filamentous hyphal form and can undergo asexual development (conidiation) to produce spores (conidia), the infectious agent. At 37° P. marneffei grows in the pathogenic yeast cell form that replicates by fission. Switching between these growth forms, known as dimorphic switching, is dependent on temperature. To understand the process of dimorphic switching and the physiological capacity of the different cell types, two microarray-based profiling experiments covering approximately 42% of the genome were performed. The first experiment compared cells from the hyphal, yeast, and conidiation phases to identify “phase or cell-state–specific” gene expression. The second experiment examined gene expression during the dimorphic switch from one morphological state to another. The data identified a variety of differentially expressed genes that have been organized into metabolic clusters based on predicted function and expression patterns. In particular, C-14 sterol reductase–encoding gene ergM of the ergosterol biosynthesis pathway showed high-level expression throughout yeast morphogenesis compared to hyphal. Deletion of ergM resulted in severe growth defects with increased sensitivity to azole-type antifungal agents but not amphotericin B. The data defined gene classes based on spatio-temporal expression such as those expressed early in the dimorphic switch but not in the terminal cell types and those expressed late. Such classifications have been helpful in linking a given gene of interest to its expression pattern throughout the P. marneffei dimorphic life cycle and its likely role in pathogenicity. PMID:24062530

The immunological characteristics and probiotic function of recombinant Bacillus subtilis spore expressing Clonorchis sinensis cysteine protease.

PubMed

Tang, Zeli; Shang, Mei; Chen, Tingjin; Ren, Pengli; Sun, Hengchang; Qu, Hongling; Lin, Zhipeng; Zhou, Lina; Yu, Jinyun; Jiang, Hongye; Zhou, Xinyi; Li, Xuerong; Huang, Yan; Xu, Jin; Yu, Xinbing

2016-12-19

Clonorchiasis, a food-borne zoonosis, is caused by Clonorchis sinensis. The intestinal tract and bile ducts are crucial places for C. sinensis metacercariae to develop into adult worms. The endospore of Bacillus subtilis is an ideal oral immunization vehicle for delivery of heterologous antigens to intestine. Cysteine protease of C. sinensis (CsCP) is an endogenous key component in the excystment of metacercariae and other physiological or pathological processes. We constructed a fusion gene of CotC (a coat protein)-CsCP and obtained B. subtilis spores with recombinant plasmid of pEB03-CotC-CsCP (B.s-CotC-CsCP). CotC-CsCP expressed on spores' surface was detected by Western blotting and immunofluorescence. Immunological characteristics of recombinant spore coat protein were evaluated in a mouse model. The levels of CsCP-specific antibodies were detected by ELISA. Effects of recombinant spores on mouse intestine were evaluated by histological staining. The activities of biochemical enzymes in serum were assayed by microplate. Liver sections of infected mice were evaluated by Ishak score after Masson's trichrome. The B.s-CotC-CsCP spores displayed CsCP on their coat. Specific IgG and isotypes were significantly induced by coat proteins of B.s-CotC-CsCP spores after subcutaneous immunization. IgA levels in intestinal mucus and bile of B.s-CotC-CsCP orally treated mice significantly increased. Additionally, more IgA-secreting cells were observed in enteraden and lamina propria regions of the mouse jejunum, and an increased amount of acidic mucins in intestines were also observed. There were no significant differences in enzyme levels of serum among groups. No inflammatory injury was observed in the intestinal tissues of each group. The degree of liver fibrosis was significantly reduced after oral immunization with B.s-CotC-CsCP spores. Bacillus subtilis spores maintained the original excellent immunogenicity of CsCP expressed on their surface. Both local and systemic specific immune responses were elicited by oral administration of B.s-CotC-CsCP spores. The spores effectively promoted intestinal health by inducing secretion of acidic mucins, with no other side effects to the liver or intestine. Oral administration of spores expressing CsCP could provide effective protection against C. sinensis. This study may be a cornerstone for development of antiparasitic agents or vaccines against clonorchiasis based on B. subtilis spore expressing CsCP on the surface.

Life History and Developmental Processes in the Basidiomycete Coprinus cinereus

PubMed Central

Kües, Ursula

2000-01-01

Coprinus cinereus has two main types of mycelia, the asexual monokaryon and the sexual dikaryon, formed by fusion of compatible monokaryons. Syngamy (plasmogamy) and karyogamy are spatially and temporally separated, which is typical for basidiomycetous fungi. This property of the dikaryon enables an easy exchange of nuclear partners in further dikaryotic-monokaryotic and dikaryotic-dikaryotic mycelial fusions. Fruiting bodies normally develop on the dikaryon, and the cytological process of fruiting-body development has been described in its principles. Within the specialized basidia, present within the gills of the fruiting bodies, karyogamy occurs in a synchronized manner. It is directly followed by meiosis and by the production of the meiotic basidiospores. The synchrony of karyogamy and meiosis has made the fungus a classical object to study meiotic cytology and recombination. Several genes involved in these processes have been identified. Both monokaryons and dikaryons can form multicellular resting bodies (sclerotia) and different types of mitotic spores, the small uninucleate aerial oidia, and, within submerged mycelium, the large thick-walled chlamydospores. The decision about whether a structure will be formed is made on the basis of environmental signals (light, temperature, humidity, and nutrients). Of the intrinsic factors that control development, the products of the two mating type loci are most important. Mutant complementation and PCR approaches identified further genes which possibly link the two mating-type pathways with each other and with nutritional regulation, for example with the cAMP signaling pathway. Among genes specifically expressed within the fruiting body are those for two galectins, β-galactoside binding lectins that probably act in hyphal aggregation. These genes serve as molecular markers to study development in wild-type and mutant strains. The isolation of genes for potential non-DNA methyltransferases, needed for tissue formation within the fruiting body, promises the discovery of new signaling pathways, possibly involving secondary fungal metabolites. PMID:10839819

Gβ-Like CpcB Plays a Crucial Role for Growth and Development of Aspergillus nidulans and Aspergillus fumigatus

PubMed Central

Kong, Qing; Wang, Long; Liu, Zengran; Kwon, Nak-Jung; Kim, Sun Chang; Yu, Jae-Hyuk

2013-01-01

Growth, development, virulence and secondary metabolism in fungi are governed by heterotrimeric G proteins (G proteins). A Gβ-like protein called Gib2 has been shown to function as an atypical Gβ in Gpa1-cAMP signaling in Cryptococcus neoformans. We found that the previously reported CpcB (cross pathway control B) protein is the ortholog of Gib2 in Aspergillus nidulans and Aspergillus fumigatus. In this report, we further characterize the roles of CpcB in governing growth, development and toxigenesis in the two aspergilli. The deletion of cpcB results in severely impaired cellular growth, delayed spore germination, and defective asexual sporulation (conidiation) in both aspergilli. Moreover, CpcB is necessary for proper expression of the key developmental activator brlA during initiation and progression of conidiation in A. nidulans and A. fumigatus. Somewhat in accordance with the previous study, the absence of cpcB results in the formation of fewer, but not micro-, cleistothecia in A. nidulans in the presence of wild type veA, an essential activator of sexual development. However, the cpcB deletion mutant cleistothecia contain no ascospores, validating that CpcB is required for progression and completion of sexual fruiting including ascosporogenesis. Furthermore, unlike the canonical GβSfaD, CpcB is not needed for the biosynthesis of the mycotoxin sterigmatocystin (ST) as the cpcB null mutant produced reduced amount of ST with unaltered STC gene expression. However, in A. fumigatus, the deletion of cpcB results in the blockage of gliotoxin (GT) production. Further genetic analyses in A. nidulans indicate that CpcB may play a central role in vegetative growth, which might be independent of FadA- and GanB-mediated signaling. A speculative model summarizing the roles of CpcB in conjunction with SfaD in A. nidulans is presented. PMID:23936193

Gβ-like CpcB plays a crucial role for growth and development of Aspergillus nidulans and Aspergillus fumigatus.

PubMed

Kong, Qing; Wang, Long; Liu, Zengran; Kwon, Nak-Jung; Kim, Sun Chang; Yu, Jae-Hyuk

2013-01-01

Growth, development, virulence and secondary metabolism in fungi are governed by heterotrimeric G proteins (G proteins). A Gβ-like protein called Gib2 has been shown to function as an atypical Gβ in Gpa1-cAMP signaling in Cryptococcus neoformans. We found that the previously reported CpcB (cross pathway control B) protein is the ortholog of Gib2 in Aspergillus nidulans and Aspergillus fumigatus. In this report, we further characterize the roles of CpcB in governing growth, development and toxigenesis in the two aspergilli. The deletion of cpcB results in severely impaired cellular growth, delayed spore germination, and defective asexual sporulation (conidiation) in both aspergilli. Moreover, CpcB is necessary for proper expression of the key developmental activator brlA during initiation and progression of conidiation in A. nidulans and A. fumigatus. Somewhat in accordance with the previous study, the absence of cpcB results in the formation of fewer, but not micro-, cleistothecia in A. nidulans in the presence of wild type veA, an essential activator of sexual development. However, the cpcB deletion mutant cleistothecia contain no ascospores, validating that CpcB is required for progression and completion of sexual fruiting including ascosporogenesis. Furthermore, unlike the canonical GβSfaD, CpcB is not needed for the biosynthesis of the mycotoxin sterigmatocystin (ST) as the cpcB null mutant produced reduced amount of ST with unaltered STC gene expression. However, in A. fumigatus, the deletion of cpcB results in the blockage of gliotoxin (GT) production. Further genetic analyses in A. nidulans indicate that CpcB may play a central role in vegetative growth, which might be independent of FadA- and GanB-mediated signaling. A speculative model summarizing the roles of CpcB in conjunction with SfaD in A. nidulans is presented.

Development of a user-friendly delivery method for the fungus Metarhizium anisopliae to control the ectoparasitic mite Varroa destructor in honey bee, Apis mellifera, colonies.

PubMed

Kanga, Lambert H B; Adamczyk, John; Patt, Joseph; Gracia, Carlos; Cascino, John

2010-12-01

A user-friendly method to deliver Metarhizium spores to honey bee colonies for control of Varroa mites was developed and tested. Patty blend formulations protected the fungal spores at brood nest temperatures and served as an improved delivery system of the fungus to bee hives. Field trials conducted in 2006 in Texas using freshly harvested spores indicated that patty blend formulations of 10 g of conidia per hive (applied twice) significantly reduced the numbers of mites per adult bee, mites in sealed brood cells, and residual mites at the end of the 47-day experimental period. Colony development in terms of adult bee populations and brood production also improved. Field trials conducted in 2007 in Florida using less virulent spores produced mixed results. Patty blends of 10 g of conidia per hive (applied twice) were less successful in significantly reducing the number of mites per adult bee. However, hive survivorship and colony strength were improved, and the numbers of residual mites were significantly reduced at the end of the 42-day experimental period. The overall results from 2003 to 2008 field trials indicated that it was critical to have fungal spores with good germination, pathogenicity and virulence. We determined that fungal spores (1 × 10(10) viable spores per gram) with 98% germination and high pathogenicity (95% mite mortality at day 7) provided successful control of mite populations in established honey bee colonies at 10 g of conidia per hive (applied twice). Overall, microbial control of Varroa mite with M. anisopliae is feasible and could be a useful component of an integrated pest management program.

Surface Bacterial-Spore Assay Using Tb3+/DPA Luminescence

NASA Technical Reports Server (NTRS)

Ponce, Adrian

2007-01-01

Equipment and a method for rapidly assaying solid surfaces for contamination by bacterial spores are undergoing development. The method would yield a total (nonviable plus viable) spore count of a surface within minutes and a viable-spore count in about one hour. In this method, spores would be collected from a surface by use of a transparent polymeric tape coated on one side with a polymeric adhesive that would be permeated with one or more reagent(s) for detection of spores by use of visible luminescence. The sticky side of the tape would be pressed against a surface to be assayed, then the tape with captured spores would be placed in a reader that illuminates the sample with ultraviolet light and counts the green luminescence spots under a microscope to quantify the number of bacterial spores per unit area. The visible luminescence spots seen through the microscope would be counted to determine the concentration of spores on the surface. This method is based on the chemical and physical principles of methods described in several prior NASA Tech Briefs articles, including Live/Dead Spore Assay Using DPA-Triggered Tb Luminescence (NPO-30444), Vol. 27, No. 3 (March 2003), page 7a. To recapitulate: The basic idea is to exploit the observations that (1) dipicolinic acid (DPA) is present naturally only in bacterial spores; and (2) when bound to Tb3+ ions, DPA triggers intense green luminescence of the ions under ultraviolet excitation; (3) DPA can be released from the viable spores by using L-alanine to make them germinate; and (4) by autoclaving, microwaving, or sonicating the sample, one can cause all the spores (non-viable as well as viable) to release their DPA. One candidate material for use as the adhesive in the present method is polydimethysiloxane (PDMS). In one variant of the method for obtaining counts of all (viable and nonviable) spores the PDMS would be doped with TbCl3. After collection of a sample, the spores immobilized on the sticky tape surface would be lysed by heating or microwaving to release their DPA. Tb3+ ions from the TbCl3 would become bound to the released DPA. The tape would then be irradiated with ultraviolet and examined as described above. In another variant of the method - for obtaining counts of viable spores only - the PDMS would be doped with L-alanine in addition to TbCl3. As now envisioned, a fully developed apparatus for implementing this method would include a pulsed source of ultraviolet light and a time-gated electronic camera to record the images seen through the microscope during a prescribed exposure interval at a prescribed short time after an ultraviolet pulse. As in the method of the second-mentioned prior article, the pulsing and time-gating would be used to discriminate between the longer-lived Tb3+/DPA luminescence and the shorter-lived background luminescence in the same wavelength range. In a time-gated image, the bright luminescence from bacterial spores could easily be seen against a dark background.

Self-Organized Criticality in an Asexual Model?

NASA Astrophysics Data System (ADS)

Chisholm, Colin; Jan, Naeem; Gibbs, Peter; Erzan, Ayşe.

Recent work has shown that the distribution of steady state mutations for an asexual ``bacteria'' model has features similar to that seen in Self-Organized Critical (SOC) sandpile model of Bak et al. We investigate this coincidence further and search for ``self-organized critical'' state for bacteria but instead find that the SOC sandpile critical behavior is very sensitive; critical behavior is destroyed with small perturbations effectively when the absorption of sand is introduced. It is only in the limit when the length of the genome of the bacteria tends to infinity that SOC properties are recovered for the asexual model.

Sex and the Imperfect Fungi.

PubMed

Dyer, Paul S; Kück, Ulrich

2017-06-01

Approximately 20% of species in the fungal kingdom are only known to reproduce by asexual means despite the many supposed advantages of sexual reproduction. However, in recent years, sexual cycles have been induced in a series of emblematic "asexual" species. We describe how these discoveries were made, building on observations of evidence for sexual potential or "cryptic sexuality" from population genetic analyses; the presence, distribution, and functionality of mating-type genes; genome analyses revealing the presence of genes linked to sexuality; the functionality of sex-related genes; and formation of sex-related developmental structures. We then describe specific studies that led to the discovery of mating and sex in certain Candida , Aspergillus , Penicillium , and Trichoderma species and discuss the implications of sex including the beneficial exploitation of the sexual cycle. We next consider whether there might be any truly asexual fungal species. We suggest that, although rare, imperfect fungi may genuinely be present in nature and that certain human activities, combined with the genetic flexibility that is a hallmark of the fungal kingdom, might favor the evolution of asexuality under certain conditions. Finally, we argue that fungal species should not be thought of as simply asexual or sexual, but rather as being composed of isolates on a continuum of sexual fertility.

Spermatozoa Production by Triploid Males in the New Zealand Freshwater Snail Potamopyrgus antipodarum

PubMed Central

Soper, D.M.; Neiman, M.; Savytskyy, O.P.; Zolan, M.E.; Lively, C.M.

2013-01-01

Asexual lineages derived from dioecious taxa are typically assumed to be all female. Even so, asexual females from a variety of animal taxa occasionally produce males. The existence of these males sets the stage for potential gene flow across asexual lineages as well as between sexual and asexual lineages. A recent study showed that asexual triploid female Potamopyrgus antipodarum, a New Zealand freshwater snail often used as a model to study sexual reproduction, occasionally produce triploid male offspring. Here, we show that these triploid male P. antipodarum 1) have testes that produce morphologically normal sperm, 2) make larger sperm cells that contain more nuclear DNA than the sperm produced by diploid sexual males, and 3) produce sperm that range in DNA content from haploid to diploid, and are often aneuploid. Analysis of meiotic chromosomes of triploid males showed that aberrant pairing during prophase I likely accounts for the high variation in DNA content among sperm. These results indicate that triploid male P. antipodarum produce sperm, but the extent to which these sperm are able to fertilize female ova remains unclear. Our results also suggest that the general assumption of sterility in triploid males should be more closely examined in other species in which such males are occasionally produced. PMID:24307744

Characterization of 21 microsatellite markers from cogongrass, Imperata cylindrica (Poaceae), a weed species distributed worldwide.

PubMed

Chiang, Yu-Chung; Tsai, Chi-Chu; Hsu, Tsai-Wen; Chou, Chang-Hung

2012-11-01

Microsatellite loci were developed from Imperata cylindrica, a traditional medicinal herb in Asia and among the top 10 worst invasive weeds in the world, to aid in the identification of the limits of asexual clonal individuals. A total of 21 microsatellite markers, including 18 polymorphic and three monomorphic loci, were developed from I. cylindrica using a magnetic bead enrichment protocol. The primers amplified dinucleotide, trinucleotide, and complex repeats. The number of alleles ranged from one to 19 per locus, with an observed heterozygosity ranging from 0.09 to 1.00. Several loci deviated significantly from the within-population Hardy-Weinberg equilibrium as a result of asexual clonal reproduction. These polymorphic markers should be useful tools in further studies on the identification of the range of clonal reproduction units and the selection and classification of the medicinal cultivar.

Mannosyltransferase is required for cell wall biosynthesis, morphology and control of asexual development in Neurospora crassa.

PubMed

Bowman, Shaun M; Piwowar, Amy; Ciocca, Maria; Free, Stephen J

2005-01-01

Two Neurospora mutants with a phenotype that includes a tight colonial growth pattern, an inability to form conidia and an inability to form protoperithecia have been isolated and characterized. The relevant mutations were mapped to the same locus on the sequenced Neurospora genome. The mutations responsible for the mutant phenotype then were identified by examining likely candidate genes from the mutant genomes at the mapped locus with PCR amplification and a sequencing assay. The results demonstrate that a map and sequence strategy is a feasible way to identify mutant genes in Neurospora. The gene responsible for the phenotype is a putative alpha-1,2-mannosyltransferase gene. The mutant cell wall has an altered composition demonstrating that the gene functions in cell wall biosynthesis. The results demonstrate that the mnt-1 gene is required for normal cell wall biosynthesis, morphology and for the regulation of asexual development.

Small-molecule xenomycins inhibit all stages of the Plasmodium life cycle.

PubMed

Erath, Jessey; Gallego-Delgado, Julio; Xu, Wenyue; Andriani, Grasiella; Tanghe, Scott; Gurova, Katerina V; Gudkov, Andrei; Purmal, Andrei; Rydkina, Elena; Rodriguez, Ana

2015-03-01

Widespread resistance to most antimalaria drugs in use has prompted the search for novel candidate compounds with activity against Plasmodium asexual blood stages to be developed for treatment. In addition, the current malaria eradication programs require the development of drugs that are effective against all stages of the parasite life cycle. We have analyzed the antimalarial properties of xenomycins, a novel subclass of small molecule compounds initially isolated for anticancer activity and similarity to quinacrine in biological effects on mammalian cells. In vitro studies show potent activity of Xenomycins against Plasmodium falciparum. Oral administration of xenomycins in mouse models result in effective clearance of liver and blood asexual and sexual stages, as well as effective inhibition of transmission to mosquitoes. These characteristics position xenomycins as antimalarial candidates with potential activity in prevention, treatment and elimination of this disease. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

A method of increasing test range and accuracy of bioindicators: Geobacillus stearothermophilus spores.

PubMed

Lundahl, Gunnel

2003-01-01

Spores of Geobacillus stearothermophilus are very sensitive to changes in temperature. When validating sterilizing processes, the most common bioindicator (BI) is spores of Geobacillus stearothermophilus ATCC12980 and ATCC7953 with about 10(6) spores /BI and a D121-value of about 2 minutes in water. Because these spores of Geobacillus stearothermophilus do not survive at a F0-value above 12 minutes, it has not been possible to evaluate the agreement between the biological F-value (F(BIO)) and physical measurements (time and temperature) when the physical F0-value exceeds that limit. However, it has been proven that glycerin substantially increases the heat resistance of the spores, and it is possible to utilize that property when manufacturing BIs suitable to use in processes with longer sterilization time or high temperature (above 121 degrees C). By the method described, it is possible to make use of the sensitivity and durability of Geobacillus stearothermophilus' spores when glycerin has increased both test range and accuracy. Experience from years of development and validation work with the use of the highly sensitive glycerin-water-spore-suspension sensor (GWS-sensor) is reported. Validation of the steam sterilization process at high temperature has been possible with the use of GWS-sensors. It has also been shown that the spores in suspension keep their characteristics for a period of 19 months when stored cold (8 degrees C).

Development and Use of Fluorescent Antibody and qPCR Protocols for the Electrostatic Spore Trap

USDA-ARS?s Scientific Manuscript database

Fluorescent antibody (FA) and qPCR protocols were evaluated for the newly developed aerobiological sampler (Ionic Spore Trap), which depends upon electrostatic deposition of particulates onto a 25 mm aluminum disk (stub). This device was originally designed for assessment of captured particulates by...

Analysis of stage-specific protein expression during babesia bovis development within female rhipicephalus microplus

USDA-ARS?s Scientific Manuscript database

Arthropod borne pathogens have a complex life cycle that includes asexual reproduction of haploid stages in mammalian erythrocytes and development of diploid stages in the vector. Transition of Apicomplexan pathogens between the mammalian host and the arthropod vector is critical for ongoing transmi...

RNA-seq analysis of an nsdC mutant in Aspergillus flavus

USDA-ARS?s Scientific Manuscript database

The C2H2-type transcription factor NsdC (Never in Sexual Development C) has been shown to play a role in asexual development and secondary metabolite production in Aspergillus flavus, an agriculturally relevant, aflatoxin-producing species. The nsdC knoackout mutant demonstrates perturbed morphologi...

An effective iodide formulation for killing Bacillus and Geobacillus spores over a wide temperature range.

PubMed

Kida, N; Mochizuki, Y; Taguchi, F

2004-01-01

To develop a sporicidal reagent which shows potent activity against bacterial spores not only at ambient temperatures but also at low temperatures. Suspension tests on spores of Bacillus and Geobacillus were conducted with the reagent based on a previously reported agent (N. Kida, Y. Mochizuki and F. Taguchi, Microbiology and Immunology 2003; 47: 279-283). The modified reagent (tentatively designated as the KMT reagent) was composed of 50 mmol l(-1) EDTA-2Na, 50 mmol l(-1) ferric chloride hexahydrate (FeCl(3).6H(2)O), 50 mmol l(-1) potassium iodide (KI) and 50% ethanol in 0.85% NaCl solution at pH 0.3. The KMT reagent showed significant sporicidal activity against three species of Bacillus and Geobacillus spores over a wide range of temperature. The KMT reagent had many practical advantages, i.e. activity was much less affected by organic substances than was sodium hypochlorite, it did not generate any harmful gas and it was stable for a long period at ambient temperatures. The mechanism(s) of sporicidal activity of the KMT reagent was considered to be based on active iodine species penetrating the spores with enhanced permeability of the spore cortex by a synergistic effect of acid, ethanol and generated active oxygen. The data suggest that the KMT reagent shows potent sporicidal activity over a wide range temperatures and possesses many advantages for practical applications. The results indicate development of a highly applicable sporicidal reagent against Bacillus and Geobacillus spores.

Fluorimetric Detection of a Bacillus stearothermophilus Spore-Bound Enzyme, α-d-Glucosidase, for Rapid Indication of Flash Sterilization Failure

PubMed Central

Vesley, Donald; Langholz, Ann C.; Rohlfing, Stephen R.; Foltz, William E.

1992-01-01

A biological indicator based on fluorimetric detection within 60 min of a Bacillus stearothermophilus spore-bound enzyme, α-d-glucosidase, has been developed. Results indicate that the enzyme survived slightly longer than spores observed after 24 h of incubation. The new system shows promise for evaluating flash sterilization cycles within 60 min compared with conventional 24-h systems. PMID:16348654

Test methods and response surface models for hot, humid air decontamination of materials contaminated with dirty spores of Bacillus anthracis ∆Sterne and Bacillus thuringiensis Al Hakam.

PubMed

Buhr, T L; Young, A A; Barnette, H K; Minter, Z A; Kennihan, N L; Johnson, C A; Bohmke, M D; DePaola, M; Cora-Laó, M; Page, M A

2015-11-01

To develop test methods and evaluate survival of Bacillus anthracis ∆Sterne or Bacillus thuringiensis Al Hakam on materials contaminated with dirty spore preparations after exposure to hot, humid air using response surface modelling. Spores (>7 log10 ) were mixed with humic acid + spent sporulation medium (organic debris) or kaolin (dirt debris). Spore samples were then dried on five different test materials (wiring insulation, aircraft performance coating, anti-skid, polypropylene, and nylon). Inoculated materials were tested with 19 test combinations of temperature (55, 65, 75°C), relative humidity (70, 80, 90%) and time (1, 2, 3 days). The slowest spore inactivation kinetics was on nylon webbing and/or after addition of organic debris. Hot, humid air effectively decontaminates materials contaminated with dirty Bacillus spore preparations; debris and material interactions create complex decontamination kinetic patterns; and B. thuringiensis Al Hakam is a realistic surrogate for B. anthracis. Response surface models of hot, humid air decontamination were developed which may be used to select decontamination parameters for contamination scenarios including aircraft. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.

Rapid identification of Bacillus anthracis spores in suspicious powder samples by using matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS).

PubMed

Dybwad, Marius; van der Laaken, Anton L; Blatny, Janet Martha; Paauw, Armand

2013-09-01

Rapid and reliable identification of Bacillus anthracis spores in suspicious powders is important to mitigate the safety risks and economic burdens associated with such incidents. The aim of this study was to develop and validate a rapid and reliable laboratory-based matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis method for identifying B. anthracis spores in suspicious powder samples. A reference library containing 22 different Bacillus sp. strains or hoax materials was constructed and coupled with a novel classification algorithm and standardized processing protocol for various powder samples. The method's limit of B. anthracis detection was determined to be 2.5 × 10(6) spores, equivalent to a 55-μg sample size of the crudest B. anthracis-containing powder discovered during the 2001 Amerithrax incidents. The end-to-end analysis method was able to successfully discriminate among samples containing B. anthracis spores, closely related Bacillus sp. spores, and commonly encountered hoax materials. No false-positive or -negative classifications of B. anthracis spores were observed, even when the analysis method was challenged with a wide range of other bacterial agents. The robustness of the method was demonstrated by analyzing samples (i) at an external facility using a different MALDI-TOF MS instrument, (ii) using an untrained operator, and (iii) using mixtures of Bacillus sp. spores and hoax materials. Taken together, the observed performance of the analysis method developed demonstrates its potential applicability as a rapid, specific, sensitive, robust, and cost-effective laboratory-based analysis tool for resolving incidents involving suspicious powders in less than 30 min.

Rapid Identification of Bacillus anthracis Spores in Suspicious Powder Samples by Using Matrix-Assisted Laser Desorption Ionization–Time of Flight Mass Spectrometry (MALDI-TOF MS)

PubMed Central

van der Laaken, Anton L.; Blatny, Janet Martha; Paauw, Armand

2013-01-01

Rapid and reliable identification of Bacillus anthracis spores in suspicious powders is important to mitigate the safety risks and economic burdens associated with such incidents. The aim of this study was to develop and validate a rapid and reliable laboratory-based matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) analysis method for identifying B. anthracis spores in suspicious powder samples. A reference library containing 22 different Bacillus sp. strains or hoax materials was constructed and coupled with a novel classification algorithm and standardized processing protocol for various powder samples. The method's limit of B. anthracis detection was determined to be 2.5 × 106 spores, equivalent to a 55-μg sample size of the crudest B. anthracis-containing powder discovered during the 2001 Amerithrax incidents. The end-to-end analysis method was able to successfully discriminate among samples containing B. anthracis spores, closely related Bacillus sp. spores, and commonly encountered hoax materials. No false-positive or -negative classifications of B. anthracis spores were observed, even when the analysis method was challenged with a wide range of other bacterial agents. The robustness of the method was demonstrated by analyzing samples (i) at an external facility using a different MALDI-TOF MS instrument, (ii) using an untrained operator, and (iii) using mixtures of Bacillus sp. spores and hoax materials. Taken together, the observed performance of the analysis method developed demonstrates its potential applicability as a rapid, specific, sensitive, robust, and cost-effective laboratory-based analysis tool for resolving incidents involving suspicious powders in less than 30 min. PMID:23811517

A fluorescence in situ staining method for investigating spores and vegetative cells of Clostridia by confocal laser scanning microscopy and structured illuminated microscopy.

PubMed

D'Incecco, P; Ong, L; Gras, S; Pellegrino, L

2018-04-18

Non-pathogenic spore-forming Clostridia are of increasing interest due to their application in biogas production and their capability to spoil different food products. The life cycle for Clostridium includes a spore stage that can assist in survival under environmentally stressful conditions, such as extremes of temperature or pH. Due to their size, spores can be investigated by a range of microscopic techniques, many of which involve sample pre-treatment. We have developed a quick, simple and non-destructive fluorescent staining procedure that allows a clear differentiation between spores and vegetative cells and effectively stains spores, allowing recovery and tracking in subsequent experiments. Hoechst 34580, Propidium iodide and wheat germ agglutinin WGA 488 were used in combination to stain four strains of Clostridia at different life cycle stages. Staining was conducted without drying the sample, preventing changes induced by dehydration and cells observed by confocal laser scanner microscopy or using a super-resolution microscope equipped with a 3D-structured illumination module. Dual staining with Hoechst/Propidium iodide differentiated spores from vegetative cells, provided information on the viability of cells and was successfully applied to follow spore production induced by heating. Super-resolution microscopy of spores probed by Hoechst 34580 also allowed chromatin to be visualised. Direct staining of a cheese specimen using Nile Red and Fast Green allowed in situ observation of spores within the cheese and their position within the cheese matrix. The proposed staining method has broad applicability and can potentially be applied to follow Clostridium spore behaviour in a range of different environments. Copyright © 2018 Elsevier Ltd. All rights reserved.

Oral delivery of Bacillus subtilis spore expressing enolase of Clonorchis sinensis in rat model: induce systemic and local mucosal immune responses and has no side effect on liver function.

PubMed

Yu, Jinyun; Chen, Tingjin; Xie, Zhizhi; Liang, Pei; Qu, Honglin; Shang, Mei; Mao, Qiang; Ning, Dan; Tang, Zeli; Shi, Mengchen; Zhou, Lina; Huang, Yan; Yu, Xinbing

2015-07-01

Caused by the consumption of raw or undercooked freshwater fish containing infective metacercariae of Clonorchis sinensis, human clonorchiasis remains a major public health problem in China. In previous study, we had expressed enolase from C. sinensis (CsENO) on the surface of Bacillus subtilis spore and the recombinant spore induced a pronounced protection in terms of reduced worm burden and eggs per gram feces, suggesting B. subtilis spore as an ideal vehicle for antigen delivery by oral treatment and CsENO as a promising vaccine candidate against clonorchiasis. In the current study, we detected CsENO-specific IgG and IgA levels both in serum and in intestinal mucus from rats orally administrated with B. subtilis spore surface expressing CsENO by ELISA. Lysozyme levels in serum and in intestinal mucus were analyzed too. In addition, IgA-secreting cells in intestine epithelium of the rats were detected by immunohistochemistry assay. The intestinal villi lengths of duodenum, jejunum, and ileum were also measured. Rats orally treated with B. subtilis spore or normal saline were used as controls. Our results showed that, compared with the control groups, oral administration of B. subtilis spore expressing CsENO induced both systemic and local mucosal immune response. The recombinant spores also enhanced non-specific immune response in rats. The spores had no side effect on liver function. Moreover, it might facilitate food utilization and digestion of the rats. Our work will pave the way to clarify the involved mechanisms of protective efficacy elicited by B. subtilis spore expressing CsENO and encourage us to carry out more assessment trails of the oral treated spore to develop vaccine against clonorchiasis.

Characterization of the yrbA Gene of Bacillus subtilis, Involved in Resistance and Germination of Spores

PubMed Central

Takamatsu, Hiromu; Kodama, Takeko; Nakayama, Tatsuo; Watabe, Kazuhito

1999-01-01

Insertional inactivation of the yrbA gene of Bacillus subtilis reduced the resistance of the mutant spores to lysozyme. The yrbA mutant spores lost their optical density at the same rate as the wild-type spores upon incubation with l-alanine but became only phase gray and did not swell. The response of the mutant spores to a combination of asparagine, glucose, fructose, and KCl was also extremely poor; in this medium yrbA spores exhibited only a small loss in optical density and gave a mixture of phase-bright, -gray, and -dark spores. Northern blot analysis of yrbA transcripts in various sig mutants indicated that yrbA was transcribed by RNA polymerase with ςE beginning at 2 h after the start of sporulation. The yrbA promoter was localized by primer extension analysis, and the sequences of the −35 (TCATAAC) and −10 (CATATGT) regions were similar to the consensus sequences of genes recognized by ςE. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of proteins solubilized from intact yrbA mutant spores showed an alteration in the protein profile, as 31- and 36-kDa proteins, identified as YrbA and CotG, respectively, were absent, along with some other minor changes. Electron microscopic examination of yrbA spores revealed changes in the spore coat, including a reduction in the density and thickness of the outer layer and the appearance of an inner coat layer-like structure around the outside of the coat. This abnormal coat structure was also observed on the outside of the developing forespores of the yrbA mutant. These results suggest that YrbA is involved in assembly of some coat proteins which have roles in both spore lysozyme resistance and germination. PMID:10438771

Effect of water temperature on the development, release and survival of the triactinomyxon stage of Myxobolus cerebralis in its oligochaete host.

PubMed

El-Matbouli, M; McDowell, T S; Antonio, D B; Andree, K B; Hedrick, R P

1999-04-01

The development of the triactinomyxon stage of Myxobolus cerebralis and release of mature spores from Tubifex tubifex were shown to be temperature dependent. In the present work, the effect of temperature over a range of 5-30 degrees C on the development and release of the triactinomyxon stages of M. cerebralis was studied. Infected T. tubifex stopped releasing triactinomyxon spores 4 days after transfer from 15 degrees C to 25 degrees C or 30 degrees C. Transmission electron microscopic examinations of the tubificids held at 25 degrees C and 30 degrees C for 3 days showed that all developmental stages degenerated and transformed to electron-dense clusters between the gut epithelial cells of T. tubifex. In contrast, tubificid worms held at 5 degrees C and 10 degrees C examined at the same time were heavily infected with many early developmental stages of triactinomyxon. At 15 degrees C, the optimal temperature for development, maturing and mature stages of the parasite were evident. Infected T. tubifex transferred from 15 degrees C to 20 degrees C stopped producing triactinomyxon spores after 15 days. However, 15 days at 20 degrees C was not sufficient to destroy all developmental stages of the parasite. When the tubificid worms were returned to 15 degrees C, the one-cell stages and the binucleate-cell stages resumed normal growth. It was also demonstrated that T. tubifex cured of infection by holding at 30 degrees C for 3 weeks and shifted to 15 degrees C could be re-infected with M. cerebralis spores. The waterborne triactinomyxon spores of M. cerebralis did not appear to be as short-lived as previously reported. More than 60% of experimentally produced waterborne triactinomyxon spores survived and maintained their infectivity for rainbow trout for 15 days at water temperatures up to 15 degrees C. In natural aquatic systems, the triactinomyxon spores may survive and keep their infectivity for periods even longer than 15 days.

Programmed cell death in vegetative development: apoptosis during the colonial life cycle of the ascidian Botryllus schlosseri.

PubMed

Tiozzo, S; Ballarin, L; Burighel, P; Zaniolo, G

2006-06-01

Programmed cell death (PCD) by apoptosis is a physiological mechanism by which cells are eliminated during embryonic and post-embryonic stages of animal life cycle. During asexual reproduction, the zooids of colonial ascidians originate from an assorted cell population instead of a single zygote, so that we assume that regulation of the equilibrium among proliferation, differentiation and cell death may follow different pathways in comparison to the embryonic development. Here we investigate the presence of apoptotic events throughout the blastogenetic life cycle of the colonial ascidian Botryllus schlosseri, by means of terminal deoxynucleotidyl transferase dUTP Nick End Labeling (TUNEL) coupled with histochemical and electron microscopy techniques. The occurrence of low levels of morphogenetic cell death suggests that, in contrast to what happens during sexual development (embryogenesis and metamorphosis), apoptosis does not play a pivotal role during asexual propagation in botryllid ascidian. Nevertheless, PCD emerges as a key force to regulate homeostasis in adult zooids and to shape and modulate the growth of the whole colony.

Development and pathology of two undescribed species of microsporidia infecting the predatory mite, Phytoseiulus persimilis Athias-Henriot.

PubMed

Bjøornson, S; Keddie, B A

2000-11-01

Two undescribed species of microsporidia were found in mass-reared Phytoseiulus persimilis Athias-Henriot from two commercial sources during a routine examination of these predators for pathogens. Both microsporidian species were described from specimens that had been prepared for transmission electron microscopy; live specimens were unavailable for examination. One microsporidium, identified as Species A, was described from two specimens obtained from a commercial insectary in North America. All observed stages of this microsporidium were uninucleate. Rounded-to-ovoid schizonts appeared to develop in direct contact with the cytoplasm of lyrate organ cells (ovarian tissue). Mature spores of Species A were elongate-ovoid and measured 2.88 x 1.21 microm. A polar filament coiled 7 to 10 times in the posterior half of the spore. Sporoblasts and spores were observed in the cytoplasm of cells of numerous tissues and in developing eggs within gravid females. A second species, identified as Species B, was described from five specimens obtained from a commercial source in Israel. All observed stages of this microsporidium were uninucleate. Schizonts of Species B were observed within the cytoplasm of cecal wall cells and within the nuclei of lyrate organ cells. Mature spores were ovoid and measured 2.65 x 1.21 microm. A polar filament coiled 3 to 4 times in the posterior half of the spore. Densely packed ribosomes often concealed the polar filament and other internal spore characteristics. Spores were observed in the cytoplasm of cells of numerous tissues and occasionally within the nuclei of lyrate organ cells. Numerous spores and presporal stages were observed within the ovary and developing eggs. The development and pathology of Species A and B were compared to those of Microsporidium phytoseiuli Bjøornson, Steiner and Keddie, a microsporidium previously described from P. persimilis obtained from a commercial source in Europe. The occurrence of three species of microsporidia within P. persimilis from three sources raises questions regarding the origin of these pathogens. Because microsporidia may have profound impact on the performance of P. persimilis, consideration must be given to the identification and exclusion of microsporidia from field-collected specimens or from predators that may be shared among commercial sources.

Re-evaluation of endogenous development of Eimeria bareillyi Gill, Chhabra and Lall, 1963 in water buffalo (Bubalus bubalis).

PubMed

Dubey, J P

2018-04-25

Water buffalo (Bubalus bubalis) is important for the economy of Asia, South America and parts of Europe. Coccidiosis is an important cause of neonatal mortality in livestock, including buffalo. Of more than 12 species of Eimeria in buffalo, Eimeria bareillyi is the most pathogenic. There are uncertainties concerning its asexual and sexual development. During a previously reported outbreak of fatal enteritis associated with E. bareillyi in buffaloes in the Netherlands, sections of small intestine were re-evaluated histologically and by transmission electron microscopy (TEM) to seek details of endogenous development. Profuse asexual multiplication occurred in the jejunum and ileum. Light microscopic examination revealed that parasites divided in two (probably endodyogeny) or more organisms. There were two or more generations of morphologically different merozoites; some of these observations were confirmed by TEM. Details of gametogonic development, including oocyst wall formation are provided. Schizogonic and gametogonic development described in the present study can serve as a guide for differential diagnosis of Eimeria species in histological sections of intestines of buffaloes.

Exploring the interaction network of the Bacillus subtilis outer coat and crust proteins.

PubMed

Krajčíková, Daniela; Forgáč, Vladimír; Szabo, Adam; Barák, Imrich

2017-11-01

Bacillus subtilis spores, representatives of an exceptionally resistant dormant cell type, are encircled by a thick proteinaceous layer called the spore coat. More than 80 proteins assemble into four distinct coat layers: a basement layer, an inner coat, an outer coat and a crust. As the spore develops inside the mother cell, spore coat proteins synthesized in the cytoplasm are gradually deposited onto the prespore surface. A small set of morphogenetic proteins necessary for spore coat morphogenesis are thought to form a scaffold to which the rest of the coat proteins are attached. Extensive localization and proteomic studies using wild type and mutant spores have revealed the arrangement of individual proteins within the spore coat layers. In this study we examined the interactions between the proteins localized to the outer coat and crust using a bacterial two hybrid system. These two layers are composed of at least 25 components. Self-interactions were observed for most proteins and numerous novel interactions were identified. The most interesting contacts are those made with the morphogenetic proteins CotE, CotY and CotZ; these could serve as a basis for understanding the specific roles of particular proteins in spore coat morphogenesis. Copyright © 2017 Elsevier GmbH. All rights reserved.

Improvement of Biological Indicators by Uniformly Distributing Bacillus subtilis Spores in Monolayers To Evaluate Enhanced Spore Decontamination Technologies

PubMed Central

Raguse, Marina; Fiebrandt, Marcel; Stapelmann, Katharina; Madela, Kazimierz; Laue, Michael; Lackmann, Jan-Wilm; Thwaite, Joanne E.; Setlow, Peter; Awakowicz, Peter

2016-01-01

Novel decontamination technologies, including cold low-pressure plasma and blue light (400 nm), are promising alternatives to conventional surface decontamination methods. However, the standardization of the assessment of such sterilization processes remains to be accomplished. Bacterial endospores of the genera Bacillus and Geobacillus are frequently used as biological indicators (BIs) of sterility. Ensuring standardized and reproducible BIs for reliable testing procedures is a significant problem in industrial settings. In this study, an electrically driven spray deposition device was developed, allowing fast, reproducible, and homogeneous preparation of Bacillus subtilis 168 spore monolayers on glass surfaces. A detailed description of the structural design as well as the operating principle of the spraying device is given. The reproducible formation of spore monolayers of up to 5 × 107 spores per sample was verified by scanning electron microscopy. Surface inactivation studies revealed that monolayered spores were inactivated by UV-C (254 nm), low-pressure argon plasma (500 W, 10 Pa, 100 standard cubic cm per min), and blue light (400 nm) significantly faster than multilayered spores were. We have thus succeeded in the uniform preparation of reproducible, highly concentrated spore monolayers with the potential to generate BIs for a variety of nonpenetrating surface decontamination techniques. PMID:26801572

Asexuality: an extreme variant of sexual desire disorder?

PubMed

Brotto, Lori A; Yule, Morag A; Gorzalka, Boris B

2015-03-01

Human asexuality is defined as a lack of sexual attraction to anyone or anything. Various theories have been proposed to explain how asexuality should best be classified, and some have maintained that asexuality is an extreme variant of hypoactive sexual desire disorder (HSDD)-a sexual dysfunction characterized by a lack of interest in sex and significant distress. To date, this has never been empirically examined. Using measures of sexual desire and behavior, sex-related distress, personality, and psychopathology, the aim of the current study was to compare individuals scoring above the cutoff for asexuality identification (AIS >40) (n = 192) to sexual individuals (n = 231). The sexual group was further divided into a control group (n = 122), a HSDD group (n = 50), and a group with symptoms of low desire that were nondistressing (n = 59). Analyses were controlled for age. Individuals in the AIS >40 group had a greater likelihood of never previously engaging in sexual intercourse, fantasies, or kissing and petting than all other groups and a lower likelihood of experiencing sex-related distress than those with HSDD. For women, those in the HSDD and AIS >40 groups had significantly lower desire than the subclinical HSDD and control groups. Men in the AIS >40 group had significantly lower desire than the other three groups. Symptoms of depression were highest among those with subclinical HSDD and HSDD, whereas there were no group differences on alexithymia or desirable responding. A binary logistic regression indicated that relationship status (long-term dating/married), sexual desire, sex-related distress, and lower alexithymia scores were the best predictors of group membership (HSDD vs. AIS >40). Taken together, these results challenge the speculation that asexuality should be classified as a sexual dysfunction of low desire. © 2014 International Society for Sexual Medicine.

Impact of three phthalate esters on the sexual reproduction of the Monogonont rotifer, Brachionus calyciflorus.

PubMed

Cruciani, V; Iovine, C; Thomé, J-P; Joaquim-Justo, C

2016-01-01

Phthalate esters are widespread contaminants that can cause endocrine disruption in vertebrates. Studies showed that molecules with hormonal activities in vertebrates and invertebrates can affect asexual and sexual reproduction in rotifers. We investigated the impact of di-hexylethyl phthalate (DEHP), di-butyl phthalate (DBP) and butylbenzyl phthalate (BBP), on the asexual and sexual reproduction of the freshwater monogonont rotifer Brachionus calyciflorus in order to determine a potential environmental risk for sexual reproduction. We observed that DEHP has no significant impact on both asexual and sexual reproduction up to 2 mg/L. DBP has a positive effect on asexual reproduction at concentrations from 0.05 to 1 mg/L, but depresses it at 2 mg/L. Sexual reproduction is only affected at 2 mg/L and the impact observed is negative. BBP displayed a negative impact on both asexual and sexual reproduction at 1 and 2 mg/L. However we showed that the impacts of BBP on mixis and fertilization rates observed are due to the decrease in population growth rates at these concentrations and not to a direct impact of BBP on the mixis and the fertilization processes. Our results show that sexual reproduction in B. calyciflorus is not more sensitive than asexual reproduction to any of the substances tested which indicates the mode of action of these molecules is related to general toxicity and not to an interference with potential endocrine regulation of sexual reproduction. Comparison of effect concentrations and surface water contamination by phthalate esters suggests these compounds do not constitute a risk for primary consumers in these environments.

In vitro production of Clostridium difficile spores for use in the efficacy evaluation of disinfectants: a precollaborative investigation.

PubMed

Hasan, Jafrul A; Japal, Knoxley M; Christensen, Erick R; Samalot-Freire, Luisa C

2011-01-01

Clostridium difficile is a strict anaerobic spore-forming bacterium, and an increasingly common nosocomial pathogen. The U.S. Environmental Protection Agency (EPA) is responsible for the registration of disinfectants, including products designed to treat environmental surfaces contaminated with spores of C. difficile. Product efficacy data are required for registration; however, there is a lack of methodology for generating high-quality spore suspensions for evaluating product performance. As such, a study was carried out to select a suitable C. difficile strain and to develop a stand-alone method to prepare a spore suspension that meets specific criteria necessary for quantitative testing of disinfectants. The criteria are: (1) a spore titer of > 8 log10/mL, (2) > or = 90% spores to vegetative cells, and (3) resistance of spores (determined by viability) to 2.5 M hydrochloric acid (HCl). Several strains of C. difficile (toxigenic and nontoxigenic) were grown on various media (solid and liquid) for varying lengths of time to determine the best combination of incubation conditions and media to optimize spore production and quality. Once the spore production procedure was optimized, a toxigenic strain of C. difficile [American Type Culture Collection (ATCC) 43598] was selected for use in trials to verify repeatability from one production run to the next. The spore suspension was initiated by spreading vegetative cells of C. difficile (ATCC 43598) on CDC anaerobic 5% sheep blood agar plates and incubating for 7-10 days at 36 +/- 1 degrees C under anaerobic conditions. Spores were harvested when > or = 90% of the cells converted to spores as determined by observation using phase-contrast microscopy. The spores were washed three times with saline-Tween-80, resuspended in cold deionized water, heated to 70 degrees C for 10 min, evaluated microscopically for quality, and enumerated on cycloserine-cefoxitin-fructose agar containing horse blood and taurocholate. The spore suspension was used to inoculate brushed stainless steel carriers (1 cm in diameter) with and without a soil load in accordance with the Standard Quantitative Carrier Disk Test Method (ASTM E-2197-02) to determine carrier load. Once it was determined that > 6 log10 spores/carrier could be recovered, spores were evaluated for resistance to HCI. The sporulation method presented in this report is simple and repeatable and results in spore suspension of high titer (> 8 log10/mL) and quality (> or = 90% spores to vegetative cells) that met acid resistance criteria (spores were resistant to 2.5 M HCI for 10 min). In addition, recovery from brushed stainless steel carriers with and without soil load was > 6 log10 spores/carrier. A 6 log10 performance standard was set forth in the EPA's interim guidance for generating data to support a label claim for effectiveness against C. difficile spores on hard, nonporous surfaces. This precollaborative investigation successfully demonstrated the use of a methodology for in vitro production of C. difficile spores (ATCC 43598) necessary for conducting efficacy tests. A proposal will be submitted to the AOAC INTERNATIONAL Methods Committee on Antimicrobial Efficacy Testing for a collaborative study; see Appendix.

One-Step Purification of Enterocytozoon bieneusi Spores from Human Stools by Immunoaffinity Expanded-Bed Adsorption

PubMed Central

Accoceberry, Isabelle; Thellier, Marc; Datry, Annick; Desportes-Livage, Isabelle; Biligui, Sylvestre; Danis, Martin; Santarelli, Xavier

2001-01-01

An original, reliable, and reproducible method for the purification of Enterocytozoon bieneusi spores from human stools is described. We recently reported the production of a species-specific monoclonal antibody (MAb) 6E52D9 immunoglobulin G2a (IgG2a) raised against the exospore of E. bieneusi spore walls. The MAb was used as a ligand to develop an immunoaffinity matrix. The mouse IgG2a MAb was bound directly to a Streamline rProtein A adsorbent, used for expanded-bed adsorption of immunoglobulins, for optimal spatial orientation of the antibody and maximum binding efficiency of the antigen. The complex was then cross-linked covalently using dimethyl pimelimidate dihydrochloride. After incubation of the immunoaffinity matrix with filtered stool samples containing numerous E. bieneusi spores and before elution with 6 M guanidine HCl, the expansion of the adsorbent bed eliminated all the fecal contaminants. The presence of spores in the elution fractions was determined by an indirect immunofluorescence antibody test (IFAT). E. bieneusi spores were found in the elution fraction in all four experiments and were still highly antigenic as indicated by IFAT. Smears examined by light microscopy contained very clean spores with no fecal debris or background bacterial and fungal contaminants. However, spore recovery rates were relatively low: an average of 107 spores were purified per run. This technique for isolating E. bieneusi spores directly from human stool samples with a high degree of purity opens up new approaches for studying this parasite. PMID:11326019

Separation of mycotoxin-containing sources in grain dust and determination of their mycotoxin potential.

PubMed Central

Palmgren, M S; Lee, L S

1986-01-01

Two distinct reservoirs of mycotoxins exist in fungal-infected cereal grains--the fungal spores and the spore-free mycelium-substrate matrix. Many fungal spores are of respirable size and the mycelium-substrate matrix can be pulverized to form particles of respirable size during routine handling of grain. In order to determine the contribution of each source to the level of mycotoxin contamination of dust, we developed techniques to harvest and separate mycelium-substrate matrices from spores of fungi. Conventional quantitative chromatographic analyses of separated materials indicated that aflatoxin from Aspergillus parasiticus, norsolorinic acid from a mutant of A. parasiticus, and secalonic acid D from Penicillium oxalicum were concentrated in the mycelium-substrate matrices and not in the spores. In contrast, spores of Aspergillus niger and Aspergillus fumigatus contained significant concentrations of aurasperone C and fumigaclavine C, respectively; only negligible amounts of the toxins were detected in the mycelium-substrate matrices of these two fungi. PMID:3709472

As you reap, so shall you sow: coupling of harvesting and inoculating stabilizes the mutualism between termites and fungi

PubMed Central

Aanen, Duur K

2006-01-01

At present there is no consensus theory explaining the evolutionary stability of mutualistic interactions. However, the question is whether there are general ‘rules’, or whether each particular mutualism needs a unique explanation. Here, I address the ultimate evolutionary stability of the ‘agricultural’ mutualism between fungus-growing termites and Termitomyces fungi, and provide a proximate mechanism for how stability is achieved. The key to the proposed mechanism is the within-nest propagation mode of fungal symbionts by termites. The termites suppress horizontal fungal transmission by consuming modified unripe mushrooms (nodules) for food. However, these nodules provide asexual gut-resistant spores that form the inoculum of new substrate. This within-nest propagation has two important consequences: (i) the mutualistic fungi undergo severe, recurrent bottlenecks, so that the fungus is likely to be in monoculture and (ii) the termites ‘artificially’ select for high nodule production, because their fungal food source also provides the inoculum for the next harvest. I also provide a brief comparison of the termite–fungus mutualism with the analogous agricultural mutualism between attine ants and fungi. This comparison shows that—although common factors for the ultimate evolutionary stability of mutualisms can be identified—the proximate mechanisms can be fundamentally different between different mutualisms. PMID:17148364

[Morphofunctional organization of reserve stem cells providing for asexual and sexual reproduction of invertebrates].

PubMed

Isaeva, V V; Akhmadieva, A V; Aleksandriova, Ia N; Shukaliuk, A I

2009-01-01

Published and original data indicating evolutionary conservation of the morphofunctional organization of reserve stem cells providing for asexual and sexual reproduction of invertebrates are reviewed. Stem cells were studied in representatives of five animal types: archeocytes in sponge Oscarella malakhovi (Porifera), large interstitial cells in colonial hydroid Obelia longissima (Cnidaria), neoblasts in an asexual race of planarian Girardia tigrina (Platyhelmintes), stem cells in colonial rhizocephalans Peltogasterella gracilis, Polyascus polygenea, and Thylacoplethus isaevae (Arthropoda), and colonial ascidian Botryllus tuberatus (Chordata). Stem cells in animals of such diverse taxa feature the presence of germinal granules, are positive for proliferating cell nuclear antigen, demonstrate alkaline phosphatase activity (at marker of embryonic stem cells and primary germ cells in vertebrates), and rhizocephalan stem cells express the vasa-like gene (such genes are expressed in germline cells of different metazoans). The self-renewing pool of stem cells is the cellular basis of the reproductive strategy including sexual and asexual reproduction.

Sex as a strategy against rapidly evolving parasites

PubMed Central

Tinkler, Shona K.; Tinsley, Matthew C.

2016-01-01

Why is sex ubiquitous when asexual reproduction is much less costly? Sex disrupts coadapted gene complexes; it also causes costs associated with mate finding and the production of males who do not themselves bear offspring. Theory predicts parasites select for host sex, because genetically variable offspring can escape infection from parasites adapted to infect the previous generations. We examine this using a facultative sexual crustacean, Daphnia magna, and its sterilizing bacterial parasite, Pasteuria ramosa. We obtained sexually and asexually produced offspring from wild-caught hosts and exposed them to contemporary parasites or parasites isolated from the same population one year later. We found rapid parasite adaptation to replicate within asexual but not sexual offspring. Moreover, sexually produced offspring were twice as resistant to infection as asexuals when exposed to parasites that had coevolved alongside their parents (i.e. the year two parasite). This fulfils the requirement that the benefits of sex must be both large and rapid for sex to be favoured by selection. PMID:28003455

Sex as a strategy against rapidly evolving parasites.

PubMed

Auld, Stuart K J R; Tinkler, Shona K; Tinsley, Matthew C

2016-12-28

Why is sex ubiquitous when asexual reproduction is much less costly? Sex disrupts coadapted gene complexes; it also causes costs associated with mate finding and the production of males who do not themselves bear offspring. Theory predicts parasites select for host sex, because genetically variable offspring can escape infection from parasites adapted to infect the previous generations. We examine this using a facultative sexual crustacean, Daphnia magna, and its sterilizing bacterial parasite, Pasteuria ramosa We obtained sexually and asexually produced offspring from wild-caught hosts and exposed them to contemporary parasites or parasites isolated from the same population one year later. We found rapid parasite adaptation to replicate within asexual but not sexual offspring. Moreover, sexually produced offspring were twice as resistant to infection as asexuals when exposed to parasites that had coevolved alongside their parents (i.e. the year two parasite). This fulfils the requirement that the benefits of sex must be both large and rapid for sex to be favoured by selection. © 2016 The Author(s).

Population genomics reveals the origin and asexual evolution of human infective trypanosomes

PubMed Central

Weir, William; Capewell, Paul; Foth, Bernardo; Clucas, Caroline; Pountain, Andrew; Steketee, Pieter; Veitch, Nicola; Koffi, Mathurin; De Meeûs, Thierry; Kaboré, Jacques; Camara, Mamadou; Cooper, Anneli; Tait, Andy; Jamonneau, Vincent; Bucheton, Bruno; Berriman, Matt; MacLeod, Annette

2016-01-01

Evolutionary theory predicts that the lack of recombination and chromosomal re-assortment in strictly asexual organisms results in homologous chromosomes irreversibly accumulating mutations and thus evolving independently of each other, a phenomenon termed the Meselson effect. We apply a population genomics approach to examine this effect in an important human pathogen, Trypanosoma brucei gambiense. We determine that T.b. gambiense is evolving strictly asexually and is derived from a single progenitor, which emerged within the last 10,000 years. We demonstrate the Meselson effect for the first time at the genome-wide level in any organism and show large regions of loss of heterozygosity, which we hypothesise to be a short-term compensatory mechanism for counteracting deleterious mutations. Our study sheds new light on the genomic and evolutionary consequences of strict asexuality, which this pathogen uses as it exploits a new biological niche, the human population. DOI: http://dx.doi.org/10.7554/eLife.11473.001 PMID:26809473

Planetary quarantine program

NASA Technical Reports Server (NTRS)

1971-01-01

A quantitative means was developed to investigate the sensitivity of current spacecraft sterilization plans to variations in D-values. A quantitative expression was derived to represent the distribution of D-values among a population of naturally occurring organisms. An investigation was made of (1) the inactivation of both Bacillus subtilis var. niger spores and Cape Kennedy soil spores by gamma-radiation at room temperature in a nitrogen environment, and (2) the thermoradiation resistance of Cape Kennedy soil spores at elevated temperatures below 125 C. The relation between standard survival experiments with bacterial spores in soils and results obtained on spacecraft surfaces is discussed. Sporocidal properties of aqueous formaldehyde can be increased by elevating the temperature.

Thermal Inactivation of Aerosolized Bacillus subtilis var. niger Spores

PubMed Central

Mullican, Charles L.; Buchanan, Lee M.; Hoffman, Robert K.

1971-01-01

A hot-air sterilizer capable of exposing airborne microorganisms to elevated temperatures with an almost instantaneous heating time was developed and evaluated. With this apparatus, aerosolized Bacillus subtilis var. niger spores were killed in about 0.02 sec when exposed to temperatures above 260 C. This is about 500 times faster than killing times reported by others. Extrapolation and comparison of data on the time and temperature required to klll B. subtilis var. niger spores on surfaces show that approximately the same killing time is required as is necessary for spores in air, if corrections are made for the heating time of the surface. PMID:5002138

Effects of fungal species, cultivation time, growth substrate, and air exposure velocity on the fluorescence properties of airborne fungal spores.

PubMed

Saari, S; Mensah-Attipoe, J; Reponen, T; Veijalainen, A M; Salmela, A; Pasanen, P; Keskinen, J

2015-12-01

Real-time bioaerosol monitoring is possible with fluorescence based instruments. This study provides information on major factors that can affect the fluorescence properties of airborne fungal spores. Two fluorescence-based bioaerosol detectors, BioScout, and ultraviolet aerodynamic particle sizer (UVAPS), were used to study fluorescent particle fractions (FPFs) of released spores of three fungal species (Aspergillus versicolor, Cladosporium cladosporioides, and Penicillium brevicompactum). Two culture media (agar and gypsum board), three ages of the culture (one week, one month, and four months), and three aerosolization air velocities (5, 15, and 27 m/s) were tested. The results showed that the FPF values for spores released from gypsum were typically lower than for those released from agar indicating that poor nutrient substrate produces spores with lower amounts of fluorescent compounds. The results also showed higher FPF values with lower air velocities in aerosolization. This indicates that easily released fully developed spores have more fluorescent compounds compared to forcibly extracted non-matured spores. The FPFs typically were lower with older samples. The FPF results between the two instruments were similar, except with four-month-old samples. The results can be utilized in field measurements of fungal spores to estimate actual concentrations and compare different instruments with fluorescence-based devices as well as in instrument calibration and testing in laboratory conditions. Fluorescence-based instruments are the only choice for real-time detection of fungal spores at the moment. In general, all fluorescence-based bioaerosol instruments are tested against known bacterial and fungal spores in laboratory conditions. This study showed that fungal species, growth substrate, age of culture, and air current exposure rate have an effect on detection efficiency of fungal spores in the fluorescence-based instruments. Therefore, these factors should be considered in the instrument calibration process. The results are also important when interpreting results of fluorescence-based field measurements of fungal spores. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Developing a Table of Forces for Human Activity as It Relates ...

EPA Pesticide Factsheets

Report The purpose of this project was to evaluate forces generated by human activity that may cause reaerosolization of Bacillus anthracis spores in the aftermath of an intentional release. Understanding human interaction is important to inform communities on activities that may cause the re-distribution of spores. This report presents the results of an extensive literature review of human-surface interaction forces with respect to Bacillus spore reaerosolization in the outdoor environment.

Picturing space for lesbian nonsexualities: rethinking sex-normative commitments through The Kids Are All Right (2010).

PubMed

Gupta, Kristina

2013-01-01

This article examines representations of lesbian nonsexuality in the film The Kids Are All Right and in responses to the film by feminist and queer scholars. In some moments, the film offers a limited endorsement of lesbian nonsexuality, placing pressure on the category lesbian to include nonsexuality and asexuality. However, in their responses to the film, many feminist and queer scholars rejected nonsexuality as an aspect of lesbian experience, placing pressure on the category lesbian to exclude nonsexual and asexual women. Asexual activism challenges scholars to question their sex-normative commitments and to keep the category lesbian open and flexible.

Parasites in sexual and asexual mollies (Poecilia, Poeciliidae, Teleostei): a case for the Red Queen?

PubMed Central

Tobler, Michael; Schlupp, Ingo

2005-01-01

The maintenance of sexual reproduction in the face of its supposed costs is a major paradox in evolutionary biology. The Red Queen hypothesis, which states that sex is an adaptation to fast-evolving parasites, is currently one of the most recognized explanations for the ubiquity of sex and predicts that asexual lineages should suffer from a higher parasite load if they coexist with closely related sexuals. We tested this prediction using four populations of the sexual fish species Poecilia latipinna and its asexual relative Poecilia formosa. Contrary to expectation, no differences in parasite load could be detected between the two species. PMID:17148156

Detection of Bacillus spores within 15 minutes by surface-enhanced Raman spectroscopy

NASA Astrophysics Data System (ADS)

Shende, Chetan; Inscore, Frank; Huang, Hermes; Farquharson, Stuart; Sengupta, Atanu

2012-06-01

Since the distribution of Bacillus anthracis causing spores through the US Postal System, there has been a persistent fear that biological warfare agents (BWAs) will be used by terrorists against our military abroad and our civilians at home. Despite the substantial effort to develop BWA analyzers, they remain either too slow, produce high falsealarm rates, lack sensitivity, or cannot be fielded. Consequently there remains a need for a portable analyzer that can overcome these limitations as expressed at the 2011 Biological Weapons Convention. To meet this need we have been developing a sample system that selectively binds BWAs and produce surface-enhanced Raman (SER) spectra using portable Raman spectrometers. Here we describe the use of a short peptide ligand functionalized on silver nanoparticles to selectively capture Bacillus cereus spores (a surrogate of B. anthracis) and their subsequent detection by SER spectroscopy. This technique was used to specifically detect B. cereus spores over closely related species like B. subtilis belonging to the same genus within 15 minutes. Sensitivity of the method was demonstrated by detecting 104 B. cereus spores/mL of water. The technology, once developed should prove invaluable for rapid monitoring of BWAs, which will immensely help first responders and emergency personnel in implementing appropriate counter measures.

Quantification of Nonproteolytic Clostridium botulinum Spore Loads in Food Materials.

PubMed

Barker, Gary C; Malakar, Pradeep K; Plowman, June; Peck, Michael W

2016-01-04

We have produced data and developed analysis to build representations for the concentration of spores of nonproteolytic Clostridium botulinum in materials that are used during the manufacture of minimally processed chilled foods in the United Kingdom. Food materials are categorized into homogenous groups which include meat, fish, shellfish, cereals, fresh plant material, dairy liquid, dairy nonliquid, mushroom and fungi, and dried herbs and spices. Models are constructed in a Bayesian framework and represent a combination of information from a literature survey of spore loads from positive-control experiments that establish a detection limit and from dedicated microbiological tests for real food materials. The detection of nonproteolytic C. botulinum employed an optimized protocol that combines selective enrichment culture with multiplex PCR, and the majority of tests on food materials were negative. Posterior beliefs about spore loads center on a concentration range of 1 to 10 spores kg(-1). Posterior beliefs for larger spore loads were most significant for dried herbs and spices and were most sensitive to the detailed results from control experiments. Probability distributions for spore loads are represented in a convenient form that can be used for numerical analysis and risk assessments. Copyright © 2016 Barker et al.

The coat morphogenetic protein SpoVID is necessary for spore encasement in Bacillus subtilis.

PubMed

Wang, Katherine H; Isidro, Anabela L; Domingues, Lia; Eskandarian, Haig A; McKenney, Peter T; Drew, Kevin; Grabowski, Paul; Chua, Ming-Hsiu; Barry, Samantha N; Guan, Michelle; Bonneau, Richard; Henriques, Adriano O; Eichenberger, Patrick

2009-11-01

Endospores formed by Bacillus subtilis are encased in a tough protein shell known as the coat, which consists of at least 70 different proteins. We investigated the process of spore coat morphogenesis using a library of 40 coat proteins fused to green fluorescent protein and demonstrate that two successive steps can be distinguished in coat assembly. The first step, initial localization of proteins to the spore surface, is dependent on the coat morphogenetic proteins SpoIVA and SpoVM. The second step, spore encasement, requires a third protein, SpoVID. We show that in spoVID mutant cells, most coat proteins assembled into a cap at one side of the developing spore but failed to migrate around and encase it. We also found that SpoIVA directly interacts with SpoVID. A domain analysis revealed that the N-terminus of SpoVID is required for encasement and is a structural homologue of a virion protein, whereas the C-terminus is necessary for the interaction with SpoIVA. Thus, SpoVM, SpoIVA and SpoVID are recruited to the spore surface in a concerted manner and form a tripartite machine that drives coat formation and spore encasement.

The coat morphogenetic protein SpoVID is necessary for spore encasement in Bacillus subtilis

PubMed Central

Wang, Katherine H.; Isidro, Anabela L.; Domingues, Lia; Eskandarian, Haig A.; McKenney, Peter T.; Drew, Kevin; Grabowski, Paul; Chua, Ming-Hsiu; Barry, Samantha N.; Guan, Michelle; Bonneau, Richard; Henriques, Adriano O.; Eichenberger, Patrick

2009-01-01

SUMMARY Endospores formed by Bacillus subtilis are encased in a tough protein shell known as the coat, which consists of at least 70 different proteins. We investigated the process of spore coat morphogenesis using a library of 40 coat proteins fused to GFP and demonstrate that two successive steps can be distinguished in coat assembly. The first step, initial localization of proteins to the spore surface, is dependent on the coat morphogenetic proteins SpoIVA and SpoVM. The second step, spore encasement, requires a third protein, SpoVID. We show that in spoVID mutant cells, most coat proteins assembled into a cap at one side of the developing spore but failed to migrate around and encase it. We also found that SpoIVA directly interacts with SpoVID. A domain analysis revealed that the N-terminus of SpoVID is required for encasement and is a structural homolog of a virion protein, whereas the C-terminus is necessary for the interaction with SpoIVA. Thus, SpoVM, SpoIVA and SpoVID are recruited to the spore surface in a concerted manner and form a tripartite machine that drives coat formation and spore encasement. PMID:19775244

Quantification of Nonproteolytic Clostridium botulinum Spore Loads in Food Materials

PubMed Central

Barker, Gary C.; Malakar, Pradeep K.; Plowman, June

2016-01-01

We have produced data and developed analysis to build representations for the concentration of spores of nonproteolytic Clostridium botulinum in materials that are used during the manufacture of minimally processed chilled foods in the United Kingdom. Food materials are categorized into homogenous groups which include meat, fish, shellfish, cereals, fresh plant material, dairy liquid, dairy nonliquid, mushroom and fungi, and dried herbs and spices. Models are constructed in a Bayesian framework and represent a combination of information from a literature survey of spore loads from positive-control experiments that establish a detection limit and from dedicated microbiological tests for real food materials. The detection of nonproteolytic C. botulinum employed an optimized protocol that combines selective enrichment culture with multiplex PCR, and the majority of tests on food materials were negative. Posterior beliefs about spore loads center on a concentration range of 1 to 10 spores kg−1. Posterior beliefs for larger spore loads were most significant for dried herbs and spices and were most sensitive to the detailed results from control experiments. Probability distributions for spore loads are represented in a convenient form that can be used for numerical analysis and risk assessments. PMID:26729721

The effects of meteorological factors on the occurrence of Ganoderma sp. spores in the air

NASA Astrophysics Data System (ADS)

Grinn-Gofroń, Agnieszka; Strzelczak, Agnieszka

2011-03-01

Ganoderma sp. is an airborne fungal spore type known to trigger respiratory allergy symptoms in sensitive patients. Aiming to reduce the risk for allergic individuals, we analysed fungal spore circulation in Szczecin, Poland, and its dependence on meteorological conditions. Statistical models for the airborne spore concentrations of Ganoderma sp.—one of the most abundant fungal taxa in the area—were developed. Aerobiological sampling was conducted over 2004-2008 using a volumetric Lanzoni trap. Simultaneously, the following meteorological parameters were recorded: daily level of precipitation, maximum and average wind speed, relative humidity and maximum, minimum, average and dew point temperatures. These data were used as the explaining variables. Due to the non-linearity and non-normality of the data set, the applied modelling techniques were artificial neural networks (ANN) and mutlivariate regression trees (MRT). The obtained classification and MRT models predicted threshold conditions above which Ganoderma sp. appeared in the air. It turned out that dew point temperature was the main factor influencing the presence or absence of Ganoderma sp. spores. Further analysis of spore seasons revealed that the airborne fungal spore concentration depended only slightly on meteorological factors.

Identification of highly variable supernumerary chromosome segments in an asexual pathogen

USDA-ARS?s Scientific Manuscript database

Supernumerary chromosome segments are known to harbor different transposons from their essential counterparts. The aim of this study was to investigate the role of transposons in the origin and evolution of supernumerary segments in the asexual fungal pathogen Fusariumvirguliforme. We compared the g...

Loss of Sexual Reproduction and Dwarfing in a Small Metazoan

PubMed Central

Stelzer, Claus-Peter; Schmidt, Johanna; Wiedlroither, Anneliese; Riss, Simone

2010-01-01

Background Asexuality has major theoretical advantages over sexual reproduction, yet newly formed asexual lineages rarely endure. The success, or failure, of such lineages is affected by their mechanism of origin, because it determines their initial genetic makeup and variability. Most previously described mechanisms imply that asexual lineages are randomly frozen subsamples of a sexual population. Methodology/Principal Findings We found that transitions to obligate parthenogenesis (OP) in the rotifer Brachionus calyciflorus, a small freshwater invertebrate which normally reproduces by cyclical parthenogenesis, were controlled by a simple Mendelian inheritance. Pedigree analysis suggested that obligate parthenogens were homozygous for a recessive allele, which caused inability to respond to the chemical signals that normally induce sexual reproduction in this species. Alternative mechanisms, such as ploidy changes, could be ruled out on the basis of flow cytometric measurements and genetic marker analysis. Interestingly, obligate parthenogens were also dwarfs (approximately 50% smaller than cyclical parthenogens), indicating pleiotropy or linkage with genes that strongly affect body size. We found no adverse effects of OP on survival or fecundity. Conclusions/Significance This mechanism of inheritance implies that genes causing OP may evolve within sexual populations and remain undetected in the heterozygous state long before they get frequent enough to actually cause a transition to asexual reproduction. In this process, genetic variation at other loci might become linked to OP genes, leading to non-random associations between asexuality and other phenotypic traits. PMID:20862222

Development of a filter to prevent infections with spore-forming bacteria in injecting drug users.

PubMed

Alhusein, Nour; Scott, Jenny; Kasprzyk-Hordern, Barbara; Bolhuis, Albert

2016-12-01

In heroin injectors, there have been a number of outbreaks caused by spore-forming bacteria, causing serious infections such as anthrax or botulism. These are, most likely, caused by injecting contaminated heroin, and our aim was to develop a filter that efficiently removes these bacteria and is also likely to be acceptable for use by people who inject drugs (i.e. quick, simple and not spoil the hit). A prototype filter was designed and different filter membranes were tested to assess the volume of liquid retained, filtration time and efficiency of the filter at removing bacterial spores. Binding of active ingredients of heroin to different types of membrane filters was determined using a highly sensitive analytical chemistry technique. Heroin samples that were tested contained up to 580 bacteria per gramme, with the majority being Bacillus spp., which are spore-forming soil bacteria. To remove these bacteria, a prototype filter was designed to fit insulin-type syringes, which are commonly used by people who inject drugs (PWIDs). Efficient filtration of heroin samples was achieved by combining a prefilter to remove particles and a 0.22 μm filter to remove bacterial spores. The most suitable membrane was polyethersulfone (PES). This membrane had the shortest filtration time while efficiently removing bacterial spores. No or negligible amounts of active ingredients in heroin were retained by the PES membrane. This study successfully produced a prototype filter designed to filter bacterial spores from heroin samples. Scaled up production could produce an effective harm reduction tool, especially during outbreaks such as occurred in Europe in 2009/10 and 2012.

Chemical and Stress Resistances of Clostridium difficile Spores and Vegetative Cells

PubMed Central

Edwards, Adrianne N.; Karim, Samiha T.; Pascual, Ricardo A.; Jowhar, Lina M.; Anderson, Sarah E.; McBride, Shonna M.

2016-01-01

Clostridium difficile is a Gram-positive, sporogenic and anaerobic bacterium that causes a potentially fatal colitis. C. difficile enters the body as dormant spores that germinate in the colon to form vegetative cells that secrete toxins and cause the symptoms of infection. During transit through the intestine, some vegetative cells transform into spores, which are more resistant to killing by environmental insults than the vegetative cells. Understanding the inherent resistance properties of the vegetative and spore forms of C. difficile is imperative for the development of methods to target and destroy the bacterium. The objective of this study was to define the chemical and environmental resistance properties of C. difficile vegetative cells and spores. We examined vegetative cell and spore tolerances of three C. difficile strains, including 630Δerm, a 012 ribotype and a derivative of a past epidemic strain; R20291, a 027 ribotype and current epidemic strain; and 5325, a clinical isolate that is a 078 ribotype. All isolates were tested for tolerance to ethanol, oxygen, hydrogen peroxide, butanol, chloroform, heat and sodium hypochlorite (household bleach). Our results indicate that 630Δerm vegetative cells (630 spo0A) are more resistant to oxidative stress than those of R20291 (R20291 spo0A) and 5325 (5325 spo0A). In addition, 5325 spo0A vegetative cells exhibited greater resistance to organic solvents. In contrast, 630Δerm spores were more sensitive than R20291 or 5325 spores to butanol. Spores from all three strains exhibited high levels of resistance to ethanol, hydrogen peroxide, chloroform and heat, although R20291 spores were more resistant to temperatures in the range of 60–75°C. Finally, household bleach served as the only chemical reagent tested that consistently reduced C. difficile vegetative cells and spores of all tested strains. These findings establish conditions that result in vegetative cell and spore elimination and illustrate the resistance of C. difficile to common decontamination methods. These results further demonstrate that the vegetative cells and spores of various C. difficile strains have different resistance properties that may impact decontamination of surfaces and hands. PMID:27833595

Three new combinations in Drepanopeziza for species on poplar

USDA-ARS?s Scientific Manuscript database

Three species of Drepanopeziza that cause diseases of poplars have been known using scientific names for their sexual and asexual morphs, which is no longer allowed with the change to one scientific name for fungi. For each species, the oldest epithet is provided by the asexual morph; however, neith...

Potato (Solanum tuberosum) greenhouse tuber production as an assay for asexual reproduction effects from herbicides

EPA Science Inventory

The present study determined whether young potato plants can be used as an assay to indicate potential effects of pesticides on asexual reproduction. Solanum tuberosum (Russet Burbank) plants were grown from seed pieces in a mineral soil in pots under greenhouse conditions. Plant...

Combined pressure-thermal inactivation effect on spores in lu-wei beef--a traditional Chinese meat product.

PubMed

Wang, B-S; Li, B-S; Du, J-Z; Zeng, Q-X

2015-08-01

This study investigated the inactivation effect and kinetics of Bacillus coagulans and Geobacillus stearothermophilus spores suspended in lu-wei beef by combining high pressure (500 and 600 MPa) and moderate heat (70 and 80 °C or 80 and 90 °C). During pressurization, the temperature of pressure-transmitting fluid was tested with a K-type thermocouple, and the number of surviving cells was determined by a plate count method. The pressure come-up time and corresponding inactivation of Bacillus coagulans and G. stearothermophilus spores were considered during the pressure-thermal treatment. For the two types of spores, the results showed a higher inactivation effect in phosphate buffer solution than that in lu-wei beef. Among the bacteria evaluated, G. stearothermophilus spores had a higher resistance than B. coagulans spores during the pressure-thermal processing. One linear model and two nonlinear models (i.e. the Weibull and log-logistic models) were fitted to the survivor data to obtain relevant kinetic parameters, and the performance of these models was compared. The results suggested that the survival curve of the spores could be accurately described utilizing the log-logistic model, which produced the best fit for all inactivation data. The compression heating characteristics of different pressure-transmitting fluids should be considered when using high pressure to sterilize spores, particularly while the pressure is increasing. Spores can be inactivated by combining high pressure and moderate heat. The study demonstrates the synergistic inactivation effect of moderate heat in combination with high pressure in real-life food. The use of mathematical models to predict the inactivation for spores could help the food industry further to develop optimum process conditions. © 2015 The Society for Applied Microbiology.

Infection of Tribolium castaneum with Bacillus thuringiensis: Quantification of Bacterial Replication within Cadavers, Transmission via Cannibalism, and Inhibition of Spore Germination

PubMed Central

Milutinović, Barbara; Höfling, Christina; Futo, Momir; Scharsack, Jörn P.

2015-01-01

Reproduction within a host and transmission to the next host are crucial for the virulence and fitness of pathogens. Nevertheless, basic knowledge about such parameters is often missing from the literature, even for well-studied bacteria, such as Bacillus thuringiensis, an endospore-forming insect pathogen, which infects its hosts via the oral route. To characterize bacterial replication success, we made use of an experimental oral infection system for the red flour beetle Tribolium castaneum and developed a flow cytometric assay for the quantification of both spore ingestion by the individual beetle larvae and the resulting spore load after bacterial replication and resporulation within cadavers. On average, spore numbers increased 460-fold, showing that Bacillus thuringiensis grows and replicates successfully in insect cadavers. By inoculating cadaver-derived spores and spores from bacterial stock cultures into nutrient medium, we next investigated outgrowth characteristics of vegetative cells and found that cadaver-derived bacteria showed reduced growth compared to bacteria from the stock cultures. Interestingly, this reduced growth was a consequence of inhibited spore germination, probably originating from the host and resulting in reduced host mortality in subsequent infections by cadaver-derived spores. Nevertheless, we further showed that Bacillus thuringiensis transmission was possible via larval cannibalism when no other food was offered. These results contribute to our understanding of the ecology of Bacillus thuringiensis as an insect pathogen. PMID:26386058

Capability of the nematode-trapping fungus Duddingtonia flagrans to reduce infective larvae of gastrointestinal nematodes in goat feces in the southeastern United States: dose titration and dose time interval studies.

PubMed

Terrill, T H; Larsen, M; Samples, O; Husted, S; Miller, J E; Kaplan, R M; Gelaye, S

2004-04-15

Infection with gastrointestinal nematodes, particularly Haemonchus contortus, is a major constraint to goat production in the southeastern United States. Non-anthelmintic control alternatives are needed due to increasing resistance of these nematodes to available anthelmintics. Two studies were completed in Central Georgia in August 1999, and April-May 2000, using Spanish does naturally infected with Haemonchus contortus, Trichostongylus colubriformis, and Cooperia spp. to evaluate effectiveness of nematode-trapping fungi as a biological control agent. In the first experiment, five levels of Duddingtonia flagrans spores were mixed with a complete diet and fed once daily to the does (three per treatment) in metabolism crates. The treatment concentrations were (1) 5 x 10(5), (2) 2.5 x 10(5), (3) 10(5), and (4) 5 x 10(4) spores per kilogram body weight (BW), and (5) no spores. Fungal spores were fed for the first 7 days of the 14-day trial, and fecal samples were collected daily from individual animals for analysis of fecal egg count and establishment of fecal cultures. Efficacy of the fungus at reducing development of infective larvae (L3) in the fecal cultures was evaluated. The mean reduction in L3 from day 2 of the treatment period until the day after treatment stopped (days 2-8) was 93.6, 80.2, 84.1, and 60.8% for animals given the highest to lowest spore doses, respectively. Within 3-6 days after termination of fungal spore feedings, reduction in L3 development was no longer apparent in any of the treated animals. In a second experiment, effectiveness of 2.5 x 10(5) spores of D. flagrans per kilogram BW fed to does every day, every second day, and every third day was evaluated. Reduction in L3 development by daily feeding was less in the second experiment than in the first experiment. Daily fungal spore feeding provided more consistent larval reduction than intermittant feeding (every second or third day). When fed daily under controlled conditions, D. flagrans was effective in significantly reducing development of L3 and appears to be an effective tool for biocontrol of parasitic nematodes in goats.

A Global Survey of ATPase Activity in Plasmodium falciparum Asexual Blood Stages and Gametocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ortega, Corrie; Frando, Andrew; Webb-Robertson, Bobbie-Jo

Effective malaria control and elimination in hyperendemic areas of the world will require treatment of disease-causing Plasmodium falciparum (Pf) blood stage infection but also blocking parasite transmission from humans to mosquito to prevent disease spread. Numerous antimalarial drugs have become ineffective due to parasite drug resistance and many currently used therapies do not kill gametocytes, highly specialized sexual parasite stages with distinct physiology that are necessary for transmission from the human host to the mosquito vector. Further confounding next generation drug development against Pf is the lack of known biochemical activity for most parasite gene products as well as themore » unknown metabolic needs of non-replicating gametocyte. Here, we take a systematic activity-based proteomics approach to survey the large and druggable ATPase family that is associated with replicating blood stage asexual parasites and transmissible gametocytes. We experimentally confirm existing annotation and predict ATPase function for 38 uncharacterized proteins. ATPase activity broadly changes during the transition from asexual schizonts to gametocytes, indicating altered metabolism and regulatory roles of ATPases specific for each lifecycle stage. By mapping the activity of ATPases associated with gametocytogenesis, we assign biochemical activity to a large number of uncharacterized proteins and identify new candidate transmission blocking targets.« less

Plasmodium falciparum Calcium-Dependent Protein Kinase 2 Is Critical for Male Gametocyte Exflagellation but Not Essential for Asexual Proliferation

PubMed Central

Molina-Cruz, Alvaro; Brzostowski, Joseph; Mu, Jianbing

2017-01-01

ABSTRACT Drug development efforts have focused mostly on the asexual blood stages of the malaria parasite Plasmodium falciparum. Except for primaquine, which has its own limitations, there are no available drugs that target the transmission of the parasite to mosquitoes. Therefore, there is a need to validate new parasite proteins that can be targeted for blocking transmission. P. falciparum calcium-dependent protein kinases (PfCDPKs) play critical roles at various stages of the parasite life cycle and, importantly, are absent in the human host. These features mark them as attractive drug targets. In this study, using CRISPR/Cas9 we successfully knocked out PfCDPK2 from blood-stage parasites, which was previously thought to be an indispensable protein. The growth rate of the PfCDPK2 knockout (KO) parasites was similar to that of wild-type parasites, confirming that PfCDPK2 function is not essential for the asexual proliferation of the parasite in vitro. The mature male and female gametocytes of PfCDPK2 KO parasites become round after induction. However, they fail to infect female Anopheles stephensi mosquitoes due to a defect(s) in male gametocyte exflagellation and possibly in female gametes. PMID:29042501

Widespread occurrence of lysine methylation in Plasmodium falciparum proteins at asexual blood stages.

PubMed

Kaur, Inderjeet; Zeeshan, Mohammad; Saini, Ekta; Kaushik, Abhinav; Mohmmed, Asif; Gupta, Dinesh; Malhotra, Pawan

2016-10-20

Post-transcriptional and post-translational modifications play a major role in Plasmodium life cycle regulation. Lysine methylation of histone proteins is well documented in several organisms, however in recent years lysine methylation of proteins outside histone code is emerging out as an important post-translational modification (PTM). In the present study we have performed global analysis of lysine methylation of proteins in asexual blood stages of Plasmodium falciparum development. We immunoprecipitated stage specific Plasmodium lysates using anti-methyl lysine specific antibodies that immunostained the asexual blood stage parasites. Using liquid chromatography and tandem mass spectrometry analysis, 570 lysine methylated proteins at three different blood stages were identified. Analysis of the peptide sequences identified 605 methylated sites within 422 proteins. Functional classification of the methylated proteins revealed that the proteins are mainly involved in nucleotide metabolic processes, chromatin organization, transport, homeostatic processes and protein folding. The motif analysis of the methylated lysine peptides reveals novel motifs. Many of the identified lysine methylated proteins are also interacting partners/substrates of PfSET domain proteins as revealed by STRING database analysis. Our findings suggest that the protein methylation at lysine residues is widespread in Plasmodium and plays an important regulatory role in diverse set of the parasite pathways.

Mapping of Proteomic Composition on the Surfaces of Bacillus spores by Atomic Force Microscopy-based Immunolabeling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Plomp, M; Malkin, A J

2008-06-02

Atomic force microscopy provides a unique capability to image high-resolution architecture and structural dynamics of pathogens (e.g. viruses, bacteria and bacterial spores) at near molecular resolution in native conditions. Further development of atomic force microscopy in order to enable the correlation of pathogen protein surface structures with specific gene products is essential to understand the mechanisms of the pathogen life cycle. We have applied an AFM-based immunolabeling technique for the proteomic mapping of macromolecular structures through the visualization of the binding of antibodies, conjugated with nanogold particles, to specific epitopes on Bacillus spore surfaces. This information is generated while simultaneouslymore » acquiring the surface morphology of the pathogen. The immunospecificity of this labeling method was established through the utilization of specific polyclonal and monoclonal antibodies that target spore coat and exosporium epitopes of Bacillus atrophaeus and Bacillus anthracis spores.« less

INCIPIENT GERMINATION IN HEAVY SUSPENSIONS OF SPORES OF BACILLUS STEAROTHERMOPHILUS AT SUBMINIMAL GROWTH TEMPERATURES

PubMed Central

Curran, Harold R.; Pallansch, Michael J.

1963-01-01

Curran, Harold R. (U.S. Department of Agriculture, Washington, D.C.), and Michael J. Pallansch. Incipient germination in heavy suspensions of spores of Bacillus stearothermophilus at subminimal growth temperatures. J. Bacteriol. 86:911–918. 1963.—By use of spore (plate) counts and permeability to stain, labilization was followed periodically in heavy suspensions of washed Bacillus stearothermophilus 1518 spores incubated at different temperatures. Although vegetative proliferation did not occur below 38 C, incipient germination was rapid down to 20 C and much slower and incomplete at 14 C. Dilution of the suspension materially reduced the degree and rate of labilization. The degree of washing and use of deionized water had no appreciable influence upon early development of the spores. The results are discussed from the point of view of the possible origin and nature of the germination stimulant. Images PMID:14080801

Inactivation Strategies for Clostridium perfringens Spores and Vegetative Cells

PubMed Central

Talukdar, Prabhat K.; Udompijitkul, Pathima; Hossain, Ashfaque

2016-01-01

ABSTRACT Clostridium perfringens is an important pathogen to human and animals and causes a wide array of diseases, including histotoxic and gastrointestinal illnesses. C. perfringens spores are crucial in terms of the pathogenicity of this bacterium because they can survive in a dormant state in the environment and return to being live bacteria when they come in contact with nutrients in food or the human body. Although the strategies to inactivate C. perfringens vegetative cells are effective, the inactivation of C. perfringens spores is still a great challenge. A number of studies have been conducted in the past decade or so toward developing efficient inactivation strategies for C. perfringens spores and vegetative cells, which include physical approaches and the use of chemical preservatives and naturally derived antimicrobial agents. In this review, different inactivation strategies applied to control C. perfringens cells and spores are summarized, and the potential limitations and challenges of these strategies are discussed. PMID:27795314

Unikaryon phyllotretae sp. n. (Protista, Microspora), a new microsporidian pathogen of Phyllotreta undulata (Coleoptera; Chrysomelidae).

PubMed

Yaman, Mustafa; Radek, Renate; Weiser, Jaroslav; Toguebaye, Bhen Sikina

2010-01-01

The microsporidium Unikaryon phyllotretae sp. n., a new pathogen of Phyllotreta undulata, is described based on light microscopic and ultrastructural characteristics. Microscopic examination of parasitized individuals revealed two types of spores. The majority of the spores were of the first type, which are oval and measured 2.74+/-0.17 x 1.93+/-0.17 microm when fresh. Fresh spores of the second type (very rare) are elongated and measured 4.39+/-0.18 x 1.61+/-0.20 microm. All life stages have single nuclei. Sporogony ends with uninucleate single sporoblasts and spores. The spores were only observed in Malpighian tubules. The isofilar polar filament of the parasite has six to eight coils, and a well-developed polaroplast was of the lamellated type, with closely packed anterior lamellae and loosely packed posterior lamellae. Copyright (c) 2009. Published by Elsevier GmbH.

Longitudinal assessment of dairy farm management practices associated with the presence of psychrotolerant Bacillales spores in bulk tank milk on 10 New York State dairy farms.

PubMed

Masiello, S N; Kent, D; Martin, N H; Schukken, Y H; Wiedmann, M; Boor, K J

2017-11-01

The ability of certain spore-forming bacteria in the order Bacillales (e.g., Bacillus spp., Paenibacillus spp.) to survive pasteurization in spore form and grow at refrigeration temperatures results in product spoilage and limits the shelf life of high temperature, short time (HTST)-pasteurized fluid milk. To facilitate development of strategies to minimize contamination of raw milk with psychrotolerant Bacillales spores, we conducted a longitudinal study of 10 New York State dairy farms, which included yearlong monthly assessments of the frequency and levels of bulk tank raw milk psychrotolerant spore contamination, along with administration of questionnaires to identify farm management practices associated with psychrotolerant spore presence over time. Milk samples were first spore pasteurized (80°C for 12 min) and then analyzed for sporeformer counts on the initial day of spore pasteurization (SP), and after refrigerated storage (6°C) for 7, 14, and 21 d after SP. Overall, 41% of samples showed sporeformer counts of >20,000 cfu/mL at d 21, with Bacillus and Paenibacillus spp. being predominant causes of high sporeformer counts. Statistical analyses identified 3 management factors (more frequent cleaning of the bulk tank area, the use of a skid steer to scrape the housing area, and segregating problem cows during milking) that were all associated with lower probabilities of d-21 Bacillales spore detection in SP-treated bulk tank raw milk. Our data emphasize that appropriate on-farm measures to improve overall cleanliness and cow hygiene will reduce the probability of psychrotolerant Bacillales spore contamination of bulk tank raw milk, allowing for consistent production of raw milk with reduced psychrotolerant spore counts, which will facilitate production of HTST-pasteurized milk with extended refrigerated shelf life. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Computational fluid dynamics modeling of Bacillus anthracis spore deposition in rabbit and human respiratory airways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kabilan, S.; Suffield, S. R.; Recknagle, K. P.

Three-dimensional computational fluid dynamics and Lagrangian particle deposition models were developed to compare the deposition of aerosolized Bacillus anthracis spores in the respiratory airways of a human with that of the rabbit, a species commonly used in the study of anthrax disease. The respiratory airway geometries for each species were derived respectively from computed tomography (CT) and µCT images. Both models encompassed airways that extended from the external nose to the lung with a total of 272 outlets in the human model and 2878 outlets in the rabbit model. All simulations of spore deposition were conducted under transient, inhalation–exhalation breathingmore » conditions using average species-specific minute volumes. Two different exposure scenarios were modeled in the rabbit based upon experimental inhalation studies. For comparison, human simulations were conducted at the highest exposure concentration used during the rabbit experimental exposures. Results demonstrated that regional spore deposition patterns were sensitive to airway geometry and ventilation profiles. Due to the complex airway geometries in the rabbit nose, higher spore deposition efficiency was predicted in the nasal sinus compared to the human at the same air concentration of anthrax spores. In contrast, higher spore deposition was predicted in the lower conducting airways of the human compared to the rabbit lung due to differences in airway branching pattern. This information can be used to refine published and ongoing biokinetic models of inhalation anthrax spore exposures, which currently estimate deposited spore concentrations based solely upon exposure concentrations and inhaled doses that do not factor in species-specific anatomy and physiology for deposition.« less

Rapid concentration of Bacillus and Clostridium spores from large volumes of milk, using continuous flow centrifugation.

PubMed

Agoston, Réka; Soni, Kamlesh A; McElhany, Katherine; Cepeda, Martha L; Zuckerman, Udi; Tzipori, Saul; Mohácsi-Farkas, Csilla; Pillai, Suresh D

2009-03-01

Deliberate or accidental contamination of foods such as milk, soft drinks, and drinking water with infectious agents or toxins is a major concern to health authorities. There is a critical need to develop technologies that can rapidly and efficiently separate and concentrate biothreat agents from food matrices. A key limitation of current centrifugation and filtration technologies is that they are batch processes with extensive hands-on involvement and processing times. The objective of our studies was to evaluate the continuous flow centrifugation (CFC) technique for the rapid separation and concentration of bacterial spores from large volumes of milk. We determined the effectiveness of the CFC technology for concentrating approximately 10(3) bacterial spores in 3.7 liters (1 gal) of whole milk and skim milk, using Bacillus subtilis, Bacillus atrophaeus, and Clostridium sporogenes spores as surrogates for biothreat agents. The spores in the concentrated samples were enumerated by using standard plating techniques. Three independent experiments were performed at 10,000 rpm and 0.7 liters/min flow rate. The mean B. subtilis spore recoveries were 71.3 and 56.5% in skim and whole milk, respectively, and those for B. atrophaeus were 55 and 59.3% in skim and whole milk, respectively. In contrast, mean C. sporogenes spore recoveries were 88.2 and 78.6% in skim and whole milk, respectively. The successful use of CFC to concentrate these bacterial spores from 3.7 liters of milk in 10 min shows promise for rapidly concentrating other spores from large volumes of milk.

Detection of Bacillus spores using PCR and FTA filters.

PubMed

Lampel, Keith A; Dyer, Deanne; Kornegay, Leroy; Orlandi, Palmer A

2004-05-01

Emphasis has been placed on developing and implementing rapid detection systems for microbial pathogens. We have explored the utility of expanding FTA filter technology for the preparation of template DNA for PCR from bacterial spores. Isolated spores from several Bacillus spp., B. subtilis, B. cereus, and B. megaterium, were applied to FTA filters, and specific DNA products were amplified by PCR. Spore preparations were examined microscopically to ensure that the presence of vegetative cells, if any, did not yield misleading results. PCR primers SRM86 and SRM87 targeted a conserved region of bacterial rRNA genes, whereas primers Bsub5F and Bsub3R amplified a product from a conserved sequence of the B. subtilis rRNA gene. With the use of the latter set of primers for nested PCR, the sensitivity of the PCR-based assay was increased. Overall, 53 spores could be detected after the first round of PCR, and the sensitivity was increased to five spores by nested PCR. FTA filters are an excellent platform to remove PCR inhibitors and have universal applications for environmental, clinical, and food samples.

Aspergillosis in the common sea fan Gorgonia ventalina: isolation of waterborne hyphae and spores.

PubMed

Troeger, Victoria J; Sammarco, Paul W; Caruso, John H

2014-07-03

The octocoral disease aspergillosis is caused by the terrestrial fungus Aspergillus sydowii. The possibility of secondary (horizontal) transmission of aspergillosis among common sea fans Gorgonia ventalina would require waterborne transmission of hyphae and/or spores. A laboratory filtration experiment confirmed that fungal hyphae and spores were shed into the water by infected fans. This suggests that secondary infection might be possible in this species. It remains to be determined whether healthy fans actually develop aspergillosis after contact with hyphae-laden water.

Transmission rates of the bacterial endosymbiont, Neorickettsia risticii, during the asexual reproduction phase of its digenean host, Plagiorchis elegans, within naturally infected lymnaeid snails.

PubMed

Greiman, Stephen E; Tkach, Vasyl V; Vaughan, Jefferson A

2013-10-22

Neorickettsia are obligate intracellular bacterial endosymbionts of digenean parasites present in all lifestages of digeneans. Quantitative information on the transmission of neorickettsial endosymbionts throughout the complex life cycles of digeneans is lacking. This study quantified the transmission of Neorickettsia during the asexual reproductive phase of a digenean parasite, Plagiorchis elegans, developing within naturally parasitized lymnaeid pond snails. Lymnaea stagnalis snails were collected from 3 ponds in Nelson County, North Dakota and screened for the presence of digenean cercariae. Cercariae were identified to species by PCR and sequencing of the 28S rRNA gene. Neorickettsia infections were initially detected using nested PCR and sequencing of a partial 16S rRNA gene of pooled cercariae shed from each parasitized snail. Fifty to 100 single cercariae or sporocysts were isolated from each of six parasitized snails and tested for the presence of Neorickettsia using nested PCR to estimate the efficiency at which Neorickettsia were transmitted to cercariae during asexual development of the digenean. A total of 616 L. stagnalis were collected and 240 (39%) shed digenean cercariae. Of these, 18 (8%) were Neorickettsia-positive. Six Neorickettsia infections were selected to determine the transmission efficiency of Neorickettsia from mother to daughter sporocyst and from daughter sporocyst to cercaria. The prevalence of neorickettsiae in cercariae varied from 11 to 91%. The prevalence of neorickettsiae in sporocysts from one snail was 100%. Prevalence of Neorickettsia infection in cercariae of Plagiorchis elegans was variable and never reached 100%. Reasons for this are speculative, however, the low prevalence of Neorickettsia observed in some of our samples (11 to 52%) differs from the high prevalence of other, related bacterial endosymbionts, e.g. Wolbachia in Wolbachia-dependent filariid nematodes, where the prevalence among progeny is universally 100%. This suggests that, unlike the Wolbachia-filaria relationship, the Neorickettsia-digenean relationship is not obligatory mutualism. Our study represents the first quantitative estimate of the Neorickettsia transmission through the asexual phase of the digenean life cycle.

A simple culture method inducing sexual reproduction by Fusarium graminearum, the primary causal agent of Fusarium head blight

USDA-ARS?s Scientific Manuscript database

The homothallic ascomycete fungus Fusarium graminearum is the primary causal agent of Fusarium head blight (FHB), a devastating disease of wheat and barley worldwide. The fungus undergoes both asexual and sexual stages in its life cycle. The asexual stage produces conidiospores, whereas the sexual s...

75 FR 3711 - Listing Endangered and Threatened Wildlife and Designating Critical Habitat; 12-month...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-01-22

... recruitment, and successful reattachment and recruitment of asexual fragments in water depths shallower than 30 meters in four areas in Florida, Puerto Rico, and the U.S. Virgin Islands. The petition seeks to... settlement and recruitment, and the reattachment and recruitment of asexual fragments in water depths...

The pea aphid uses a version of the terminal system during oviparous, but not viviparous, development

PubMed Central

2013-01-01

Background In most species of aphid, female nymphs develop into either sexual or asexual adults depending on the length of the photoperiod to which their mothers were exposed. The progeny of these sexual and asexual females, in turn, develop in dramatically different ways. The fertilized oocytes of sexual females begin embryogenesis after being deposited on leaves (oviparous development) while the oocytes of asexual females complete embryogenesis within the mother (viviparous development). Compared with oviparous development, viviparous development involves a smaller transient oocyte surrounded by fewer somatic epithelial cells and a smaller early embryo that comprises fewer cells. To investigate whether patterning mechanisms differ between the earliest stages of the oviparous and viviparous modes of pea aphid development, we examined the expression of pea aphid orthologs of genes known to specify embryonic termini in other insects. Results Here we show that pea aphid oviparous ovaries express torso-like in somatic posterior follicle cells and activate ERK MAP kinase at the posterior of the oocyte. In addition to suggesting that some posterior features of the terminal system are evolutionarily conserved, our detection of activated ERK in the oocyte, rather than in the embryo, suggests that pea aphids may transduce the terminal signal using a mechanism distinct from the one used in Drosophila. In contrast with oviparous development, the pea aphid version of the terminal system does not appear to be used during viviparous development, since we did not detect expression of torso-like in the somatic epithelial cells that surround either the oocyte or the blastoderm embryo and we did not observe restricted activated ERK in the oocyte. Conclusions We suggest that while oviparous oocytes and embryos may specify posterior fate through an aphid terminal system, viviparous oocytes and embryos employ a different mechanism, perhaps one that does not rely on an interaction between the oocyte and surrounding somatic cells. Together, these observations provide a striking example of a difference in the fundamental events of early development that is both environmentally induced and encoded by the same genome. PMID:23552511

Effects of steam autoclave treatment on Geobacillus stearothermophilus spores.

PubMed

Huesca-Espitia, L C; Suvira, M; Rosenbeck, K; Korza, G; Setlow, B; Li, W; Wang, S; Li, Y-Q; Setlow, P

2016-11-01

To determine the mechanism of autoclave killing of Geobacillus stearothermophilus spores used in biological indicators (BIs) for steam autoclave sterilization, and rates of loss of spore viability and a spore enzyme used in BIs. Spore viability, dipicolinic acid (DPA) release, nucleic acid staining, α-glucosidase activity, protein structure and mutagenesis were measured during autoclaving of G. stearothermophilus spores. Loss of DPA and increases in spore core nucleic acid staining were slower than loss of spore viability. Spore core α-glucosidase was also lost more slowly than spore viability, although soluble α-glucosidase in spore preparations was lost more rapidly. However, spores exposed to an effective autoclave sterilization lost all viability and α-glucosidase activity. Apparently killed autoclaved spores were not recovered by artificial germination in supportive media, much spore protein was denatured during autoclaving, and partially killed autoclave-treated spore preparations did not acquire mutations. These results indicate that autoclave-killed spores cannot be revived, spore killing by autoclaving is likely by protein damage, and spore core α-glucosidase activity is lost more slowly than spore viability. This work provides insight into the mechanism of autoclave killing of spores of an organism used in BIs, and that a spore enzyme in a BI is more stable to autoclaving than spore viability. © 2016 The Society for Applied Microbiology.

SporeWeb: an interactive journey through the complete sporulation cycle of Bacillus subtilis.

PubMed

Eijlander, Robyn T; de Jong, Anne; Krawczyk, Antonina O; Holsappel, Siger; Kuipers, Oscar P

2014-01-01

Bacterial spores are a continuous problem for both food-based and health-related industries. Decades of scientific research dedicated towards understanding molecular and gene regulatory aspects of sporulation, spore germination and spore properties have resulted in a wealth of data and information. To facilitate obtaining a complete overview as well as new insights concerning this complex and tightly regulated process, we have developed a database-driven knowledge platform called SporeWeb (http://sporeweb.molgenrug.nl) that focuses on gene regulatory networks during sporulation in the Gram-positive bacterium Bacillus subtilis. Dynamic features allow the user to navigate through all stages of sporulation with review-like descriptions, schematic overviews on transcriptional regulation and detailed information on all regulators and the genes under their control. The Web site supports data acquisition on sporulation genes and their expression, regulon network interactions and direct links to other knowledge platforms or relevant literature. The information found on SporeWeb (including figures and tables) can and will be updated as new information becomes available in the literature. In this way, SporeWeb offers a novel, convenient and timely reference, an information source and a data acquisition tool that will aid in the general understanding of the dynamics of the complete sporulation cycle.

A combination of a SEM technique and X-ray microanalysis for studying the spore germination process of Clostridium tyrobutyricum.

PubMed

Bassi, Daniela; Cappa, Fabrizio; Cocconcelli, Pier Sandro

2009-06-01

Clostridium tyrobutyricum is an anaerobic bacterium responsible for late blowing defects during cheese ripening and it is of scientific interest for biological hydrogen production. A scanning electron microscopy (SEM) coating technique and X-ray microanalysis were developed to analyze the architecture and chemical composition of spores upon germination in response to environmental changes. In addition, we investigated the effects of different compounds on this process. Agents and environmental conditions inducing germination were characterized monitoring changes in optical density (OD). Among all tested conditions, the greatest drop in OD(625) (57.4%) was obtained when spores were incubated in l-alanine/l-lactate buffer, pH 4.6. In addition, a carbon-coating SEM technique and X-ray microanalysis were used to observe the architecture of spores and to examine calcium dipicolinate release. Conditions inducing C. tyrobutyricum spore germination were identified and SEM X-ray microanalysis clearly distinguished germinating from dormant spores. We confirmed that calcium dipicolinate release is one of the first events occurring. These microscopy methods could be considered sensitive tools for evaluating morphological and chemical changes in spores of C. tyrobutyricum during the initial phase of germination. Information gathered from this work may provide new data for further research on germination.

Identifying and Inactivating Bacterial Spores

NASA Technical Reports Server (NTRS)

Newcombe, David; Dekas, Anne; Venkateswaran, Kasthuri

2009-01-01

Problems associated with, and new strategies for, inactivating resistant organisms like Bacillus canaveralius (found at Kennedy Space Center during a survey of three NASA cleanrooms) have been defined. Identifying the particular component of the spore that allows its heightened resistance can guide the development of sterilization procedures that are targeted to the specific molecules responsible for resistance, while avoiding using unduly harsh methods that jeopardize equipment. The key element of spore resistance is a multilayered protein shell that encases the spore called the spore coat. The coat of the best-studied spore-forming microbe, B. subtilis, consists of at least 45 proteins, most of which are poorly characterized. Several protective roles for the coat are well characterized including resistance to desiccation, large toxic molecules, ortho-phthalaldehyde, and ultraviolet (UV) radiation. One important long-term specific goal is an improved sterilization procedure that will enable NASA to meet planetary protection requirements without a terminal heat sterilization step. This would support the implementation of planetary protection policies for life-detection missions. Typically, hospitals and government agencies use biological indicators to ensure the quality control of sterilization processes. The spores of B. canaveralius that are more resistant to osmotic stress would serve as a better biological indicator for potential survival than those in use currently.

Quantitative Method To Determine Sporicidal Decontamination of Building Surfaces by Gaseous Fumigants, and Issues Related to Laboratory-Scale Studies▿

PubMed Central

Rastogi, Vipin K.; Wallace, Lalena; Smith, Lisa S.; Ryan, Shawn P.; Martin, Blair

2009-01-01

Chlorine dioxide gas and vaporous hydrogen peroxide sterilant have been used in the cleanup of building interiors contaminated with spores of Bacillus anthracis. A systematic study, in collaboration with the U.S. Environmental Protection Agency, was jointly undertaken by the U.S. Army-Edgewood Chemical Biological Center to determine the sporicidal efficacies of these two fumigants on six building structural materials: carpet, ceiling tile, unpainted cinder block, painted I-beam steel, painted wallboard, and unpainted pinewood. Critical issues related to high-throughput sample processing and spore recovery from porous and nonporous surfaces included (i) the extraction of spores from complex building materials, (ii) the effects of titer challenge levels on fumigant efficacy, and (iii) the impact of bioburden inclusion on spore recovery from surfaces and spore inactivation. Small pieces (1.3 by 1.3 cm of carpet, ceiling tile, wallboard, I-beam steel, and pinewood and 2.5 by 1.3 cm for cinder block) of the materials were inoculated with an aliquot of 50 μl containing the target number (1 × 106, 1 × 107, or 1 × 108) of avirulent spores of B. anthracis NNR1Δ1. The aliquot was dried overnight in a biosafety cabinet, and the spores were extracted by a combination of a 10-min sonication and a 2-min vortexing using 0.5% buffered peptone water as the recovery medium. No statistically significant drop in the kill efficacies of the fumigants was observed when the spore challenge level was increased from 6 log units to 8 log units, even though a general trend toward inhibition of fumigant efficacy was evident. The organic burden (0 to 5%) in the spore inoculum resulted in a statistically significant drop in spore recovery (at the 2 or 5% level). The effect on spore killing was a function of the organic bioburden amount and the material type. In summary, a high-throughput quantitative method was developed for determining the efficacies of fumigants, and the spore recoveries from five porous materials and one nonporous material ranged between 20 and 80%. PMID:19346341

Quantitative method to determine sporicidal decontamination of building surfaces by gaseous fumigants, and issues related to laboratory-scale studies.

PubMed

Rastogi, Vipin K; Wallace, Lalena; Smith, Lisa S; Ryan, Shawn P; Martin, Blair

2009-06-01

Chlorine dioxide gas and vaporous hydrogen peroxide sterilant have been used in the cleanup of building interiors contaminated with spores of Bacillus anthracis. A systematic study, in collaboration with the U.S. Environmental Protection Agency, was jointly undertaken by the U.S. Army-Edgewood Chemical Biological Center to determine the sporicidal efficacies of these two fumigants on six building structural materials: carpet, ceiling tile, unpainted cinder block, painted I-beam steel, painted wallboard, and unpainted pinewood. Critical issues related to high-throughput sample processing and spore recovery from porous and nonporous surfaces included (i) the extraction of spores from complex building materials, (ii) the effects of titer challenge levels on fumigant efficacy, and (iii) the impact of bioburden inclusion on spore recovery from surfaces and spore inactivation. Small pieces (1.3 by 1.3 cm of carpet, ceiling tile, wallboard, I-beam steel, and pinewood and 2.5 by 1.3 cm for cinder block) of the materials were inoculated with an aliquot of 50 microl containing the target number (1 x 10(6), 1 x 10(7), or 1 x 10(8)) of avirulent spores of B. anthracis NNR1Delta1. The aliquot was dried overnight in a biosafety cabinet, and the spores were extracted by a combination of a 10-min sonication and a 2-min vortexing using 0.5% buffered peptone water as the recovery medium. No statistically significant drop in the kill efficacies of the fumigants was observed when the spore challenge level was increased from 6 log units to 8 log units, even though a general trend toward inhibition of fumigant efficacy was evident. The organic burden (0 to 5%) in the spore inoculum resulted in a statistically significant drop in spore recovery (at the 2 or 5% level). The effect on spore killing was a function of the organic bioburden amount and the material type. In summary, a high-throughput quantitative method was developed for determining the efficacies of fumigants, and the spore recoveries from five porous materials and one nonporous material ranged between 20 and 80%.

Is meiosis a fundamental cause of inviability among sexual and asexual plants and animals?

PubMed

Levitis, Daniel A; Zimmerman, Kolea; Pringle, Anne

2017-08-16

Differences in viability between asexually and sexually generated offspring strongly influence the selective advantage and therefore the prevalence of sexual reproduction (sex). However, no general principle predicts when sexual offspring will be more viable than asexual offspring. We hypothesize that when any kind of reproduction is based on a more complex cellular process, it will encompass more potential failure points, and therefore lower offspring viability. Asexual reproduction (asex) can be simpler than sex, when offspring are generated using only mitosis. However, when asex includes meiosis and meiotic restitution, gamete production is more complex than in sex. We test our hypothesis by comparing the viability of asexual and closely related sexual offspring across a wide range of plants and animals, and demonstrate that meiotic asex does result in lower viability than sex; without meiosis, asex is mechanistically simple and provides higher viability than sex. This phylogenetically robust pattern is supported in 42 of 44 comparisons drawn from diverse plants and animals, and is not explained by the other variables included in our model. Other mechanisms may impact viability, such as effects of reproductive mode on heterozygosity and subsequent viability, but we propose the complexity of cellular processes of reproduction, particularly meiosis, as a fundamental cause of early developmental failure and mortality. Meiosis, the leading cause of inviability in humans, emerges as a likely explanation of offspring inviability among diverse eukaryotes. © 2017 The Author(s).

Geographic parthenogenesis and plant-enemy interactions in the common dandelion.

PubMed

Verhoeven, Koen J F; Biere, Arjen

2013-01-28

Many species with sexual and asexual variants show a pattern of geographic parthenogenesis where asexuals have broader and higher-latitude distribution than sexuals. Because sexual reproduction is often considered a costly evolutionary strategy that is advantageous in the face of selection by coevolving pests and pathogens, one possible explanation for geographic parthenogenesis is that populations at higher latitudes are exposed to fewer pests and pathogens. We tested this hypothesis in the common dandelion (Taraxacum officinale), a species with well-established geographic parthenogenesis, by screening prevalence and effects of several specialized pests and pathogens in natural dandelion populations. We did a population survey of 18 dandelion populations along a geographic transect that ranged from the area where sexual and asexual dandelions co-occur northward into the area where only asexuals occur. In addition we used four southern and four northern populations in a 8x8 cross-inoculation greenhouse experiment in which plants were exposed experimentally to each other's natural field soil microbial communities. The cross-inoculation experiment indicated a higher pathogenicity of soil microbial communities from the southern, mostly sexual, populations compared to soil microbial communities from the northern asexual populations. Northern dandelion populations also showed reduced infestation by a specialized seed-eating weevil. A similar trend of reduced rust fungus infection in northern populations was observed but this trend was not statistically significant. The prevalence of pests and pathogens decreased along the south-to-north axis of geographic parthenogenesis. This highlights the potential of biotic interactions in shaping patterns of geographic parthenogenesis.

Development of Spore Protein of Myxobolus koi as an Immunostimulant for Prevent of Myxobolusis on Gold Fish (Cyprinus carpio Linn) by Oral Immunisation

NASA Astrophysics Data System (ADS)

Mahasri, Gunanti

2017-02-01

Production of Gold fish (Cyprinus carpio Linn) in Indonesia has always increased from 2013 to 2015 year by year with increasing average 2% per year. The amount of production was respectively 571.892 tonnes, 1129.273 tonnes, and 1186.674 tonnes. There were almost no problems to sale of gold fish because it had a good enough prospect. The aims of this research were Isolation of spore protein of Myxobolus koi by using SDS-PAGE to analyze immun respons and survival rate gold fish that immunized with spore protein of Myxobolus koi. The method of this research used experimental method, and belonged to 4 treatments that are: Controle = the group of gold fish not immunized with protein spore of Myxobolus koi neither infected by Myxobolus koi (T1). The group immunized and infested by spore of Myxobolus koi (T2), The group which immunized and not infested by Myxobolus koi (T3), and The group only infested by Myxobolus koi (T4). The dose of immunostimulant was 5 ml in 1 kg of food. The result showed that there were two bands of whole spore protein with molecule weight (MW) 150 kDa and 72 kDa and one band of crude protein Myxobolus koi with molecule weight 73 kD and the optical density point was 0.132 on the first day and increased to 0.769 on the 56 th day. The result also showed that the immun respons and survival rate increased from 27% to 86% in chellence test. The protein spore of Myxobolus koi can used to develops material for immunostimulant and to prevent the myxobolusis.

Comparative transcriptome analysis of obligately asexual and cyclically sexual rotifers reveals genes with putative functions in sexual reproduction, dormancy, and asexual egg production.

PubMed

Hanson, Sara J; Stelzer, Claus-Peter; Welch, David B Mark; Logsdon, John M

2013-06-19

Sexual reproduction is a widely studied biological process because it is critically important to the genetics, evolution, and ecology of eukaryotes. Despite decades of study on this topic, no comprehensive explanation has been accepted that explains the evolutionary forces underlying its prevalence and persistence in nature. Monogonont rotifers offer a useful system for experimental studies relating to the evolution of sexual reproduction due to their rapid reproductive rate and close relationship to the putatively ancient asexual bdelloid rotifers. However, little is known about the molecular underpinnings of sex in any rotifer species. We generated mRNA-seq libraries for obligate parthenogenetic (OP) and cyclical parthenogenetic (CP) strains of the monogonont rotifer, Brachionus calyciflorus, to identify genes specific to both modes of reproduction. Our differential expression analysis identified receptors with putative roles in signaling pathways responsible for the transition from asexual to sexual reproduction. Differential expression of a specific copy of the duplicated cell cycle regulatory gene CDC20 and specific copies of histone H2A suggest that such duplications may underlie the phenotypic plasticity required for reproductive mode switch in monogononts. We further identified differential expression of genes involved in the formation of resting eggs, a process linked exclusively to sex in this species. Finally, we identified transcripts from the bdelloid rotifer Adineta ricciae that have significant sequence similarity to genes with higher expression in CP strains of B. calyciflorus. Our analysis of global gene expression differences between facultatively sexual and exclusively asexual populations of B. calyciflorus provides insights into the molecular nature of sexual reproduction in rotifers. Furthermore, our results offer insight into the evolution of obligate asexuality in bdelloid rotifers and provide indicators important for the use of monogononts as a model system for investigating the evolution of sexual reproduction.

Evolution: sociality as a driver of unorthodox reproduction.

PubMed

Schwander, Tanja; Keller, Laurent

2012-07-10

An unusual reproductive system was discovered in desert ants, in which daughter queens are produced asexually via parthenogenesis, whereas workers develop from hybrid crosses between genetically divergent lineages. The system appears to be doomed to extinction. Copyright © 2012 Elsevier Ltd. All rights reserved.

Ethanol, vinegar and Origanum vulgare oil vapour suppress the development of anthracnose rot in tomato fruit.

PubMed

Tzortzakis, Nikos G

2010-08-15

Anthracnose rot (Colletotrichum coccodes) development in vitro or in tomato (Lycopersicon esculentum L.) fruit was evaluated after treatment with absolute ethyl alcohol (AEA), vinegar (VIN), chlorine (CHL) or origanum oil (ORI) and storage at 12 degrees C and 95% relative humidity during or following exposure to the volatiles. Fruit treated with vapours reduced fungal spore germination/production, but in the case of AEA- and VIN-treated fruits, fungal mycelium development was accelerated. Fruit lesion development was suppressed after fruit exposure to pure (100% v/v) AEA or ORI vapours which were accompanied by increased fruit cracking. Exposure to pure VIN-, CHL- and ORI vapours reduced (up to 92%) spore germination in vitro, but no differences were observed in the AEA treatment. The benefits associated with volatiles-enrichment were maintained in fruit pre-exposed to vapours, resulting in suppression in spore germination and spore production. However, studies performed on fungi grown on Potato Dextrose Agar revealed fewer direct effects of volatiles on fungal colony development and spore germination per se, implying that suppression of pathogen development was due in a large part to the impact of volatiles on fruit-pathogen interactions and/or 'memory' effects on fruit tissue. Work is currently focussing on the mechanisms underlying the impacts of volatiles on fruit quality related attributes. The results of this study indicate that volatiles may be considered as an alternative to the traditional postharvest sanitizing techniques. Each commodity needs to be individually assessed, and the volatile concentration and sanitising technique optimised, before the volatile treatment is used commercially. Copyright 2010 Elsevier B.V. All rights reserved.

A Quantitative Dynamic Simulation of Bremia lactucae Airborne Conidia Concentration above a Lettuce Canopy.

PubMed

Fall, Mamadou Lamine; Van der Heyden, Hervé; Carisse, Odile

2016-01-01

Lettuce downy mildew, caused by the oomycete Bremia lactucae Regel, is a major threat to lettuce production worldwide. Lettuce downy mildew is a polycyclic disease driven by airborne spores. A weather-based dynamic simulation model for B. lactucae airborne spores was developed to simulate the aerobiological characteristics of the pathogen. The model was built using the STELLA platform by following the system dynamics methodology. The model was developed using published equations describing disease subprocesses (e.g., sporulation) and assembled knowledge of the interactions among pathogen, host, and weather. The model was evaluated with four years of independent data by comparing model simulations with observations of hourly and daily airborne spore concentrations. The results show an accurate simulation of the trend and shape of B. lactucae temporal dynamics of airborne spore concentration. The model simulated hourly and daily peaks in airborne spore concentrations. More than 95% of the simulation runs, the daily-simulated airborne conidia concentration was 0 when airborne conidia were not observed. Also, the relationship between the simulated and the observed airborne spores was linear. In more than 94% of the simulation runs, the proportion of the linear variation in the hourly-observed values explained by the variation in the hourly-simulated values was greater than 0.7 in all years except one. Most of the errors came from the deviation from the 1:1 line, and the proportion of errors due to the model bias was low. This model is the only dynamic model developed to mimic the dynamics of airborne inoculum and represents an initial step towards improved lettuce downy mildew understanding, forecasting and management.

A Quantitative Dynamic Simulation of Bremia lactucae Airborne Conidia Concentration above a Lettuce Canopy

PubMed Central

Fall, Mamadou Lamine; Van der Heyden, Hervé; Carisse, Odile

2016-01-01

Lettuce downy mildew, caused by the oomycete Bremia lactucae Regel, is a major threat to lettuce production worldwide. Lettuce downy mildew is a polycyclic disease driven by airborne spores. A weather-based dynamic simulation model for B. lactucae airborne spores was developed to simulate the aerobiological characteristics of the pathogen. The model was built using the STELLA platform by following the system dynamics methodology. The model was developed using published equations describing disease subprocesses (e.g., sporulation) and assembled knowledge of the interactions among pathogen, host, and weather. The model was evaluated with four years of independent data by comparing model simulations with observations of hourly and daily airborne spore concentrations. The results show an accurate simulation of the trend and shape of B. lactucae temporal dynamics of airborne spore concentration. The model simulated hourly and daily peaks in airborne spore concentrations. More than 95% of the simulation runs, the daily-simulated airborne conidia concentration was 0 when airborne conidia were not observed. Also, the relationship between the simulated and the observed airborne spores was linear. In more than 94% of the simulation runs, the proportion of the linear variation in the hourly-observed values explained by the variation in the hourly-simulated values was greater than 0.7 in all years except one. Most of the errors came from the deviation from the 1:1 line, and the proportion of errors due to the model bias was low. This model is the only dynamic model developed to mimic the dynamics of airborne inoculum and represents an initial step towards improved lettuce downy mildew understanding, forecasting and management. PMID:26953691

Development and Application of a New Method for Specific and Sensitive Enumeration of Spores of Nonproteolytic Clostridium botulinum Types B, E, and F in Foods and Food Materials ▿

PubMed Central

Peck, Michael W.; Plowman, June; Aldus, Clare F.; Wyatt, Gary M.; Penaloza Izurieta, Walter; Stringer, Sandra C.; Barker, Gary C.

2010-01-01

The highly potent botulinum neurotoxins are responsible for botulism, a severe neuroparalytic disease. Strains of nonproteolytic Clostridium botulinum form neurotoxins of types B, E, and F and are the main hazard associated with minimally heated refrigerated foods. Recent developments in quantitative microbiological risk assessment (QMRA) and food safety objectives (FSO) have made food safety more quantitative and include, as inputs, probability distributions for the contamination of food materials and foods. A new method that combines a selective enrichment culture with multiplex PCR has been developed and validated to enumerate specifically the spores of nonproteolytic C. botulinum. Key features of this new method include the following: (i) it is specific for nonproteolytic C. botulinum (and does not detect proteolytic C. botulinum), (ii) the detection limit has been determined for each food tested (using carefully structured control samples), and (iii) a low detection limit has been achieved by the use of selective enrichment and large test samples. The method has been used to enumerate spores of nonproteolytic C. botulinum in 637 samples of 19 food materials included in pasta-based minimally heated refrigerated foods and in 7 complete foods. A total of 32 samples (5 egg pastas and 27 scallops) contained spores of nonproteolytic C. botulinum type B or F. The majority of samples contained <100 spores/kg, but one sample of scallops contained 444 spores/kg. Nonproteolytic C. botulinum type E was not detected. Importantly, for QMRA and FSO, the construction of probability distributions will enable the frequency of packs containing particular levels of contamination to be determined. PMID:20709854

Development and application of a new method for specific and sensitive enumeration of spores of nonproteolytic Clostridium botulinum types B, E, and F in foods and food materials.

PubMed

Peck, Michael W; Plowman, June; Aldus, Clare F; Wyatt, Gary M; Izurieta, Walter Penaloza; Stringer, Sandra C; Barker, Gary C

2010-10-01

The highly potent botulinum neurotoxins are responsible for botulism, a severe neuroparalytic disease. Strains of nonproteolytic Clostridium botulinum form neurotoxins of types B, E, and F and are the main hazard associated with minimally heated refrigerated foods. Recent developments in quantitative microbiological risk assessment (QMRA) and food safety objectives (FSO) have made food safety more quantitative and include, as inputs, probability distributions for the contamination of food materials and foods. A new method that combines a selective enrichment culture with multiplex PCR has been developed and validated to enumerate specifically the spores of nonproteolytic C. botulinum. Key features of this new method include the following: (i) it is specific for nonproteolytic C. botulinum (and does not detect proteolytic C. botulinum), (ii) the detection limit has been determined for each food tested (using carefully structured control samples), and (iii) a low detection limit has been achieved by the use of selective enrichment and large test samples. The method has been used to enumerate spores of nonproteolytic C. botulinum in 637 samples of 19 food materials included in pasta-based minimally heated refrigerated foods and in 7 complete foods. A total of 32 samples (5 egg pastas and 27 scallops) contained spores of nonproteolytic C. botulinum type B or F. The majority of samples contained <100 spores/kg, but one sample of scallops contained 444 spores/kg. Nonproteolytic C. botulinum type E was not detected. Importantly, for QMRA and FSO, the construction of probability distributions will enable the frequency of packs containing particular levels of contamination to be determined.

Infection of Tribolium castaneum with Bacillus thuringiensis: quantification of bacterial replication within cadavers, transmission via cannibalism, and inhibition of spore germination.

PubMed

Milutinović, Barbara; Höfling, Christina; Futo, Momir; Scharsack, Jörn P; Kurtz, Joachim

2015-12-01

Reproduction within a host and transmission to the next host are crucial for the virulence and fitness of pathogens. Nevertheless, basic knowledge about such parameters is often missing from the literature, even for well-studied bacteria, such as Bacillus thuringiensis, an endospore-forming insect pathogen, which infects its hosts via the oral route. To characterize bacterial replication success, we made use of an experimental oral infection system for the red flour beetle Tribolium castaneum and developed a flow cytometric assay for the quantification of both spore ingestion by the individual beetle larvae and the resulting spore load after bacterial replication and resporulation within cadavers. On average, spore numbers increased 460-fold, showing that Bacillus thuringiensis grows and replicates successfully in insect cadavers. By inoculating cadaver-derived spores and spores from bacterial stock cultures into nutrient medium, we next investigated outgrowth characteristics of vegetative cells and found that cadaver-derived bacteria showed reduced growth compared to bacteria from the stock cultures. Interestingly, this reduced growth was a consequence of inhibited spore germination, probably originating from the host and resulting in reduced host mortality in subsequent infections by cadaver-derived spores. Nevertheless, we further showed that Bacillus thuringiensis transmission was possible via larval cannibalism when no other food was offered. These results contribute to our understanding of the ecology of Bacillus thuringiensis as an insect pathogen. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Hot, humid air decontamination of a C-130 aircraft contaminated with spores of two acrystalliferous Bacillus thuringiensis strains, surrogates for Bacillus anthracis.

PubMed

Buhr, T L; Young, A A; Bensman, M; Minter, Z A; Kennihan, N L; Johnson, C A; Bohmke, M D; Borgers-Klonkowski, E; Osborn, E B; Avila, S D; Theys, A M G; Jackson, P J

2016-04-01

To develop test methods and evaluate survival of Bacillus thuringiensis kurstaki cry(-) HD-1 and B. thuringiensis Al Hakam spores after exposure to hot, humid air inside of a C-130 aircraft. Bacillus thuringiensis spores were either pre-inoculated on 1 × 2 or 2 × 2 cm substrates or aerosolized inside the cargo hold of a C-130 and allowed to dry. Dirty, complex surfaces (10 × 10 cm) swabbed after spore dispersal showed a deposition of 8-10 log10 m(-2) through the entire cargo hold. After hot, humid air decontamination at 75-80°C, 70-90% relative humidity for 7 days, 87 of 98 test swabs covering 0·98 m(2) , showed complete spore inactivation. There was a total of 1·67 log10 live CFU detected in 11 of the test swabs. Spore inactivation in the 98 test swabs was measured at 7·06 log10 m(-2) . Laboratory test methods for hot, humid air decontamination were scaled for a large-scale aircraft field test. The C-130 field test demonstrated that hot, humid air can be successfully used to decontaminate an aircraft. Transition of a new technology from research and development to acquisition at a Technology Readiness Level 7 is unprecedented. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

Early changes in arbuscular mycorrhiza development in sugarcane under two harvest management systems.

PubMed

de Azevedo, Lucas Carvalho Basilio; Stürmer, Sidney Luiz; Lambais, Marcio Rodrigues

2014-01-01

Sugarcane (Saccharum spp.) is grown on over 8 million ha in Brazil and is used to produce ethanol and sugar. Some sugarcane fields are burned to facilitate harvesting, which can affect the soil microbial community. However, whether sugarcane pre-harvest burning affects the community of arbuscular mycorrhizal fungi (AMF) and symbioses development is not known. In this study, we investigated the early impacts of harvest management on AMF spore communities and root colonization in three sugarcane varieties, under two harvest management systems (no-burning and pre-harvest burning). Soil and root samples were collected in the field after the first harvest of sugarcane varieties SP813250, SP801842, and RB72454, and AMF species were identified based on spore morphology. Diversity indices were determined based on spore populations and root colonization determined as an indicator of symbioses development. Based on the diversity indices, spore number and species occurrence in soil, no significant differences were observed among the AMF communities, regardless of harvest management type, sugarcane variety or interactions between harvest management type and sugarcane variety. However, mycorrhiza development was stimulated in sugarcane under the no-burning management system. Our data suggest that the sugarcane harvest management system may cause early changes in arbuscular mycorrhiza development.

Early changes in arbuscular mycorrhiza development in sugarcane under two harvest management systems

PubMed Central

de Azevedo, Lucas Carvalho Basilio; Stürmer, Sidney Luiz; Lambais, Marcio Rodrigues

2014-01-01

Sugarcane (Saccharum spp.) is grown on over 8 million ha in Brazil and is used to produce ethanol and sugar. Some sugarcane fields are burned to facilitate harvesting, which can affect the soil microbial community. However, whether sugarcane pre-harvest burning affects the community of arbuscular mycorrhizal fungi (AMF) and symbioses development is not known. In this study, we investigated the early impacts of harvest management on AMF spore communities and root colonization in three sugarcane varieties, under two harvest management systems (no-burning and pre-harvest burning). Soil and root samples were collected in the field after the first harvest of sugarcane varieties SP813250, SP801842, and RB72454, and AMF species were identified based on spore morphology. Diversity indices were determined based on spore populations and root colonization determined as an indicator of symbioses development. Based on the diversity indices, spore number and species occurrence in soil, no significant differences were observed among the AMF communities, regardless of harvest management type, sugarcane variety or interactions between harvest management type and sugarcane variety. However, mycorrhiza development was stimulated in sugarcane under the no-burning management system. Our data suggest that the sugarcane harvest management system may cause early changes in arbuscular mycorrhiza development. PMID:25477936

Comparative genomics of geographically distant Fusarium fujikuroi isolates revealed two distinct pathotypes correlating with secondary metabolite profiles

PubMed Central

Arndt, Birgit; Kalinina, Svetlana A.; Houterman, Petra M.; Ahn, Il-Pyung; Tonti, Stefano; Sieber, Christian M. K.

2017-01-01

Fusarium fujikuroi causes bakanae (“foolish seedling”) disease of rice which is characterized by hyper-elongation of seedlings resulting from production of gibberellic acids (GAs) by the fungus. This plant pathogen is also known for production of harmful mycotoxins, such as fusarins, fusaric acid, apicidin F and beauvericin. Recently, we generated the first de novo genome sequence of F. fujikuroi strain IMI 58289 combined with extensive transcriptional, epigenetic, proteomic and chemical product analyses. GA production was shown to provide a selective advantage during infection of the preferred host plant rice. Here, we provide genome sequences of eight additional F. fujikuroi isolates from distant geographic regions. The isolates differ in the size of chromosomes, most likely due to variability of subtelomeric regions, the type of asexual spores (microconidia and/or macroconidia), and the number and expression of secondary metabolite gene clusters. Whilst most of the isolates caused the typical bakanae symptoms, one isolate, B14, caused stunting and early withering of infected seedlings. In contrast to the other isolates, B14 produced no GAs but high amounts of fumonisins during infection on rice. Furthermore, it differed from the other isolates by the presence of three additional polyketide synthase (PKS) genes (PKS40, PKS43, PKS51) and the absence of the F. fujikuroi-specific apicidin F (NRPS31) gene cluster. Analysis of additional field isolates confirmed the strong correlation between the pathotype (bakanae or stunting/withering), and the ability to produce either GAs or fumonisins. Deletion of the fumonisin and fusaric acid-specific PKS genes in B14 reduced the stunting/withering symptoms, whereas deletion of the PKS51 gene resulted in elevated symptom development. Phylogenetic analyses revealed two subclades of F. fujikuroi strains according to their pathotype and secondary metabolite profiles. PMID:29073267

Label-free Chemical Imaging of Fungal Spore Walls by Raman Microscopy and Multivariate Curve Resolution Analysis

PubMed Central

Noothalapati, Hemanth; Sasaki, Takahiro; Kaino, Tomohiro; Kawamukai, Makoto; Ando, Masahiro; Hamaguchi, Hiro-o; Yamamoto, Tatsuyuki

2016-01-01

Fungal cell walls are medically important since they represent a drug target site for antifungal medication. So far there is no method to directly visualize structurally similar cell wall components such as α-glucan, β-glucan and mannan with high specificity, especially in a label-free manner. In this study, we have developed a Raman spectroscopy based molecular imaging method and combined multivariate curve resolution analysis to enable detection and visualization of multiple polysaccharide components simultaneously at the single cell level. Our results show that vegetative cell and ascus walls are made up of both α- and β-glucans while spore wall is exclusively made of α-glucan. Co-localization studies reveal the absence of mannans in ascus wall but are distributed primarily in spores. Such detailed picture is believed to further enhance our understanding of the dynamic spore wall architecture, eventually leading to advancements in drug discovery and development in the near future. PMID:27278218

Gravity and light control of the developmental polarity of regenerating protoplasts isolated from prothallial cells of the fern Ceratopteris richardii

NASA Technical Reports Server (NTRS)

Edwards, E. S.; Roux, S. J.

1998-01-01

A procedure has been developed for isolating protoplasts from prothalli of Ceratopteris richardii which can be cultured and are capable of regeneration. Protoplasts were isolated from 2-week-old gametophytes in a medium containing wall-digesting enzymes in 0.5 M sucrose, followed by purification of the released protoplasts by floating them up into a 0.5 M sorbitol layer. Regeneration occurred over a period of 10-24 days, and, under optimal osmotic conditions, followed the developmental pattern seen during spore germination, in that the first division gave rise to a primary rhizoid. Thus, prothallial protoplasts are comparable to germinating spores as suitable models for studies of developmental polarity in single cells. As in germinating spores, the polarity of development in regenerating protoplasts is influenced by the vectors of gravity and unilateral light. However, the relative influence of light in fixing this polarity is greater in regenerating protoplasts, while in germinating spores, the influence of gravity is greater.

Anti-Aspergillus Activities of the Respiratory Epithelium in Health and Disease.

PubMed

Bertuzzi, Margherita; Hayes, Gemma E; Icheoku, Uju J; van Rhijn, Norman; Denning, David W; Osherov, Nir; Bignell, Elaine M

2018-01-08

Respiratory epithelia fulfil multiple roles beyond that of gaseous exchange, also acting as primary custodians of lung sterility and inflammatory homeostasis. Inhaled fungal spores pose a continual antigenic, and potentially pathogenic, challenge to lung integrity against which the human respiratory mucosa has developed various tolerance and defence strategies. However, respiratory disease and immune dysfunction frequently render the human lung susceptible to fungal diseases, the most common of which are the aspergilloses, a group of syndromes caused by inhaled spores of Aspergillus fumigatus . Inhaled Aspergillus spores enter into a multiplicity of interactions with respiratory epithelia, the mechanistic bases of which are only just becoming recognized as important drivers of disease, as well as possible therapeutic targets. In this mini-review we examine current understanding of Aspergillus -epithelial interactions and, based upon the very latest developments in the field, we explore two apparently opposing schools of thought which view epithelial uptake of Aspergillus spores as either a curative or disease-exacerbating event.

Gravity-directed calcium current in germinating spores of Ceratopteris richardii

NASA Technical Reports Server (NTRS)

Chatterjee, A.; Porterfield, D. M.; Smith, P. S.; Roux, S. J.

2000-01-01

Gravity directs the early polar development in single cells of Ceratopteris richardii Brogn. It acts over a limited period of time during which it irreversibly determines the axis of the spore cell's development. A self-referencing calcium selective electrode was utilized to record the net movement of calcium across the cell membrane at different positions around the periphery of the spore during the period in which gravity orients the polarity of the spore. A movement of calcium into the cell along the bottom and out of the cell along the top was detected. This movement was specific, polarized, and strongest in a direction that opposed the vector of gravity. Treatment with nifedipine, a calcium-channel blocker, diminished the calcium current and caused the cell to lose its responsiveness to the orienting influence of gravity. Results shown suggest that calcium plays a crucial role in the ability of a single cell to respond to gravity and in the subsequent establishment of its polarity.

Molecular evidence for the first records of facultative parthenogenesis in elapid snakes

PubMed Central

Sanders, K. L.

2018-01-01

Parthenogenesis is a form of asexual reproduction by which embryos develop from unfertilized eggs. Parthenogenesis occurs in reptiles; however, it is not yet known to occur in the widespread elapid snakes (Elapidae), which include well-known taxa such as cobras, mambas, taipans and sea snakes. Here, we describe the production of viable parthenogens in two species of Australo-Papuan elapids with divergent reproductive modes: the oviparous coastal/Papuan taipan (Oxyuranus scutellatus) and the viviparous southern death adder (Acanthophis antarcticus). Analyses of nuclear SNP data excluded paternity for putative fathers and convincingly demonstrated asexual reproduction, thus representing the first evidence of facultative parthenogenesis in Elapidae. Our finding has broad implications for understanding the evolution of reproductive diversity in snakes, as well as managing the conservation of genetic diversity in wild and captive populations. PMID:29515892

Occurrence of Parthenogenesis in Potato Tuber Moth

PubMed Central

Liu, Yan; Hu, Chun-Hua; Wang, Chun-Ya; Xiong, Yan; Li, Zong-Kai; Xiao, Chun

2018-01-01

Abstract Parthenogenesis, a natural form of asexual reproduction produced from unfertilized eggs, occurs in many insects in Hemiptera and Hymenoptera, but very rarely in Lepidoptera. The current study aimed to test the larval density dependent occurrence of parthenogenesis in potato tuber moth, Phthorimaea operculella (Zeller; Lepidoptera: Gelechiidae) under laboratory conditions. More than 10% of females out of 25 tested females that developed from the high larval density treatment at 45 larvae per tuber were capable to reproduce asexually. Both male and female offspring were produced parthenogenetically. The sexually reproductive offspring of a laboratory parthenogenetic population had a lower egg hatch rate, shorter larval stage, and shorter male life span when compared with the non-parthenogenetic population. This suggests that the sexually reproductive offspring of parthenogenetic population have a decreased overall fitness compared to the sexually reproductive offspring of non-parthenogenetic population.

Molecular evidence for the first records of facultative parthenogenesis in elapid snakes.

PubMed

Allen, L; Sanders, K L; Thomson, V A

2018-02-01

Parthenogenesis is a form of asexual reproduction by which embryos develop from unfertilized eggs. Parthenogenesis occurs in reptiles; however, it is not yet known to occur in the widespread elapid snakes (Elapidae), which include well-known taxa such as cobras, mambas, taipans and sea snakes. Here, we describe the production of viable parthenogens in two species of Australo-Papuan elapids with divergent reproductive modes: the oviparous coastal/Papuan taipan ( Oxyuranus scutellatus ) and the viviparous southern death adder ( Acanthophis antarcticus ). Analyses of nuclear SNP data excluded paternity for putative fathers and convincingly demonstrated asexual reproduction, thus representing the first evidence of facultative parthenogenesis in Elapidae. Our finding has broad implications for understanding the evolution of reproductive diversity in snakes, as well as managing the conservation of genetic diversity in wild and captive populations.

Comparative analysis of immune effects in mice model: Clonorchis sinensis cysteine protease generated from recombinant Escherichia coli and Bacillus subtilis spores.

PubMed

Wu, Zhanshuai; Tang, Zeli; Shang, Mei; Zhao, Lu; Zhou, Lina; Kong, Xiangzhan; Lin, Zhipeng; Sun, Hengchang; Chen, Tingjin; Xu, Jin; Li, Xuerong; Huang, Yan; Yu, Xinbing

2017-07-01

Clonorchiasis remains a nonnegligible public health problem in endemic areas. Cysteine protease of Clonorchis sinensis (CsCP) plays indispensable roles in the parasitic physiology and pathology, and has been exploited as a promising drug and vaccine candidate. In recent years, development of spore-based vaccines against multiple pathogens has attracted many investigators' interest. In previous studies, the recombinant Escherichia coli (BL21) and Bacillus subtilis spores expressing CsCP have been successfully constructed, respectively. In this study, the immune effects of CsCP protein purified from recombinant BL21 (rCsCP) and B. subtilis spores presenting CsCP (B.s-CsCP) in Balb/c mice model were conducted with comparative analysis. Levels of specific IgG, IgG1 and IgG2a were significantly increased in sera from both rCsCP and B.s-CsCP intraperitoneally immunized mice. Additionally, recombinant spores expressing abundant fusion CsCP (0.03125 pg/spore) could strongly enhance the immunogenicity of CsCP with significantly higher levels of IgG and isotypes. Compared with rCsCP alone, intraperitoneal administration of mice with spores expressing CsCP achieved a better effect of fighting against C. sinensis infection by slowing down the process of fibrosis. Our results demonstrated that a combination of Th1/Th2 immune responses could be elicited by rCsCP, while spores displaying CsCP prominently induced Th1-biased specific immune responses, and the complex cytokine network maybe mediates protective immune responses against C. sinensis. This work further confirmed that the usage of B. subtilis spores displaying CsCP is an effective way to against C. sinensis.

Spore test parameters matter: Mesophilic and thermophilic spore counts detected in raw milk and dairy powders differ significantly by test method.

PubMed

Kent, D J; Chauhan, K; Boor, K J; Wiedmann, M; Martin, N H

2016-07-01

United States dairy industry exports have steadily risen in importance over the last 10yr, with dairy powders playing a particularly critical role. Currently, approximately half of US-produced nonfat dry milk and skim milk powder is exported. Reaching new and expanding existing export markets relies in part on the control of endospore-forming bacteria in dairy powders. This study reports baseline mesophilic and thermophilic spore counts and spore populations from 55 raw material samples (primarily raw milk) and 33 dairy powder samples from dairy powder processors across the United States. Samples were evaluated using various spore testing methodologies and included initial heat treatments of (1) 80°C for 12 min; (2) 100°C for 30 min; and (3) 106°C for 30 min. Results indicate that significant differences in both the level and population of spores were found for both raw milk and dairy powders with the various testing methods. Additionally, on average, spore counts were not found to increase significantly from the beginning to the end of dairy powder processing, most likely related to the absence of biofilm formation by processing plant-associated sporeformers (e.g., Anoxybacillus sp.) in the facilities sampled. Finally, in agreement with other studies, Bacillus licheniformis was found to be the most prevalent sporeformer in both raw materials and dairy powders, highlighting the importance of this organism in developing strategies for control and reduction of spore counts in dairy powders. Overall, this study emphasizes the need for standardization of spore enumeration methodologies in the dairy powder industry. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Detection of Biomass in New York City Aerosols: Light Scattering and Optical Fluorescence Techniques

NASA Astrophysics Data System (ADS)

Niebauer, M.; Alimova, A.; Katz, A.; Xu, M.; Rudolph, E.; Steiner, J.; Alfano, R. R.

2005-12-01

Optical spectroscopy is an ideal method for detecting bacteria and spores in real time. Optical fluorescence spectroscopy examination of New York City aerosols is used to quantify the mass of bacteria spores present in air masses collected at 14 liters/minute onto silica fiber filters, and on silica fiber ribbons using an Environmental Beta Attenuation Monitor manufactured by MetOne Instruments configured for the PM2.5 fraction. Dipicolinic acid (DPA), a molecule found primarily in bacterial spores, is the most characteristic component of spores in trial experiments on over 200 collected aerosol samples. DPA is extracted from the spores using a heat bath and chelated with Terbium. The DPA:Tb is detected by measuring its characteristic fluorescence with emission bands at 490, 545 and 585 nm for 270 nm excitation. Light scattering also measures the size distribution for a number of a variety of bacteria - Bacillus subtilis (rod shaped), Staphylococcus aureus (spherical) and Pseudomonas aeruginosa (short rods) establishing that optical techniques satisfactorily distinguish populations based on their variable morphology. Size and morphology are obtained by applying a variation of the Gaussian Ray Approximation theory of anomalous diffraction theory to an analysis of the transmission spectra in the range of 0.4 to 1.0 microns. In test experiments, the refractive index of the inner spore core of Bacillus subtilis decreases from 1.51 to 1.39 while the spore radius enlarges from 0.38 to 0.6 micrometers. Optical determinations are verified by oil-immersion techniques and by scanning electron microscope measurements. Characterization of spores, germinating spore materials, and bacteria is considered vital to tracing bacteria in the environment, for the development of life-detection systems for planetary exploration, monitoring pathogens in environmental systems, and for the preparation of anti-terrorism strategies.

The infrared spectral transmittance of Aspergillus niger spore aggregated particle swarm

NASA Astrophysics Data System (ADS)

Zhao, Xinying; Hu, Yihua; Gu, Youlin; Li, Le

2015-10-01

Microorganism aggregated particle swarm, which is quite an important composition of complex media environment, can be developed as a new kind of infrared functional materials. Current researches mainly focus on the optical properties of single microorganism particle. As for the swarm, especially the microorganism aggregated particle swarm, a more accurate simulation model should be proposed to calculate its extinction effect. At the same time, certain parameters deserve to be discussed, which helps to better develop the microorganism aggregated particle swarm as a new kind of infrared functional materials. In this paper, take Aspergillus Niger spore as an example. On the one hand, a new calculation model is established. Firstly, the cluster-cluster aggregation (CCA) model is used to simulate the structure of Aspergillus Niger spore aggregated particle. Secondly, the single scattering extinction parameters for Aspergillus Niger spore aggregated particle are calculated by using the discrete dipole approximation (DDA) method. Thirdly, the transmittance of Aspergillus Niger spore aggregated particle swarm is simulated by using Monte Carlo method. On the other hand, based on the model proposed above, what influences can wavelength causes has been studied, including the spectral distribution of scattering intensity of Aspergillus Niger spore aggregated particle and the infrared spectral transmittance of the aggregated particle swarm within the range of 8-14μm incident infrared wavelengths. Numerical results indicate that the scattering intensity of Aspergillus Niger spore aggregated particle reduces with the increase of incident wavelengths at each scattering angle. Scattering energy mainly concentrates on the scattering angle between 0-40°, forward scattering has an obvious effect. In addition, the infrared transmittance of Aspergillus Niger spore aggregated particle swarm goes up with the increase of incident wavelengths. However, some turning points of the trend are associated with the absorption capacity of the swarm. When parameters of the swarm are set as follows: each Aspergillus Niger spore aggregated particle contains 40 original particles, the radius of original particle is 1.5μm, the density of aggregated particles is around 200/cm3, the measurement area is 4 meters thick, under conditions mentioned above, the infrared transmittance can be less than 10% between the incident wavelengths of 9.5-13μm. In the end, all the results provide the basis for better developing the microorganism aggregated particle swarm as a new kind of infrared functional materials and precisely choosing the effective defiladed infrared band.

Competing sexual and asexual generic names in Pucciniomycotina and Ustilaginomycotina (Basidiomycota) and recommendations for use

USDA-ARS?s Scientific Manuscript database

With the change to one scientific name for fungi, generic names representing sexual and asexual morphs have been evaluated to determine which name to use when two names represent the same genus and thus compete for use. In this paper generic names of the Pucciniomycotina and Ustilaginomycotina are e...

Development of a user-friendly delivery method for the fungus Metarhizium anisopliac to control the ectoparasitic mite Varroa destructor in honey bee, Apis mellifera, colonies

USDA-ARS?s Scientific Manuscript database

A user-friendly method to deliver Metarhizium spores to honey bee colonies for control of Varroa mites was developed and tested. Patty blend formulations protected the fungal spores at brood nest temperatures and served as an improved delivery system of the fungus to bee hives. Field trials conducte...

The PsB glycoprotein complex is secreted as a preassembled precursor of the spore coat in Dictyostelium discoideum.

PubMed

Watson, N; McGuire, V; Alexander, S

1994-09-01

The PsB glycoprotein in Dictyostelium discoideum is one of a diverse group of developmentally regulated, prespore-cell-specific proteins, that contain a common O-linked oligosaccharide. This post-translational modification is dependent on the wild-type modB allele. The PsB protein exists as part of a multiprotein complex of six different proteins, which have different post-translational modifications and are held together by both covalent and non-covalent interactions (Watson et al. (1993). J. Biol. Chem. 268, 22634-22641). In this study we have used microscopic and biochemical analyses to examine the cellular localization and function of the PsB complex during development. We found that the PsB complex first accumulates in prespore vesicles in slug cells and is secreted later during culmination and becomes localized to both the extracellular matrix of the apical spore mass of mature fruiting bodies and to the inner layer of the spore coat. The PsB associated with the spore coat is covalently bound by disulfide bridges. The PsB protein always exists in a multiprotein complex, but the composition of the PsB complex changes during secretion and spore maturation. Some of the PsB complex proteins have been identified as spore coat proteins. These data demonstrate that some of the proteins that form the spore coat exist as a preassembled precursor complex. The PsB complex is secreted in a developmentally regulated manner during the process of spore differentiation, at which time proteins of the complex, as well as additional spore coat proteins, become covalently associated in at least two forms of extracellular matrix: the interspore matrix and the spore coat. These and other studies show that proteins with modB dependent O-linked oligosaccharides are involved in a wide variety of processes underlying morphogenesis in this organism. These developmental processes are the direct result of cellular mechanisms regulating protein targeting, assembly and secretion, and the assembly of specific extracellular matrices.

The impact of inducing germination of Bacillus anthracis and Bacillus thuringiensis spores on potential secondary decontamination strategies.

PubMed

Omotade, T O; Bernhards, R C; Klimko, C P; Matthews, M E; Hill, A J; Hunter, M S; Webster, W M; Bozue, J A; Welkos, S L; Cote, C K

2014-12-01

Decontamination and remediation of a site contaminated by the accidental or intentional release of fully virulent Bacillus anthracis spores are difficult, costly and potentially damaging to the environment. Development of novel decontamination strategies that have minimal environmental impacts remains a high priority. Although ungerminated spores are amongst the most resilient organisms known, once exposed to germinants, the germinating spores, in some cases, become susceptible to antimicrobial environments. We evaluated the concept that once germinated, B. anthracis spores would be less hazardous and significantly easier to remediate than ungerminated dormant spores. Through in vitro germination and sensitivity assays, we demonstrated that upon germination, B. anthracis Ames spores and Bacillus thuringiensis Al Hakam spores (serving as a surrogate for B. anthracis) become susceptible to environmental stressors. The majority of these germinated B. anthracis and B. thuringiensis spores were nonviable after exposure to a defined minimal germination-inducing solution for prolonged periods of time. Additionally, we examined the impact of potential secondary disinfectant strategies including bleach, hydrogen peroxide, formaldehyde and artificial UV-A, UV-B and UV-C radiation, employed after a 60-min germination-induction step. Each secondary disinfectant employs a unique mechanism of killing; as a result, germination-induction strategies are better suited for some secondary disinfectants than others. These results provide evidence that the deployment of an optimal combination strategy of germination-induction/secondary disinfection may be a promising aspect of wide-area decontamination following a B. anthracis contamination event. By inducing spores to germinate, our data confirm that the resulting cells exhibit sensitivities that can be leveraged when paired with certain decontamination measures. This increased susceptibility could be exploited to devise more efficient and safe decontamination measures and may obviate the need for more stringent methods that are currently in place. Published 2014. This article is a U.S. Government work and is in the public domain in the USA.

Removal of Bacillus anthracis sterne spore from commercial unpasteurized liquid egg white

USDA-ARS?s Scientific Manuscript database

Thermal pasteurization used by the egg industry for controlling vegetative cells of pathogens is ineffective for destroying endospores. There is a strong need in the agri-industries to develop effective intervention strategies to eliminate the possible bioterrorism threat from spore forming bacteria...

Mechanism of Bacillus subtilis spore inactivation by and resistance to supercritical CO2 plus peracetic acid.

PubMed

Setlow, B; Korza, G; Blatt, K M S; Fey, J P; Setlow, P

2016-01-01

Determine how supercritical CO2 (scCO2 ) plus peracetic acid (PAA) inactivates Bacillus subtilis spores, factors important in spore resistance to scCO2 -PAA, and if spores inactivated by scCO2 -PAA are truly dead. Spores of wild-type B. subtilis and isogenic mutants lacking spore protective proteins were treated with scCO2 -PAA in liquid or dry at 35°C. Wild-type wet spores (aqueous suspension) were more susceptible than dry spores. Treated spores were examined for viability (and were truly dead), dipicolinic acid (DPA), mutations, permeability to nucleic acid stains, germination under different conditions, energy metabolism and outgrowth. ScCO2 -PAA-inactivated spores retained DPA, and survivors had no notable DNA damage. However, DPA was released from inactivated spores at a normally innocuous temperature (85°C), and colony formation from treated spores was salt sensitive. The inactivated spores germinated but did not outgrow, and these germinated spores had altered plasma membrane permeability and defective energy metabolism. Wet or dry coat-defective spores had increased scCO2 -PAA sensitivity, and dry spores but not wet spores lacking DNA protective proteins were more scCO2 -PAA sensitive. These findings suggest that scCO2 -PAA inactivates spores by damaging spores' inner membrane. The spore coat provided scCO2 -PAA resistance for both wet and dry spores. DNA protective proteins provided scCO2 -PAA resistance only for dry spores. These results provide information on mechanisms of spore inactivation of and resistance to scCO2 -PAA, an agent with increasing use in sterilization applications. © 2015 The Society for Applied Microbiology.

Endotrophic Calcium, Strontium, and Barium Spores of Bacillus megaterium and Bacillus cereus1

PubMed Central

Foerster, Harold F.; Foster, J. W.

1966-01-01

Foerster, Harold F. (The University of Texas, Austin), and J. W. Foster. Endotrophic calcium, strontium, and barium spores of Bacillus megaterium and Bacillus cereus. J. Bacteriol. 91:1333–1345. 1966.—Spores were produced by washed vegetative cells suspended in deionized water supplemented with CaCl2, SrCl2, or BaCl2. Normal, refractile spores were produced in each case; a portion of the barium spores lost refractility and darkened. Thin-section electron micrographs revealed no apparent anatomical differences among the three types of spores. Analyses revealed that the different spore types were enriched specifically in the metal to which they were exposed during sporogenesis. The calcium content of the strontium and the barium spores was very small. From binary equimolar mixtures of the metal salts, endotrophic spores accumulated both metals to nearly the same extent. Viability of the barium spores was considerably less than that of the other two types. Strontium and barium spores were heat-resistant; however, calcium was essential for maximal heat resistance. Significant differences existed in the rates of germination; calcium spores germinated fastest, strontium spores were slower, and barium spores were slowest. Calcium-barium and calcium-strontium spores germinated readily. Endotrophic calcium and strontium spores germinated without the prior heat activation essential for growth spores. Chemical germination of the different metal-type spores with n-dodecylamine took place at the same relative rates as physiological germination. Heat-induced release of dipicolinic acid occurred much faster with barium and strontium spores than with calcium spores. The washed “coat fraction” from disrupted spores contained little of the spore calcium but most of the spore barium. The metal in this fraction was released by dilute acid. The demineralized coats reabsorbed calcium and barium at neutral pH. Images PMID:4956334

In vitro propagation of Cyathea atrovirens (Cyatheaceae): spore storage and sterilization conditions.

PubMed

Vargas, Isabel Beatriz de; Droste, Annette

2014-03-01

Cyathea atrovirens occurs in a wide range of habitats in Brazil, Paraguay, Uruguay and Argentina. In the Brazilian State of Rio Grande do Sul, this commonly found species is a target of intense exploitation, because of its ornamental characteristics. The in vitro culture is an important tool for propagation which may contribute toward the reduction of extractivism. However, exogenous contamination of spores is an obstacle for the success of aseptic long-term cultures. This study evaluated the influence of different sterilization methods combined with storage conditions on the contamination of the in vitro cultures and the gametophytic development of C. atrovirens, in order to establish an efficient propagation protocol. Spores were obtained from plants collected in Novo Hamburgo, State of Rio Grande do Sul, Brazil. In the first experiment, spores stored at 7 degrees C were surface sterilized with 0.5, 0.8 and 2% of sodium hypochlorite (NaClO) for 15 minutes and sown in Meyer's culture medium. The cultures were maintained in a growth room at 26 +/- 1 degrees C for a 12-h photoperiod and photon flux density of 100 micromol/m2/s provided by cool white fluorescent light. Contamination was assessed at 60 days, and gametophytic development was scored at 30, 60, 120 and 130 days of in vitro culture, analyzing 300 individuals for each treatment. There was no significant difference in culture contamination among the different sodium hypochlorite concentrations tested, and all treatments allowed for the development of cordiform gametophytes at 130 days of culture. In the second experiment, spores stored at 7 and -20 degrees C were divided into two groups. Half of the spores were surface sterilized with 2% of NaClO for 15 minutes and the other half was not sterilized. All spores were sown in Meyer's medium supplemented with one of the following antibiotics: nystatin, Micostatin and actidione. The culture conditions and the procedures used for evaluating contamination and gametophytic development were the same described for the first experiment. No contamination was observed in spores stored at -20 degrees C and treated with NaCIO and actidione. In all treatments, cordiform gametophytes presenting antheridia were observed at 120 days. The percentages of these gametophytes increased from 120 to 130 days and no significant differences were observed among treatments. Archegonia were observed on cordiform gametophytes at 130 days. The findings provide data relevant to in vitro propagation procedures of this species, which may increase the availability of plants for ornamental purposes, therefore contributing to the reduction of the exploitation of endangered tree ferns species.

The use of spore strips for monitoring the sterilization of bottled fluids.

PubMed Central

Selkon, J. B.; Sisson, P. R.; Ingham, H. R.

1979-01-01

A bacterial spore test has been developed which enables the efficacy of the sterilizing cycle recommended by the British Pharmaceutical Codex (1973) for bottled fluids to be accurately monitored. During a 14-month period this test detected faults in 3.3% of the sterilizing cycles, representing five distinct episodes of sterilization failure that passed unnoticed by the conventional controls of physical measurements and sterility testing. There were no failures of sterilization as detected by conventional techniques which were not indicated by the spore test. PMID:458140

Growth and sporulation of a pyrimidine spore color mutant of Sordaria fimicola.

PubMed

el-Ani, A S

1967-04-07

A nonautonomous spore color mutant of Sordaria fimicola is a pyrimidine auxotroph that produces hyaline nonviable ascospores. Uracil, uridine, and cytidine are more effective growth factors than cytosine and thymine and, in high concentrations, render the mutant self-fertile by inducing the ascospores to resume development and maturation. Crosses with the unlinked arginine non-autonomus spore color mutant st-59 yielded the double mutant st-59 pyr that requires both arginine and a pyrimidine for growth, which indicates a lack of suppression of the pyrimidine requirement by the arginine locus.

A Waking Review: Old and Novel Insights into the Spore Germination in Streptomyces.

PubMed

Bobek, Jan; Šmídová, Klára; Čihák, Matouš

2017-01-01

The complex development undergone by Streptomyces encompasses transitions from vegetative mycelial forms to reproductive aerial hyphae that differentiate into chains of single-celled spores. Whereas their mycelial life - connected with spore formation and antibiotic production - is deeply investigated, spore germination as the counterpoint in their life cycle has received much less attention. Still, germination represents a system of transformation from metabolic zero point to a new living lap. There are several aspects of germination that may attract our attention: (1) Dormant spores are strikingly well-prepared for the future metabolic restart; they possess stable transcriptome, hydrolytic enzymes, chaperones, and other required macromolecules stabilized in a trehalose milieu; (2) Germination itself is a specific sequence of events leading to a complete morphological remodeling that include spore swelling, cell wall reconstruction, and eventually germ tube emergences; (3) Still not fully unveiled are the strategies that enable the process, including a single cell's signal transduction and gene expression control, as well as intercellular communication and the probability of germination across the whole population. This review summarizes our current knowledge about the germination process in Streptomyces , while focusing on the aforementioned points.

Prophenoloxidase-Mediated Ex Vivo Immunity to Delay Fungal Infection after Insect Ecdysis.

PubMed

Zhang, Jie; Huang, Wuren; Yuan, Chuanfei; Lu, Yuzhen; Yang, Bing; Wang, Cheng-Yuan; Zhang, Peng; Dobens, Leonard; Zou, Zhen; Wang, Chengshu; Ling, Erjun

2017-01-01

Skin immunity protects animals from airborne pathogen infection. Unlike mammals, arthropods, including insects, undergo periodic ecdysis to grow and develop. Newly molted insects emerge with unsclerotized thin cuticles but successfully escape pathogenic infections during the post-molt period. Here we show that prophenoloxidases (PPOs) in molting fluids remain bioactive on the integument and impede fungal infection after ecdysis. We found that the purified plasma PPOs or recombinant PPOs could effectively bind to fungal spores (conidia) by targeting the cell wall components chitin and β-1,3-glucan. Pretreatment of the spores of the fungal pathogen Beauveria bassiana with PPOs increased spore hydrophilicity and reduced spore adhesion activity, resulting in a significant decrease in virulence as compared with mock infection. We also identified a spore-secreted protease BPS8, a member of peptidase S8 family of protease that degrade PPOs at high levels to benefit fungal infection, but which at lower doses activate PPOs to inhibit spore germination after melanization. These data indicate that insects have evolved a distinct strategy of ex vivo immunity to survive pathogen infections after ecdysis using PPOs in molting fluids retained on the underdeveloped and tender integument of newly molted insects for protection against airborne fungal infection.

Response of AM fungi spore population to elevated temperature and nitrogen addition and their influence on the plant community composition and productivity

PubMed Central

Zhang, Tao; Yang, Xue; Guo, Rui; Guo, Jixun

2016-01-01

To examine the influence of elevated temperature and nitrogen (N) addition on species composition and development of arbuscular mycorrhizal fungi (AMF) and the effect of AMF on plant community structure and aboveground productivity, we conducted a 5-year field experiment in a temperate meadow in northeast China and a subsequent greenhouse experiment. In the field experiment, N addition reduced spore population diversity and richness of AMF and suppressed the spore density and the hyphal length density (HLD). Elevated temperature decreased spore density and diameter and increased the HLD, but did not affect AMF spore population composition. In the greenhouse experiment, AMF altered plant community composition and increased total aboveground biomass in both elevated temperature and N addition treatments; additionally, AMF also increased the relative abundance and aboveground biomass of the grasses Leymus chinensis (Poaceae) and Setaria viridis (Gramineae) and significantly reduced the relative abundance and aboveground biomass of the Suaeda corniculata (Chenopodiaceae). Although elevated temperature and N addition can affect species composition or suppress the development of AMF, AMF are likely to play a vital role in increasing plant diversity and productivity. Notably, AMF might reduce the threat of climate change induced degradation of temperate meadow ecosystems. PMID:27098761

Response of AM fungi spore population to elevated temperature and nitrogen addition and their influence on the plant community composition and productivity

NASA Astrophysics Data System (ADS)

Zhang, Tao; Yang, Xue; Guo, Rui; Guo, Jixun

2016-04-01

To examine the influence of elevated temperature and nitrogen (N) addition on species composition and development of arbuscular mycorrhizal fungi (AMF) and the effect of AMF on plant community structure and aboveground productivity, we conducted a 5-year field experiment in a temperate meadow in northeast China and a subsequent greenhouse experiment. In the field experiment, N addition reduced spore population diversity and richness of AMF and suppressed the spore density and the hyphal length density (HLD). Elevated temperature decreased spore density and diameter and increased the HLD, but did not affect AMF spore population composition. In the greenhouse experiment, AMF altered plant community composition and increased total aboveground biomass in both elevated temperature and N addition treatments; additionally, AMF also increased the relative abundance and aboveground biomass of the grasses Leymus chinensis (Poaceae) and Setaria viridis (Gramineae) and significantly reduced the relative abundance and aboveground biomass of the Suaeda corniculata (Chenopodiaceae). Although elevated temperature and N addition can affect species composition or suppress the development of AMF, AMF are likely to play a vital role in increasing plant diversity and productivity. Notably, AMF might reduce the threat of climate change induced degradation of temperate meadow ecosystems.

A rapid and repeatable method to deposit bioaerosols on material surfaces.

PubMed

Calfee, M Worth; Lee, Sang Don; Ryan, Shawn P

2013-03-01

A simple method for repeatably inoculating surfaces with a precise quantity of aerosolized spores was developed. Laboratory studies were conducted to evaluate the variability of the method within and between experiments, the spatial distribution of spore deposition, the applicability of the method to complex surface types, and the relationship between material surface roughness and spore recoveries. Surface concentrations, as estimated by recoveries from wetted-wipe sampling, were between 5×10(3) and 1.5×10(4)CFUcm(-2) across the entire area (930cm(2)) inoculated. Between-test variability (Cv) in spore recoveries was 40%, 81%, 66%, and 20% for stainless steel, concrete, wood, and drywall, respectively. Within-test variability was lower, and did not exceed 33%, 47%, 52%, and 20% for these materials. The data demonstrate that this method is repeatable, is effective at depositing spores across a target surface area, and can be used to dose complex materials such as concrete, wood, and drywall. In addition, the data demonstrate that surface sampling recoveries vary by material type, and this variability can partially be explained by the material surface roughness index. This deposition method was developed for use in biological agent detection, sampling, and decontamination studies, however, is potentially beneficial to any scientific discipline that investigates surfaces containing aerosol-borne particles. Published by Elsevier B.V.

Inactivation Strategies for Clostridium perfringens Spores and Vegetative Cells.

PubMed

Talukdar, Prabhat K; Udompijitkul, Pathima; Hossain, Ashfaque; Sarker, Mahfuzur R

2017-01-01

Clostridium perfringens is an important pathogen to human and animals and causes a wide array of diseases, including histotoxic and gastrointestinal illnesses. C. perfringens spores are crucial in terms of the pathogenicity of this bacterium because they can survive in a dormant state in the environment and return to being live bacteria when they come in contact with nutrients in food or the human body. Although the strategies to inactivate C. perfringens vegetative cells are effective, the inactivation of C. perfringens spores is still a great challenge. A number of studies have been conducted in the past decade or so toward developing efficient inactivation strategies for C. perfringens spores and vegetative cells, which include physical approaches and the use of chemical preservatives and naturally derived antimicrobial agents. In this review, different inactivation strategies applied to control C. perfringens cells and spores are summarized, and the potential limitations and challenges of these strategies are discussed. Copyright © 2016 American Society for Microbiology.

Investigation of spore coat display of Bacillus subtilis β-galactosidase for developing of whole cell biocatalyst.

PubMed

Tavassoli, Setareh; Hinc, Krzysztof; Iwanicki, Adam; Obuchowski, Michal; Ahmadian, Gholamreza

2013-03-01

The production of highly efficient, recyclable and cost-effective enzymes is one of the most important goals in industrial biotechnology. Bacterial spores are highly resistant to harsh environmental conditions, easy to produce and are suitable for manipulation of genetic materials. These features make them a very efficient tool for biotechnology. Here, we show the use bacterial spores for presentation of functional enzyme. Spore coat display was used to produce a biocatalyst, which expresses β-galactiosidase (LacA). This enzyme is commonly used to produce lactose-free milk for lactose intolerant individuals. The lacA gene from Bacillus subtilis strain 168 was expressed on the surface of B. subtilis RH101(ΔcotC) spores using CotC as protein carrier. Presence of LacA protein is verified by western blotting. Results of β-galactiosidase assay show that the expressed enzyme retained its activity in condition of freezing and drying, as well as after recovery from the reaction's mixture.

Spontaneous hybrids between native and exotic Rubus in the Western United States produce offspring both by apomixis and by sexual recombination

PubMed Central

Clark, L V; Jasieniuk, M

2012-01-01

Facultative asexual reproduction is a trait commonly found in invasive species. With a combination of sexual and asexual reproductive modes, such species may adapt to new environments via sexual recombination during range expansion, while at the same time having the benefits of asexuality such as the maintenance of fitness effects that depend upon heterozygosity. In the Western United States, native species of Rubus (Rosaceae) reproduce sexually whereas exotic naturalized Rubus species reproduce by pseudogamous apomixis. We hypothesized that new asexual lineages of Rubus could arise from hybridization in this range. To detect hybridization between native and exotic Rubus, we genotyped 579 individuals collected across California, Oregon and Washington with eight nuclear microsatellites and two chloroplast markers. Principal Coordinate Analysis and Bayesian clustering revealed a limited amount of hybridization of the native R. ursinus with the exotic R. armeniacus and R. pensilvanicus, as well as cultivated varieties. Genetic distances between these hybrids and their offspring indicated that both R. ursinus × R. armeniacus and R. ursinus × R. pensilvanicus produced a mix of apomictic and sexual seeds, with sexual seeds being more viable. Although neither of these hybrid types is currently considered invasive, they model the early stages of evolution of new invasive lineages, given the potential for fixed heterosis and the generation of novel genotypes. The hybrids also retain the ability to increase their fitness via sexual recombination and natural selection. Mixed reproductive systems such as those described here may be an important step in the evolution of asexual invasive species. PMID:22850699

The role of sexual vs. asexual recruitment of Artemisia wudanica in transition zone habitats between inter-dune lowlands and active dunes in Inner Mongolia, China

NASA Astrophysics Data System (ADS)

Wang, Yongcui; Alberto, Busso Carlos; Jiang, Deming; Ala, Musa; Li, Xuehua; Zhou, Quanlai; Lin, Jixiang; Ren, Guohua; Jia, Lian

2016-04-01

Artemisia wudanica is an endemic, perennial, pioneering psammophyte species in the sand dune ecosystems of western Horqin Sand Land in northern China. However, no studies have addressed how sexual and asexual reproduction modes of A. wudanica perform at the transitional zones between active dune inter-dune lowlands and active dunes. In early spring, quadrats were randomly set up in the study area to monitor surviving seedling and/or ramet density and frequency coming from sexual/asexual reproduction of A. wudanica. Iron sticks were inserted near each quadrat to determine wind erosion intensity (WE). Additionally, soil samples were collected nearby each quadrat to test for soil moisture (SM), organic matter (OM) and pH. Surviving seedlings of A. wudanica showed an inverse response in comparison with ramets to SM, OM and WE. Soil moisture showed the most positive effect, and WE the negative effect, on surviving, sexual reproduction seedlings. Contrarily, WE had the most positive effect, and SM the negative effect, on asexual reproduction ramets. This suggests that increases in SM and decreases in WE should benefit recruitment of A. wudanica seedlings. On the contrary, ramets coming from asexual reproduction showed a different response to environmental factors in transition zone habitats. While SM was not a key constraint for the survival of seedlings, they showed a better, positive response to wind erosion environments. Overall, various study environmental parameters could be improved to foster A. wudanica invasion and settlement in the plant community through different reproductive modes, thereby promoting vegetation restoration and rehabilitation.

Geographic parthenogenesis and plant-enemy interactions in the common dandelion

PubMed Central

2013-01-01

Background Many species with sexual and asexual variants show a pattern of geographic parthenogenesis where asexuals have broader and higher-latitude distribution than sexuals. Because sexual reproduction is often considered a costly evolutionary strategy that is advantageous in the face of selection by coevolving pests and pathogens, one possible explanation for geographic parthenogenesis is that populations at higher latitudes are exposed to fewer pests and pathogens. We tested this hypothesis in the common dandelion (Taraxacum officinale), a species with well-established geographic parthenogenesis, by screening prevalence and effects of several specialized pests and pathogens in natural dandelion populations. Results We did a population survey of 18 dandelion populations along a geographic transect that ranged from the area where sexual and asexual dandelions co-occur northward into the area where only asexuals occur. In addition we used four southern and four northern populations in a 8x8 cross-inoculation greenhouse experiment in which plants were exposed experimentally to each other’s natural field soil microbial communities. The cross-inoculation experiment indicated a higher pathogenicity of soil microbial communities from the southern, mostly sexual, populations compared to soil microbial communities from the northern asexual populations. Northern dandelion populations also showed reduced infestation by a specialized seed-eating weevil. A similar trend of reduced rust fungus infection in northern populations was observed but this trend was not statistically significant. Conclusions The prevalence of pests and pathogens decreased along the south-to-north axis of geographic parthenogenesis. This highlights the potential of biotic interactions in shaping patterns of geographic parthenogenesis. PMID:23356700

Mechanism of Bacillus subtilis Spore Inactivation by and Resistance to Supercritical CO2 plus Peracetic Acid

PubMed Central

Setlow, Barbara; Korza, George; Blatt, Kelly M.S.; Fey, Julien P.; Setlow, Peter

2015-01-01

Aims Determine how supercritical CO2 (scCO2) plus peracetic acid (PAA) inactivates Bacillus subtilis spores, factors important in spore resistance to scCO2-PAA, and if spores inactivated by scCO2-PAA are truly dead. Methods and Results Spores of wild-type B. subtilis and isogenic mutants lacking spore protective proteins were treated with scCO2-PAA in liquid or dry at 35°C. Wild-type wet spores (aqueous suspension) were more susceptible than dry spores. Treated spores were examined for viability (and were truly dead), dipicolinic acid (DPA), mutations, permeability to nucleic acid stains, germination under different conditions, energy metabolism and outgrowth. ScCO2-PAA-inactivated spores retained DPA, and survivors had no notable DNA damage. However, DPA was released from inactivated spores at a normally innocuous temperature (85°C), and colony formation from treated spores was salt sensitive. The inactivated spores germinated but did not outgrow, and these germinated spores had altered plasma membrane permeability and defective energy metabolism. Wet or dry coat-defective spores had increased scCO2-PAA sensitivity, and dry spores but not wet spores lacking DNA protective proteins were more scCO2-PAA sensitive. Conclusions These findings suggest that scCO2-PAA inactivates spores by damaging spores’ inner membrane. The spore coat provided scCO2-PAA resistance for both wet and dry spores. DNA protective proteins provided scCO2-PAA resistance only for dry spores. Significance and Impact of Study These results provide information on mechanisms of spore inactivation of and resistance to scCO2-PAA, an agent with increasing use in sterilization applications. PMID:26535794

PfCDPK1 is critical for malaria parasite gametogenesis and mosquito infection

PubMed Central

Bansal, Abhisheka; Molina-Cruz, Alvaro; Brzostowski, Joseph; Liu, Poching; Luo, Yan; Gunalan, Karthigayan; Li, Yuesheng; Ribeiro, José M. C.; Miller, Louis H.

2018-01-01

Efforts to knock out Plasmodium falciparum calcium-dependent protein kinase 1 (PfCDPK1) from asexual erythrocytic stage have not been successful, indicating an indispensable role of the enzyme in asexual growth. We recently reported generation of a transgenic parasite with mutant CDPK1 [Bansal A, et al. (2016) MBio 7:e02011-16]. The mutant CDPK1 (T145M) had reduced activity of transphosphorylation. We reasoned that CDPK1 could be disrupted in the mutant parasites. Consistent with this assumption, CDPK1 was successfully disrupted in the mutant parasites using CRISPR/Cas9. We and others could not disrupt PfCDPK1 in the WT parasites. The CDPK1 KO parasites show a slow growth rate compared with the WT and the CDPK1 T145M parasites. Additionally, the CDPK1 KO parasites show a defect in both male and female gametogenesis and could not establish an infection in mosquitoes. Complementation of the KO parasite with full-length PfCDPK1 partially rescued the asexual growth defect and mosquito infection. Comparative global transcriptomics of WT and the CDPK1 KO schizonts using RNA-seq show significantly high transcript expression of gametocyte-specific genes in the CDPK1 KO parasites. This study conclusively demonstrates that CDPK1 is a good target for developing transmission-blocking drugs. PMID:29311293

PfCDPK1 is critical for malaria parasite gametogenesis and mosquito infection.

PubMed

Bansal, Abhisheka; Molina-Cruz, Alvaro; Brzostowski, Joseph; Liu, Poching; Luo, Yan; Gunalan, Karthigayan; Li, Yuesheng; Ribeiro, José M C; Miller, Louis H

2018-01-23

Efforts to knock out Plasmodium falciparum calcium-dependent protein kinase 1 ( Pf CDPK1) from asexual erythrocytic stage have not been successful, indicating an indispensable role of the enzyme in asexual growth. We recently reported generation of a transgenic parasite with mutant CDPK1 [Bansal A, et al. (2016) MBio 7:e02011-16]. The mutant CDPK1 (T145M) had reduced activity of transphosphorylation. We reasoned that CDPK1 could be disrupted in the mutant parasites. Consistent with this assumption, CDPK1 was successfully disrupted in the mutant parasites using CRISPR/Cas9. We and others could not disrupt Pf CDPK1 in the WT parasites. The CDPK1 KO parasites show a slow growth rate compared with the WT and the CDPK1 T145M parasites. Additionally, the CDPK1 KO parasites show a defect in both male and female gametogenesis and could not establish an infection in mosquitoes. Complementation of the KO parasite with full-length Pf CDPK1 partially rescued the asexual growth defect and mosquito infection. Comparative global transcriptomics of WT and the CDPK1 KO schizonts using RNA-seq show significantly high transcript expression of gametocyte-specific genes in the CDPK1 KO parasites. This study conclusively demonstrates that CDPK1 is a good target for developing transmission-blocking drugs. Copyright © 2018 the Author(s). Published by PNAS.

Sexual Plasticity and Self-Fertilization in the Sea Anemone Aiptasia diaphana

PubMed Central

Schlesinger, Ami; Kramarsky-Winter, Esti; Rosenfeld, Hanna; Armoza-Zvoloni, Rachel; Loya, Yossi

2010-01-01

Traits that influence reproductive success and contribute to reproductive isolation in animal and plant populations are a central focus of evolutionary biology. In the present study we used an experimental approach to demonstrate the occurrence of environmental effects on sexual and asexual reproduction, and provide evidence for sexual plasticity and inter-clonal fertilization in laboratory-cultured lines of the sea anemone Aiptasia diaphana. We showed that in A. diaphana, both asexual reproduction by pedal laceration, and sexual reproduction have seasonal components. The rate of pedal laceration was ten-fold higher under summer photoperiod and water temperature conditions than under winter conditions. The onset of gametogenesis coincided with the rising water temperatures occurring in spring, and spawning occurred under parameters that emulated summer photoperiod and temperature conditions. In addition, we showed that under laboratory conditions, asexually produced clones derived from a single founder individual exhibit sexual plasticity, resulting in the development of both male and female individuals. Moreover, a single female founder produced not only males and females but also hermaphrodite individuals. We further demonstrated that A. diaphana can fertilize within and between clone lines, producing swimming planula larvae. These diverse reproductive strategies may explain the species success as invader of artificial marine substrates. We suggest that these diverse reproductive strategies, together with their unique evolutionary position, make Aiptasia diaphana an excellent model for studying the evolution of sex. PMID:20686700

Health hazards related to conidia of Cladosporium-biological air pollutants in Poland, central Europe.

PubMed

Weryszko-Chmielewska, Elzbieta; Kasprzyk, Idalia; Nowak, Malgorzata; Sulborska, Aneta; Kaczmarek, Joanna; Szymanska, Agata; Haratym, Weronika; Gilski, Miroslaw; Jedryczka, Malgorzata

2018-03-01

The spores of Cladosporium Link. are often present in the air in high quantities and produce many allergenic proteins, which may lead to asthma. An aerobiological spore monitoring program can inform patients about the current spore concentration in air and help their physicians determine the spore dose that is harmful for a given individual. This makes it possible to develop optimized responses and propose personalized therapy for a particular sensitive patient. The aim of this study was to assess the extent of the human health hazard posed by the fungal genus Cladosporium. For the first time, we have determined the number of days on which air samples in Poland exceeded the concentrations linked to allergic responses of sensitive patients, according to thresholds established by three different groups (2800/3000/4000 spores per 1m 3 of the air). The survey was conducted over three consecutive growing seasons (April-September, 2010-2012) in three cities located in different climate zones of Poland (Poznan, Lublin and Rzeszow). The average number of days exceeding 2800 spores per cubic meter (the lowest threshold) ranged from 61 (2010) through 76 (2011) to 93 (2012), though there was significant variation between cities. In each year the highest concentration of spores in the air was detected in either Poznan or Lublin, both located on large plains with intensive agriculture. We have proposed that an effective, science-based software platform to support policy-making on air quality should incorporate biological air pollutant data, such as allergenic fungal spores and pollen grains. Copyright © 2017. Published by Elsevier B.V.

Computational Fluid Dynamics Modeling of Bacillus anthracis Spore Deposition in Rabbit and Human Respiratory Airways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kabilan, Senthil; Suffield, Sarah R.; Recknagle, Kurtis P.

Three-dimensional computational fluid dynamics and Lagrangian particle deposition models were developed to compare the deposition of aerosolized Bacillus anthracis spores in the respiratory airways of a human with that of the rabbit, a species commonly used in the study of anthrax disease. The respiratory airway geometries for each species were derived from computed tomography (CT) or µCT images. Both models encompassed airways that extended from the external nose to the lung with a total of 272 outlets in the human model and 2878 outlets in the rabbit model. All simulations of spore deposition were conducted under transient, inhalation-exhalation breathing conditionsmore » using average species-specific minute volumes. The highest exposure concentration was modeled in the rabbit based upon prior acute inhalation studies. For comparison, human simulation was also conducted at the same concentration. Results demonstrated that regional spore deposition patterns were sensitive to airway geometry and ventilation profiles. Due to the complex airway geometries in the rabbit nose, higher spore deposition efficiency was predicted in the upper conducting airways compared to the human at the same air concentration of anthrax spores. As a result, higher particle deposition was predicted in the conducting airways and deep lung of the human compared to the rabbit lung due to differences in airway branching pattern. This information can be used to refine published and ongoing biokinetic models of inhalation anthrax spore exposures, which currently estimate deposited spore concentrations based solely upon exposure concentrations and inhaled doses that do not factor in species-specific anatomy and physiology.« less

Cytometric Approach for Detection of Encephalitozoon intestinalis, an Emergent Agent▿

PubMed Central

Barbosa, Joana; Rodrigues, Acácio Gonçalves; Pina-Vaz, Cidália

2009-01-01

Encephalitozoon intestinalis is responsible for intestinal disease in patients with AIDS and immunocompetent patients. The infectious form is a small spore that is resistant to water treatment procedures. Its detection is very important, but detection is very cumbersome and time-consuming. Our main objective was to develop and optimize a specific flow cytometric (FC) protocol for the detection of E. intestinalis in hospital tap water and human feces. To determine the optimal specific antibody (Microspor-FA) concentration, a known concentration of E. intestinalis spores (Waterborne, Inc.) was suspended in hospital tap water and stool specimens with different concentrations of Microspor-FA, and the tap water and stool specimens were incubated under different conditions. The sensitivity limit and specificity were also evaluated. To study spore infectivity, double staining with propidium iodide (PI) and Microspor-FA was undertaken. Distinct approaches for filtration and centrifugation of the stool specimens were used. E. intestinalis spores stained with 10 μg/ml of Microspor-FA at 25°C overnight provided the best results. The detection limit was 5 × 104 spores/ml, and good specificity was demonstrated. Simultaneous staining with Microspor-FA and PI ensured that the E. intestinalis spores were dead and therefore noninfectious. With the stool specimens, better spore recovery was observed with a saturated solution of NaCl and centrifugation at 1,500 × g for 15 min. A new approach for the detection of E. intestinalis from tap water or human feces that ensures that the spores are not viable is now available and represents an important step for the prevention of this threat to public health. PMID:19439525

Sensitive, Rapid Detection of Bacterial Spores

NASA Technical Reports Server (NTRS)

Kern, Roger G.; Venkateswaran, Kasthuri; Chen, Fei; Pickett, Molly; Matsuyama, Asahi

2009-01-01

A method of sensitive detection of bacterial spores within delays of no more than a few hours has been developed to provide an alternative to a prior three-day NASA standard culture-based assay. A capability for relatively rapid detection of bacterial spores would be beneficial for many endeavors, a few examples being agriculture, medicine, public health, defense against biowarfare, water supply, sanitation, hygiene, and the food-packaging and medical-equipment industries. The method involves the use of a commercial rapid microbial detection system (RMDS) that utilizes a combination of membrane filtration, adenosine triphosphate (ATP) bioluminescence chemistry, and analysis of luminescence images detected by a charge-coupled-device camera. This RMDS has been demonstrated to be highly sensitive in enumerating microbes (it can detect as little as one colony-forming unit per sample) and has been found to yield data in excellent correlation with those of culture-based methods. What makes the present method necessary is that the specific RMDS and the original protocols for its use are not designed for discriminating between bacterial spores and other microbes. In this method, a heat-shock procedure is added prior to an incubation procedure that is specified in the original RMDS protocols. In this heat-shock procedure (which was also described in a prior NASA Tech Briefs article on enumerating sporeforming bacteria), a sample is exposed to a temperature of 80 C for 15 minutes. Spores can survive the heat shock, but nonspore- forming bacteria and spore-forming bacteria that are not in spore form cannot survive. Therefore, any colonies that grow during incubation after the heat shock are deemed to have originated as spores.

A Spore Coat Protein, CotS, of Bacillus subtilis Is Synthesized under the Regulation of ςK and GerE during Development and Is Located in the Inner Coat Layer of Spores

PubMed Central

Takamatsu, Hiromu; Chikahiro, Yukari; Kodama, Takeko; Koide, Hidekatsu; Kozuka, Satoshi; Tochikubo, Kunio; Watabe, Kazuhito

1998-01-01

The spore coat of Bacillus subtilis has a unique morphology and consists of polypeptides of different sizes, whose synthesis and assembly are precisely regulated by a cascade of transcription factors and regulatory proteins. We examined the factors that regulate cotS gene expression and CotS assembly into the coat layer of B. subtilis by Northern blot and Western blot analysis. Transcription of cotS mRNA was not detected in sporulating cells of ςK and gerE mutants by Northern blot analysis. By Western blot analysis using anti-CotS antibody, CotS was first detected in protein samples solubilized from wild-type cells at 5 h after the start of sporulation. CotS was not detected in the vegetative cells and spores of a gerE mutant or in the spores of mutants deficient in ςE, ςF, ςG, or ςK. CotS was detected in the sporangium but not in the spores of a cotE mutant. The sequence of the promoter region of cotS was similar to the consensus sequences for binding of ςK and GerE. These results demonstrate that ςK and GerE are required for cotS expression and that CotE is essential for the assembly of CotS in the coat. Immunoelectron microscopic observation using anti-CotS antibody revealed that CotS is located within the spore coat, in particular in the inner coats of dormant spores. PMID:9603889

Discourses Governing Lesbian, Gay, Bisexual, Transgender, Queer, Intersex, and Asexual Teachers' Disclosure of Sexual Orientation and Gender History

ERIC Educational Resources Information Center

Bower-Phipps, Laura

2017-01-01

Significant progress has been made in equal rights for lesbian, gay, bisexual, transgender, queer, intersex, and asexual (LGBTQIA) individuals, yet schools remain institutions where sexual and gender diversity are marginalized and/or silenced. Queer theory, a non-linear theory that disrupts dominant beliefs about gender and sexuality and what…

In Vitro Activities of Primaquine-Schizonticide Combinations on Asexual Blood Stages and Gametocytes of Plasmodium falciparum

PubMed Central

Cabrera, Mynthia

2015-01-01

Currently, the World Health Organization recommends addition of a 0.25-mg base/kg single dose of primaquine (PQ) to artemisinin combination therapies (ACTs) for Plasmodium falciparum malaria as a gametocytocidal agent for reducing transmission. Here, we investigated the potential interactions of PQ with the long-lasting components of the ACT drugs for eliminating the asexual blood stages and gametocytes of in vitro-cultured P. falciparum strains. Using the SYBR green I assay for asexual parasites and a flow cytometry-based assay for gametocytes, we determined the interactions of PQ with the schizonticides chloroquine, mefloquine, piperaquine, lumefantrine, and naphthoquine. With the sums of fractional inhibitory concentrations and isobolograms, we were able to determine mostly synergistic interactions for the various PQ and schizonticide combinations on the blood stages of P. falciparum laboratory strains. The synergism in inhibiting asexual stages and gametocytes was highly evident with PQ-naphthoquine, whereas synergism was moderate for the PQ-piperaquine, PQ-chloroquine, and PQ-mefloquine combinations. We have detected potentially antagonistic interactions between PQ and lumefantrine under certain drug combination ratios, suggesting that precautions might be needed when PQ is added as the gametocytocide to the artemether-lumefantrine ACT (Coartem). PMID:26416869

Genome structure of bdelloid rotifers: shaped by asexuality or desiccation?

PubMed

Gladyshev, Eugene A; Arkhipova, Irina R

2010-01-01

Bdelloid rotifers are microscopic invertebrate animals best known for their ancient asexuality and the ability to survive desiccation at any life stage. Both factors are expected to have a profound influence on their genome structure. Recent molecular studies demonstrated that, although the gene-rich regions of bdelloid genomes are organized as colinear pairs of closely related sequences and depleted in repetitive DNA, subtelomeric regions harbor diverse transposable elements and horizontally acquired genes of foreign origin. Although asexuality is expected to result in depletion of deleterious transposons, only desiccation appears to have the power to produce all the uncovered genomic peculiarities. Repair of desiccation-induced DNA damage would require the presence of a homologous template, maintaining colinear pairs in gene-rich regions and selecting against insertion of repetitive DNA that might cause chromosomal rearrangements. Desiccation may also induce a transient state of competence in recovering animals, allowing them to acquire environmental DNA. Even if bdelloids engage in rare or obscure forms of sexual reproduction, all these features could still be present. The relative contribution of asexuality and desiccation to genome organization may be clarified by analyzing whole-genome sequences and comparing foreign gene and transposon content in species which lost the ability to survive desiccation.

First real-time observation of transverse division in azooxanthellate scleractinian corals

PubMed Central

Tokuda, Yuki; Haraguchi, Hiroko; Ezaki, Yoichi

2017-01-01

Asexual reproduction is one of the most important traits in the evolutionary history of corals. No real-time observations of asexual reproduction in azooxanthellate solitary scleractinian corals have been conducted to date. Here, we describe previously unknown aspects of asexual reproduction by using Truncatoflabellum spheniscus (Family Flabellidae) based on observations of transverse division conducted over 1200 days. The findings revealed that (1) transverse division was caused by decalcification; (2) compared to the anthocyathus (upper part of the divided corallum), the soft parts of the anthocaulus (lower part of the divided corallum) were severely damaged and injured during division; (3) these injuries were repaired rapidly; and (4) the anthocaulus regrew and repeatedly produced anthocyathi by means of transverse division. Differences in the patterns of soft-part regeneration and repair, as well as differences in skeletal growth rates between the anthocaulus and the anthocyathus imply that the ecological requirements and reproductive success are different from each other immediately after division. The findings provide important clues for unravelling why asexual reproduction appeared frequently in free-living corals, and the extent to which those modes of reproduction has affected the adaptive and evolutionary success of scleractinian corals throughout the Phanerozoic. PMID:28150745

First real-time observation of transverse division in azooxanthellate scleractinian corals

NASA Astrophysics Data System (ADS)

Tokuda, Yuki; Haraguchi, Hiroko; Ezaki, Yoichi

2017-02-01

Asexual reproduction is one of the most important traits in the evolutionary history of corals. No real-time observations of asexual reproduction in azooxanthellate solitary scleractinian corals have been conducted to date. Here, we describe previously unknown aspects of asexual reproduction by using Truncatoflabellum spheniscus (Family Flabellidae) based on observations of transverse division conducted over 1200 days. The findings revealed that (1) transverse division was caused by decalcification; (2) compared to the anthocyathus (upper part of the divided corallum), the soft parts of the anthocaulus (lower part of the divided corallum) were severely damaged and injured during division; (3) these injuries were repaired rapidly; and (4) the anthocaulus regrew and repeatedly produced anthocyathi by means of transverse division. Differences in the patterns of soft-part regeneration and repair, as well as differences in skeletal growth rates between the anthocaulus and the anthocyathus imply that the ecological requirements and reproductive success are different from each other immediately after division. The findings provide important clues for unravelling why asexual reproduction appeared frequently in free-living corals, and the extent to which those modes of reproduction has affected the adaptive and evolutionary success of scleractinian corals throughout the Phanerozoic.

Genetic diversity of Toxoplasma gondii in animals and humans

USDA-ARS?s Scientific Manuscript database

Toxoplasma gondii is one of the most common parasites of domestic, wild, and companion animals, and it also infects approximately 25% of the world’s human population. T. gondii has a complex life cycle. Sexual development occurs only in the cat gut, while asexual replication and transmission occur i...

Wellness for All: Improving Campus Climate for LGBTQA Students as Prevention

ERIC Educational Resources Information Center

Manning, Patricia; Pring, Lauren; Glider, Peggy

2014-01-01

While it is widely recognized that campus climate can have a profound impact on students' academic performance, mental/behavioral health, and character development, campus is not experienced equally across identity groups. For lesbian, gay, bisexual, transgender, queer, questioning, and asexual (LGBTQA) students, campus often includes added…

Inhibition of Ageratina adenophora on spore germination and gametophyte development of Macrothelypteris torresiana.

PubMed

Zhang, Kai-Mei; Shi, Lei; Jiang, Chuang-Dao; Li, Zhen-Yu

2008-05-01

Allelopathy of Ageratina adenophora plays an important role in its invasion. However, we have little knowledge of its allelpathic effects on ferns. In Petri dish bioassays, the inhibitory potential of aqueous leachates from roots, stems and leaves of A. adenophora was studied on the spore germination and gametophyte development of Macrothelypteris torresiana. All leachates inhibited the spore germination and growth of the first rhizoid of M. torresiana and inhibitory effects increased with increasing leachate concentrations. Root leachates proved most inhibitory. Gametophyte rhizoids of M. torresiana treated with stem and leaf leachates of A. adenophora were erect, which was similar to those of the control. However, gametophyte rhizoids of M. torresiana treated with root leachates of A. adenophora were erect, but also curving or swollen. Moreover, curving and swollen rhizoids increased with increasing concentrations. As time went by, rhizoids treated with root leachates were not so curved and the swelling almost disappeared. Possible causes are discussed in the present study. The increasing concentrations of leaf leachates also delayed the stages of gametophyte development. With the treatment of root leachates, the delay was more obvious. Thus A. adenophora inhibited the spore germination and gametophyte development of M. torresiana and the root leachates were most inhibitory.

Multi-Probe Investigation of Proteomic Structure of Pathogens

DOE Office of Scientific and Technical Information (OSTI.GOV)

Malkin, A J; Plomp, M; Leighton, T J

Complete genome sequences are available for understanding biotransformation, environmental resistance and pathogenesis of microbial, cellular and pathogen systems. The present technological and scientific challenges are to unravel the relationships between the organization and function of protein complexes at cell, microbial and pathogens surfaces, to understand how these complexes evolve during the bacterial, cellular and pathogen life cycles, and how they respond to environmental changes, chemical stimulants and therapeutics. In particular, elucidating the molecular structure and architecture of human pathogen surfaces is essential to understanding mechanisms of pathogenesis, immune response, physicochemical interactions, environmental resistance and development of countermeasures against bioterrorist agents.more » The objective of this project was to investigate the architecture, proteomic structure, and function of bacterial spores through a combination of high-resolution in vitro atomic force microscopy (AFM) and AFM-based immunolabeling with threat-specific antibodies. Particular attention in this project was focused on spore forming Bacillus species including the Sterne vaccine strain of Bacillus anthracis and the spore forming near-neighbor of Clostridium botulinum, C. novyi-NT. Bacillus species, including B. anthracis, the causative agent of inhalation anthrax are laboratory models for elucidating spore structure/function. Even though the complete genome sequence is available for B. subtilis, cereus, anthracis and other species, the determination and composition of spore structure/function is not understood. Prof. B. Vogelstein and colleagues at the John Hopkins University have recently developed a breakthrough bacteriolytic therapy for cancer treatment (1). They discovered that intravenously injected Clostridium novyi-NT spores germinate exclusively within the avascular regions of tumors in mice and destroy advanced cancerous lesions. The bacteria were also found to significantly improve the efficacy of chemotherapeutic drugs and radiotherapy (2,3). Currently, there is no understanding of the structure-function relationships of Clostridium novyi-NT spores. As well as their therapeutic interest, studies of Clostridium noyii spores could provide a model for further studies of human pathogenic spore formers including Clostridium botulinum and Clostridium perfringens. This project involved a multi-institutional collaboration of our LLNL group with the groups of Prof. T.J. Leighton (Children's Hospital Oakland Research Institute) and Prof. B. Vogelstein (The Howard Hughes Medical Institute and the Ludwig Center for Cancer Genetics and Therapeutics at The John Hopkins Sidney Kimmel Comprehensive Cancer Center).« less

Decontamination of Anthrax spores in critical infrastructure and critical assets.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boucher, Raymond M.; Crown, Kevin K.; Tucker, Mark David

2010-05-01

Decontamination of anthrax spores in critical infrastructure (e.g., subway systems, major airports) and critical assets (e.g., the interior of aircraft) can be challenging because effective decontaminants can damage materials. Current decontamination methods require the use of highly toxic and/or highly corrosive chemical solutions because bacterial spores are very difficult to kill. Bacterial spores such as Bacillus anthracis, the infectious agent of anthrax, are one of the most resistant forms of life and are several orders of magnitude more difficult to kill than their associated vegetative cells. Remediation of facilities and other spaces (e.g., subways, airports, and the interior of aircraft)more » contaminated with anthrax spores currently requires highly toxic and corrosive chemicals such as chlorine dioxide gas, vapor- phase hydrogen peroxide, or high-strength bleach, typically requiring complex deployment methods. We have developed a non-toxic, non-corrosive decontamination method to kill highly resistant bacterial spores in critical infrastructure and critical assets. A chemical solution that triggers the germination process in bacterial spores and causes those spores to rapidly and completely change to much less-resistant vegetative cells that can be easily killed. Vegetative cells are then exposed to mild chemicals (e.g., low concentrations of hydrogen peroxide, quaternary ammonium compounds, alcohols, aldehydes, etc.) or natural elements (e.g., heat, humidity, ultraviolet light, etc.) for complete and rapid kill. Our process employs a novel germination solution consisting of low-cost, non-toxic and non-corrosive chemicals. We are testing both direct surface application and aerosol delivery of the solutions. A key Homeland Security need is to develop the capability to rapidly recover from an attack utilizing biological warfare agents. This project will provide the capability to rapidly and safely decontaminate critical facilities and assets to return them to normal operations as quickly as possible, sparing significant economic damage by re-opening critical facilities more rapidly and safely. Facilities and assets contaminated with Bacillus anthracis (i.e., anthrax) spores can be decontaminated with mild chemicals as compared to the harsh chemicals currently needed. Both the 'germination' solution and the 'kill' solution are constructed of 'off-the-shelf,' inexpensive chemicals. The method can be utilized by directly spraying the solutions onto exposed surfaces or by application of the solutions as aerosols (i.e., small droplets), which can also reach hidden surfaces.« less

Studies on mould growth and biomass production using waste banana peel.

PubMed

Essien, J P; Akpan, E J; Essien, E P

2005-09-01

Hyphomycetous (Aspergillus fumigatus) and Phycomycetous (Mucor hiemalis) moulds were cultivated in vitro at room temperature (28 + 20 degrees C) to examined their growth and biomass production on waste banana peel agar (BPA) and broth (BPB) using commercial malt extract agar (MEA) and broth (MEB) as control. The moulds grew comparatively well on banana peel substrates. No significant difference (p > 0.05) in radial growth rates was observed between moulds cultivated on PBA and MEA, although growth rates on MEA were slightly better. Slight variations in sizes of asexual spores and reproductive hyphae were also observed between moulds grown on MEA and BPA. Smaller conidia and sporangiospores, and shorter aerial hyphae (conidiophores and sporangiophores) were noticed in moulds grown on BPA than on MEA. The biomass weight of the test moulds obtained after one month of incubation with BPB were only about 1.8 mg and 1.4 mg less than values recorded for A. fumigatus and M. hiemalis respectively, grown on MEB. The impressive performance of the moulds on banana peel substrate may be attributed to the rich nutrient (particularly the crude protein 7.8% and crude fat 11.6% contents) composition of banana peels. The value of this agricultural waste can therefore be increased by its use not only in the manufacture of mycological medium but also in the production of valuable microfungal biomass which is rich in protein and fatty acids.

Complementary symbiont contributions to plant decomposition in a fungus-farming termite

PubMed Central

Hu, Haofu; Li, Cai; Chen, Zhensheng; Otani, Saria; Nygaard, Sanne; Nobre, Tania; Klaubauf, Sylvia; Schindler, Philipp M.; Hauser, Frank; Pan, Hailin; Yang, Zhikai; Sonnenberg, Anton S. M.; de Beer, Z. Wilhelm; Zhang, Yong; Wingfield, Michael J.; Grimmelikhuijzen, Cornelis J. P.; de Vries, Ronald P.; Korb, Judith; Aanen, Duur K.; Wang, Jun; Boomsma, Jacobus J.; Zhang, Guojie

2014-01-01

Termites normally rely on gut symbionts to decompose organic matter but the Macrotermitinae domesticated Termitomyces fungi to produce their own food. This transition was accompanied by a shift in the composition of the gut microbiota, but the complementary roles of these bacteria in the symbiosis have remained enigmatic. We obtained high-quality annotated draft genomes of the termite Macrotermes natalensis, its Termitomyces symbiont, and gut metagenomes from workers, soldiers, and a queen. We show that members from 111 of the 128 known glycoside hydrolase families are represented in the symbiosis, that Termitomyces has the genomic capacity to handle complex carbohydrates, and that worker gut microbes primarily contribute enzymes for final digestion of oligosaccharides. This apparent division of labor is consistent with the Macrotermes gut microbes being most important during the second passage of comb material through the termite gut, after a first gut passage where the crude plant substrate is inoculated with Termitomyces asexual spores so that initial fungal growth and polysaccharide decomposition can proceed with high efficiency. Complex conversion of biomass in termite mounds thus appears to be mainly accomplished by complementary cooperation between a domesticated fungal monoculture and a specialized bacterial community. In sharp contrast, the gut microbiota of the queen had highly reduced plant decomposition potential, suggesting that mature reproductives digest fungal material provided by workers rather than plant substrate. PMID:25246537

Relation of indoor and outdoor airborne fungal spore levels in the Kansas City metropolitan area.

PubMed

Jara, David; Portnoy, Jay; Dhar, Minati; Barnes, Charles

2017-03-01

Environmental control is an important component of asthma management for persons with asthma. A damp indoor environment and elevated airborne spore levels are factors in housing environmental control. We investigated if indoor airborne fungal spore levels correlated with outdoor ground-level airborne fungal spores or outdoor centrally collected spore levels as to types and abundance. Air collections were taken from home interiors, outdoor areas adjacent to the homes, and at a central location in the metropolitan area at the approximate same time. All air collections were examined and enumerated microscopically, and airborne spore estimates per cubic meter of air were reported for total fungal spores and for 11 identifiable spore groups. The 244 homes in the study were typical of the North American Midwest. The overall mean total spore counts in spores per cubic meter of air was indoors (4076 spores/m3), outdoors at ground level (8899 spores/m3), and outdoor metropolitan area (8342 spores/m3). All of the major indoor taxa were strongly correlated with the mean total spores present in the home. Total outdoor ground spore levels were highly correlated with levels of major outdoor taxa, such as ascospores and Cladosporium. Correlations of indoor spore levels with outdoor spore levels are strong for most major outdoor taxa. Indoor Aspergillus-Penicillium and Chaetomium are significantly correlated between indoor and local ground-level outdoor air. Although conditions may exist where indoor or outdoor spore levels were not well aligned, in most circumstances, the outdoor airborne spore community was reflected in the indoor airborne spore community.

On the neutralization of bacterial spores in post-detonation flows

NASA Astrophysics Data System (ADS)

Gottiparthi, K. C.; Schulz, J. C.; Menon, S.

2014-09-01

In multiple operational scenarios, explosive charges are used to neutralize confined or unconfined stores of bacterial spores. The spore destruction is achieved by post-detonation combustion and mixing of hot detonation product gases with the ambient flow and spore clouds. In this work, blast wave interaction with bacterial spore clouds and the effect of post-detonation combustion on spore neutralization are investigated using numerical simulations. Spherical explosive charges (radius, = 5.9 cm) comprising of nitromethane are modeled in the vicinity of a spore cloud, and the spore kill in the post-detonation flow is quantified. The effect of the mass of the spores and the initial distance, , of the spore cloud from the explosive charge on the percentage of spores neutralized is investigated. When the spores are initially placed within a distance of 3.0, within 0.1 ms after detonation of the charge, all the spores are neutralized by the blast wave and the hot detonation product gases. In contrast, almost all the spores survived the explosion when is greater than 8.0. The percentage of intact spores varied from 0 to 100 for 3.0 8.0 with spore neutralization dependent on time spent by the spores in the post-detonation mixing/combustion zone.

Distinction of broken cellular wall Ganoderma lucidum spores and G. lucidum spores using FTIR microspectroscopy

NASA Astrophysics Data System (ADS)

Chen, Xianliang; Liu, Xingcun; Sheng, Daping; Huang, Dake; Li, Weizu; Wang, Xin

2012-11-01

In this paper, FTIR microspectroscopy was used to identify broken cellular wall Ganoderma lucidum spores and G. lucidum spores. For IR spectra, broken cellular wall G. lucidum spores and G. lucidum spores were mainly different in the regions of 3000-2800, 1660-1600, 1400-1200 and 1100-1000 cm-1. For curve fitting, the results showed the differences in the protein secondary structures and the polysaccharide structures/content between broken cellular wall G. lucidum spores and G. lucidum spores. Moreover, the value of A1078/A1741 might be a potentially useful factor to distinguish broken cellular wall G. lucidum spores from G. lucidum spores. Additionally, FTIR microspectroscopy could identify broken cellular wall G. lucidum spores and G. lucidum spores accurately when it was combined with hierarchical cluster analysis. The result suggests FTIR microspectroscopy is very simple and efficient for distinction of broken cellular wall G. lucidum spores and G. lucidum spores. The result also indicates FTIR microspectroscopy may be useful for TCM identification.

PATHWAYS OF GLUCOSE CATABOLISM IN BACILLUS CEREUS1

PubMed Central

Goldman, Manuel; Blumenthal, Harold J.

1964-01-01

Goldman, Manuel (The University of Michigan, Ann Arbor), and Harold J. Blumenthal. Pathways of glucose catabolism in Bacillus cereus. J. Bacteriol. 87:377–386. 1964.—Estimates by a radiorespirometric method of the pathways of glucose catabolism of resting-cell suspensions of Bacillus cereus strain terminalis indicate that the Embden-Meyerhof pathway predominates at every stage of development, including the sporogenic and germinative phases. At the filamentous, granular, forespore, and transitional stages, 98% of the glucose was catabolized by the Embden-Meyerhof pathway, and the remainder by the hexose monophosphate oxidative pathway. Estimates of the pathways in resting spore-suspensions arrested at defined stages of development indicate that 20% of the glucose was catabolized through the hexose monophosphate pathway in germinated spores, and 10% in the swollen and elongated stages of postgermination. In cells which had completed the first cell division, the figure fell to about 2%, a level similar to that found for vegetative cells at later stages of development. The key Embden-Meyerhof enzymes, hexokinase, phosphohexoisomerase, phosphofructokinase, and aldolase, as well as several other enzymes, were present at all stages of germination and postgerminative development, supporting the radioisotopic data obtained with whole cells. As indicated by the release of C14O2 from glucose-6-C14, terminal respiration of resting-cell suspensions operates maximally in vegetative cells at the granular, fore-spore, and transitional stages. There was marked inhibition of terminal respiration during the development of spores into vegetative cells. Only slight activity occurred in the earliest vegetative stages, and maximal operation developed after about ten cell divisions. Fumarase was absent in spores until sometime late in the elongation stage. At this point, a weak but definite activity appeared which increased during later stages of development so that, by the end of about the sixth cell division, fumarase had a specific activity about 80 times that observed at elongation. PMID:14151060

Inhibiting Inosine Hydrolase and Alanine Racemase to Enhance the Germination of Bacillus anthracis Sterne Spores: Potential Spore Decontamination Strategies

DTIC Science & Technology

2015-06-19

animal waste an~ decompositiOn DISTRIBUTION STATEMENT A: Approved for public release; distribution is unlimited. UNCLASSIFIED PR-15-306 Anthrax...influx of water. Ungerminated spore Germination Germinated spore Spore hydratation ~ Non-refractile spore Refractile spore • Fluorescence

Purification and partial characterization of PfHRP-II protein of Plasmodium falciparum.

PubMed

Ghimire, Prakash; Samantaray, J C; Mirdha, B R; Patra, A K; Panda, A K

2003-12-01

The human malarial parasite Plasmodium falciparum secretes various intra-and extra-cellular proteins during its asexual life cycle in human RBC. Histidine rich protein-II (HRP-II) is one of the most prominent proteins, found to be secreted by P. falciparum throughout the asexual cycle with the peak during mature schizont stage of the parasite development in human IRBC. The high histidine content (35% of the total amino acids in protein) of this protein suggested the potential to bind divalent metal ions. We have demonstrated by metal chelate chromatography, an extraordinary capacity of HRP-II to bind nickel ions (Ni++) and employed this characteristic to purify the extra-cellular HRP-II protein secreted by P. falciparum from culture supernatant. The identity of the purified protein was verified by the relative molecular weight on SDS-PAGE, by reacting with polyclonal antibodies directed against it using Western blot technique.

Chemical signatures and new drug targets for gametocytocidal drug development

NASA Astrophysics Data System (ADS)

Sun, Wei; Tanaka, Takeshi Q.; Magle, Crystal T.; Huang, Wenwei; Southall, Noel; Huang, Ruili; Dehdashti, Seameen J.; McKew, John C.; Williamson, Kim C.; Zheng, Wei

2014-01-01

Control of parasite transmission is critical for the eradication of malaria. However, most antimalarial drugs are not active against P. falciparum gametocytes, responsible for the spread of malaria. Consequently, patients can remain infectious for weeks after the clearance of asexual parasites and clinical symptoms. Here we report the identification of 27 potent gametocytocidal compounds (IC50 < 1 μM) from screening 5,215 known drugs and compounds. All these compounds were active against three strains of gametocytes with different drug sensitivities and geographical origins, 3D7, HB3 and Dd2. Cheminformatic analysis revealed chemical signatures for P. falciparum sexual and asexual stages indicative of druggability and suggesting potential targets. Torin 2, a top lead compound (IC50 = 8 nM against gametocytes in vitro), completely blocked oocyst formation in a mouse model of transmission. These results provide critical new leads and potential targets to expand the repertoire of malaria transmission-blocking reagents.

The powdery mildews: a review of the world's most familiar (yet poorly known) plant pathogens.

PubMed

Glawe, Dean A

2008-01-01

The past decade has seen fundamental changes in our understanding of powdery mildews (Erysiphales). Research on molecular phylogeny demonstrated that Erysiphales are Leotiomycetes (inoperculate discomycetes) rather than Pyrenomycetes or Plectomycetes. Life cycles are surprisingly variable, including both sexual and asexual states, or only sexual states, or only asexual states. At least one species produces dematiaceous conidia. Analyses of rDNA sequences indicate that major lineages are more closely correlated with anamorphic features such as conidial ontogeny and morphology than with teleomorph features. Development of molecular clock models is enabling researchers to reconstruct patterns of coevolution and host-jumping, as well as ancient migration patterns. Geographic distributions of some species appear to be increasing rapidly but little is known about species diversity in many large areas, including North America. Powdery mildews may already be responding to climate change, suggesting they may be useful models for studying effects of climate change on plant diseases.

Effects of meteorological conditions on spore plumes

NASA Astrophysics Data System (ADS)

Burch, M.; Levetin, E.

2002-05-01

Fungal spores are an ever-present component of the atmosphere, and have long been known to trigger asthma and hay fever symptoms in sensitive individuals. The atmosphere around Tulsa has been monitored for airborne spores and pollen with Burkard spore traps at several sampling stations. This study involved the examination of the hourly spore concentrations on days that had average daily concentrations near 50,000 spores/m3 or greater. Hourly concentrations of Cladosporium, Alternaria, Epicoccum, Curvularia, Pithomyces, Drechslera, smut spores, ascospores, basidiospores, other, and total spores were determined on 4 days at three sites and then correlated with hourly meteorological data including temperature, rainfall, wind speed, dew point, air pressure, and wind direction. On each of these days there was a spore plume, a phenomenon in which spore concentrations increased dramatically over a very short period of time. Spore plumes generally occurred near midday, and concentrations were seen to increase from lows around 20,000 total spores/m3 to highs over 170,000 total spores/m3 in 2 h. Multiple regression analysis of the data indicated that increases in temperature, dew point, and air pressure correlated with the increase in spore concentrations, but no single weather variable predicted the appearance of a spore plume. The proper combination of changes in these meteorological parameters that result in a spore plume may be due to the changing weather conditions associated with thunderstorms, as on 3 of the 4 days when spore plumes occurred there were thunderstorms later that evening. The occurrence of spore plumes may have clinical significance, because other studies have shown that sensitization to certain spore types can occur during exposure to high spore concentrations.

Spore collection and elimination apparatus and method

DOEpatents

Czajkowski, Carl [South Jamesport, NY; Warren, Barbara Panessa [Port Jefferson, NY

2007-04-03

The present invention is for a spore collection apparatus and its method of use. The portable spore collection apparatus includes a suction source, a nebulizer, an ionization chamber and a filter canister. The suction source collects the spores from a surface. The spores are activated by heating whereby spore dormancy is broken. Moisture is then applied to the spores to begin germination. The spores are then exposed to alpha particles causing extinction.

An insight into spore dispersal of Ganoderma boninense on oil palm.

PubMed

Sanderson, F R

2005-01-01

The disease of oil palm caused by Ganoderma boninense, although universally referred to as Ganoderma basal stem rot, occurs in three very distinct phases, with basal stem rot only part of the disease cycle. G. boninense also causes a seedling disease and an upper stem rot. An understanding of spore dispersal provides an insight into where spores of G. boninense have a role in the infection process. This role will be discussed in relation to each of these three infection phases. This understanding is a critical component of developing a successful disease control strategy.

A rapid colorimetric assay for mold spore germination using XTT tetrazolium salt

Treesearch

Carol A. Clausen; Vina W. Yang

2011-01-01

Current laboratory test methods to measure efficacy of new mold inhibitors are time consuming, some require specialized test equipment and ratings are subjective. Rapid, simple quantitative assays to measure the efficacy of mold inhibitors are needed. A quantitative, colorimetric microassay was developed using XTT tetrazolium salt to metabolically assess mold spore...

Distinction of broken cellular wall Ganoderma lucidum spores and G. lucidum spores using FTIR microspectroscopy.

PubMed

Chen, Xianliang; Liu, Xingcun; Sheng, Daping; Huang, Dake; Li, Weizu; Wang, Xin

2012-11-01

In this paper, FTIR microspectroscopy was used to identify broken cellular wall Ganoderma lucidum spores and G. lucidum spores. For IR spectra, broken cellular wall G. lucidum spores and G. lucidum spores were mainly different in the regions of 3000-2800, 1660-1600, 1400-1200 and 1100-1000 cm(-1). For curve fitting, the results showed the differences in the protein secondary structures and the polysaccharide structures/content between broken cellular wall G. lucidum spores and G. lucidum spores. Moreover, the value of A1078/A1741 might be a potentially useful factor to distinguish broken cellular wall G. lucidum spores from G. lucidum spores. Additionally, FTIR microspectroscopy could identify broken cellular wall G. lucidum spores and G. lucidum spores accurately when it was combined with hierarchical cluster analysis. The result suggests FTIR microspectroscopy is very simple and efficient for distinction of broken cellular wall G. lucidum spores and G. lucidum spores. The result also indicates FTIR microspectroscopy may be useful for TCM identification. Copyright © 2012 Elsevier B.V. All rights reserved.

Engineering Rugged Field Assays to Detect Hazardous Chemicals Using Spore-Based Bacterial Biosensors.

PubMed

Wynn, Daniel; Deo, Sapna; Daunert, Sylvia

2017-01-01

Bacterial whole cell-based biosensors have been genetically engineered to achieve selective and reliable detection of a wide range of hazardous chemicals. Although whole-cell biosensors demonstrate many advantages for field-based detection of target analytes, there are still some challenges that need to be addressed. Most notably, their often modest shelf life and need for special handling and storage make them challenging to use in situations where access to reagents, instrumentation, and expertise are limited. These problems can be circumvented by developing biosensors in Bacillus spores, which can be engineered to address all of these concerns. In its sporulated state, a whole cell-based biosensor has a remarkably long life span and is exceptionally resistant to environmental insult. When these spores are germinated for use in analytical techniques, they show no loss in performance, even after long periods of storage under harsh conditions. In this chapter, we will discuss the development and use of whole cell-based sensors, their adaptation to spore-based biosensors, their current applications, and future directions in the field. © 2017 Elsevier Inc. All rights reserved.

Monitoring Rates and Heterogeneity of High-Pressure Germination of Bacillus Spores by Phase-Contrast Microscopy of Individual Spores

DTIC Science & Technology

2014-01-01

wild-type spores but ~15-fold higher deltaTrelease values; v ) germination kinetics of wild-type spores given a ? 30 sec 140 MPa HP pulse followed by...15-fold longer than those for wild-type spores, but the two types of spores exhibited similar average Tlag values; and ( v ) the germination of wild-type...committed spores, as it does for nutrient-committed spores (14)? ( v ) Can these HP-com- mitted spores be isolated under conditions that do not allow

Area and Carbon Content of Sphagnum Since Last Glacial Maximum

DOE Data Explorer

Gajewski, K. [University of Ottawa, Ottawa, Ontario (Canada); Viau, A. [University of Ottawa, Ottawa, Ontario (Canada); Sawada, M. [University of Ottawa, Ottawa, Ontario (Canada); Atkinson, D. [University of Ottawa, Ottawa, Ontario (Canada); Wilson, S. [University of Ottawa, Ottawa, Ontario (Canada)

2002-01-01

The distribution and abundance of Sphagnum spores in North America and Eurasia are mapped for the past 21ka, as described in Gajewski et al. (2002). In summary, spore data were taken from existing pollen data bases, as were radiocarbon chronologies. The abundance of Sphagnum spores was mapped at 2000-year intervals beginning 21000 years BP (before present). The present-day distribution of abundant Sphagnum spores corresponds closely to areas with peatland development, with maximum Sphagnum abundance between 630 and 1300 mm annual precipitation and between -2° and 60°C mean annual air temperature. Carbon content of peatlands was generated from estimated peatland area, calculated values of peat thickness, and specified values of bulk density (112 × 10³ g m^-3) and fraction of carbon (51.7%).

Immunoproteomics Approach for Screening of Vaccine Candidates against Intestinal Botulism.

PubMed

Sharma, Arti; Rani, Sarita; Alam, Syed Imteyaz; Ponmariappan, Sarkaraisamy

2017-01-01

Intestinal botulism is an infectious form of botulism in which disease results from ingesting spores, which is followed by spore germination and intraluminal production of botulinum neurotoxins over an extended period. Botulinum neurotoxin is produced by endospore forming bacteria called C. botulinum. Immunoproteomic study was used to screen the cross reactive immunogenic proteins of Clostridium botulinum type B using C. botulinum type B live spore antiserum. The whole cell proteins were separated by two dimensional gel electrophoresis and transferred to polyvinylidene difluoride membranes. Further, the Western blotting was performed with mouse pups immune serum against C. botulinum type B live spores. Eight predominant cross immunoreactive proteins were identified by mass spectrometry. These immunogenic proteins might be used to develop novel subunit vaccine candidates against the intestinal botulism. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Germination and Outgrowth of Single Spores of Saccharomyces cerevisiae Viewed by Scanning Electron and Phase-Contrast Microscopy

PubMed Central

Rousseau, Paul; Halvorson, Harlyn O.; Bulla, Lee A.; Julian, Grant St.

1972-01-01

Single spores of Saccharomyces cerevisiae were examined during germination and outgrowth by scanning electron and phase-contrast microscopy. Also determined were changes in cell weight and light absorbance, trehalose utilization, and synthesis of protein and KOH-soluble carbohydrates. These studies reveal that development of the vegetative cell from a spore follows a definite sequence of events involving dramatic physical and chemical modifications. These changes are: initial rapid loss in cellular absorbance followed later by an abrupt gain in absorbance; reduction in cell weight and a subsequent progressive increase; modification of the spore surface with concomitant diminution in refractility; elongation of the cell and augmentation of surface irregularities; rapid decline in trehalose content of the cell accompanied by extensive formation of KOH-soluble carbohydrates; and bud formation. Images PMID:4551750

Detection of Bacillus anthracis spores by super-paramagnetic lateral-flow immunoassays based on "Road Closure".

PubMed

Wang, Dian-Bing; Tian, Bo; Zhang, Zhi-Ping; Wang, Xu-Ying; Fleming, Joy; Bi, Li-Jun; Yang, Rui-Fu; Zhang, Xian-En

2015-05-15

Detection of Bacillus anthracis in the field, whether as a natural infection or as a biothreat remains challenging. Here we have developed a new lateral-flow immunochromatographic assay (LFIA) for B. anthracis spore detection based on the fact that conjugates of B. anthracis spores and super-paramagnetic particles labeled with antibodies will block the pores of chromatographic strips and form retention lines on the strips, instead of the conventionally reported test lines and control lines in classic LFIA. As a result, this new LFIA can simultaneously realize optical, magnetic and naked-eye detection by analyzing signals from the retention lines. As few as 500-700 pure B. anthracis spores can be recognized with CV values less than 8.31% within 5 min of chromatography and a total time of 20 min. For powdery sample tests, this LFIA can endure interference from 25% (w/v) milk, 10% (w/v) baking soda and 10% (w/v) starch without any sample pre-treatment, and has a corresponding detection limit of 6×10(4) spores/g milk powder, 2×10(5) spores/g starch and 5×10(5) spores/g baking soda. Compared with existing methods, this new approach is very competitive in terms of sensitivity, specificity, cost and ease of operation. This proof-of-concept study can also be extended for detection of many other large-sized analytes. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Evaluation of Stachybotrys chartarum in the house of an infant with pulmonary hemorrhage: quantitative assessment before, during, and after remediation.

PubMed

Vesper, S; Dearborn, D G; Yike, I; Allan, T; Sobolewski, J; Hinkley, S F; Jarvis, B B; Haugland, R A

2000-03-01

Stachybotrys chartarum is an indoor mold that has been associated with pulmonary hemorrhage cases in the Cleveland, Ohio, area. This study applied two new quantitative measurements to air samples from a home in which an infant developed PH. Quantitative polymerase chain reaction and a protein synthesis inhibition assay were used to determine the level of S. chartarum spores and their toxicity in air samples taken before, during, and after a remediation program was implemented to remove the fungus. Initial spore concentrations were between 0.1 and 9.3 spores/m3 of air, and the toxicity of air particulates was correspondingly low. However, the dust in the house contained between 0.4 and 2.1 x 10(3) spores/mg (as determined by hemocytometer counts). The remediation program removed all contaminated wallboard, paneling, and carpeting in the water-damaged areas of the home. In addition, a sodium hypochlorite solution was used to spray all surfaces during remediation. Although spore counts and toxicity were high during remediation, air samples taken postremediation showed no detectable levels of S. chartarum or related toxicity. Nine isolates of S. chartarum obtained from the home were analyzed for spore toxicity, hemolytic activity, and random amplified polymorphic DNA banding patterns. None of the isolates produced highly toxic spores (>90 microg T2 toxin equivalents per gram wet weight spores) after growth for 10 and 30 days on wet wallboard, but three isolates were hemolytic consistently. DNA banding patterns suggested that at least one of these isolates was related to isolates from homes of infants with previously investigated cases.

The inhibitory effects of essential oil constituents against germination, outgrowth and vegetative growth of spores of Clostridium perfringens type A in laboratory medium and chicken meat.

PubMed

Alanazi, Saud; Alnoman, Maryam; Banawas, Saeed; Saito, Ryoichi; Sarker, Mahfuzur R

2018-08-01

C. perfringens type A is the causative agent of C. perfringens type A food poisoning (FP) and non-food-borne (NFB) human gastrointestinal diseases. Due to its ability to form highly heat-resistant spores, it is of great interest to develop strategies alternative to thermal processing to inactivate C. perfrinegens. Thus, in this study we evaluated the inhibitory effects of essential oil constituents (EOCs) (cinnamaldehyde, eugenol, allyl isothiocyanate (AITC), and carvacrol) against germination, outgrowth and vegetative growth of spores of C. perfringens FP and NFB disease isolates in laboratory medium and chicken meat. The cinnamaldehyde, eugenol and carvacrol, but not AITC, all at 0.05-0.1%, inhibited the germination of spores of all tested C. perfringens isolates in Tripticase-glucose-yeast extract (TGY) medium. Furthermore, all tested EOCs at 0.05-0.1% arrested the outgrowth and vegetative growth of C. perfringens spores in TGY, with AITC and carvacrol being the most effective. However, among four tested EOCs, only AITC (at 0.5%-2.0%) was able to inhibit the growth of C. perfringens spores in chicken meat and no such inhibitory effect was observed even with a 10-fold higher concentration (5%) of carvacrol. In conclusion, our current work identified AITC as an effective EOC to control spores and vegetative cells of C. perfringens isolates in laboratory medium and chicken meat. Further studies on evaluating the effectiveness of different combination of EOCs against C. perfringens spore growth in different meat products should establish an effective use of EOCs to control the risk of C. perfringens-mediated illnesses. Copyright © 2018 Elsevier Ltd. All rights reserved.

Computational Fluid Dynamics Modeling of Bacillus anthracis ...

EPA Pesticide Factsheets

Journal Article Three-dimensional computational fluid dynamics and Lagrangian particle deposition models were developed to compare the deposition of aerosolized Bacillus anthracis spores in the respiratory airways of a human with that of the rabbit, a species commonly used in the study of anthrax disease. The respiratory airway geometries for each species were derived from computed tomography (CT) or µCT images. Both models encompassed airways that extended from the external nose to the lung with a total of 272 outlets in the human model and 2878 outlets in the rabbit model. All simulations of spore deposition were conducted under transient, inhalation-exhalation breathing conditions using average species-specific minute volumes. Four different exposure scenarios were modeled in the rabbit based upon experimental inhalation studies. For comparison, human simulations were conducted at the highest exposure concentration used during the rabbit experimental exposures. Results demonstrated that regional spore deposition patterns were sensitive to airway geometry and ventilation profiles. Despite the complex airway geometries in the rabbit nose, higher spore deposition efficiency was predicted in the upper conducting airways of the human at the same air concentration of anthrax spores. This greater deposition of spores in the upper airways in the human resulted in lower penetration and deposition in the tracheobronchial airways and the deep lung than that predict

Arbuscular mycorrhizal fungi spore propagation using single spore as starter inoculum and a plant host.

PubMed

Selvakumar, G; Shagol, C C; Kang, Y; Chung, B N; Han, S G; Sa, T M

2018-06-01

The propagation of pure cultures of arbuscular mycorrhizal fungal (AMF) is an essential requirement for their large-scale agricultural application and commercialization as biofertilizers. The present study aimed to propagate AMF using the single-spore inoculation technique and compare their propagation ability with the known reference spores. Arbuscular mycorrhizal fungal spores were collected from salt-affected Saemangeum reclaimed soil in South Korea. The technique involved inoculation of sorghum-sudangrass (Sorghum bicolor L.) seedlings with single, healthy spores on filter paper followed by the transfer of successfully colonized seedlings to 1-kg capacity pots containing sterilized soil. After the first plant cycle, the contents were transferred to 2·5-kg capacity pots containing sterilized soil. Among the 150 inoculated seedlings, only 27 seedlings were colonized by AMF spores. After 240 days, among the 27 seedlings, five inoculants resulted in the production of over 500 spores. The 18S rDNA sequencing of spores revealed that the spores produced through single-spore inoculation method belonged to Gigaspora margarita, Claroideoglomus lamellosum and Funneliformis mosseae. Furthermore, indigenous spore F. mosseae M-1 reported a higher spore count than the reference spores. The AMF spores produced using the single-spore inoculation technique may serve as potential bio-inoculants with an advantage of being more readily adopted by farmers due to the lack of requirement of a skilled technique in spore propagation. The results of the current study describe the feasible and cost-effective method to mass produce AMF spores for large-scale application. The AMF spores obtained from this method can effectively colonize plant roots and may be easily introduced to the new environment. © 2018 The Society for Applied Microbiology.

Bestatin Inhibits Cell Growth, Cell Division, and Spore Cell Differentiation in Dictyostelium discoideum

PubMed Central

Poloz, Yekaterina; Catalano, Andrew

2012-01-01

Bestatin methyl ester (BME) is an inhibitor of Zn2+-binding aminopeptidases that inhibits cell proliferation and induces apoptosis in normal and cancer cells. We have used Dictyostelium as a model organism to study the effects of BME. Only two Zn2+-binding aminopeptidases have been identified in Dictyostelium to date, puromycin-sensitive aminopeptidase A and B (PsaA and PsaB). PSA from other organisms is known to regulate cell division and differentiation. Here we show that PsaA is differentially expressed throughout growth and development of Dictyostelium, and its expression is regulated by developmental morphogens. We present evidence that BME specifically interacts with PsaA and inhibits its aminopeptidase activity. Treatment of cells with BME inhibited the rate of cell growth and the frequency of cell division in growing cells and inhibited spore cell differentiation during late development. Overexpression of PsaA-GFP (where GFP is green fluorescent protein) also inhibited spore cell differentiation but did not affect growth. Using chimeras, we have identified that nuclear versus cytoplasmic localization of PsaA affects the choice between stalk or spore cell differentiation pathway. Cells that overexpressed PsaA-GFP (primarily nuclear) differentiated into stalk cells, while cells that overexpressed PsaAΔNLS2-GFP (cytoplasmic) differentiated into spores. In conclusion, we have identified that BME inhibits cell growth, division, and differentiation in Dictyostelium likely through inhibition of PsaA. PMID:22345351

Evolutionary dynamics of phenotype-structured populations: from individual-level mechanisms to population-level consequences

NASA Astrophysics Data System (ADS)

Chisholm, Rebecca H.; Lorenzi, Tommaso; Desvillettes, Laurent; Hughes, Barry D.

2016-08-01

Epigenetic mechanisms are increasingly recognised as integral to the adaptation of species that face environmental changes. In particular, empirical work has provided important insights into the contribution of epigenetic mechanisms to the persistence of clonal species, from which a number of verbal explanations have emerged that are suited to logical testing by proof-of-concept mathematical models. Here, we present a stochastic agent-based model and a related deterministic integrodifferential equation model for the evolution of a phenotype-structured population composed of asexually-reproducing and competing organisms which are exposed to novel environmental conditions. This setting has relevance to the study of biological systems where colonising asexual populations must survive and rapidly adapt to hostile environments, like pathogenesis, invasion and tumour metastasis. We explore how evolution might proceed when epigenetic variation in gene expression can change the reproductive capacity of individuals within the population in the new environment. Simulations and analyses of our models clarify the conditions under which certain evolutionary paths are possible and illustrate that while epigenetic mechanisms may facilitate adaptation in asexual species faced with environmental change, they can also lead to a type of "epigenetic load" and contribute to extinction. Moreover, our results offer a formal basis for the claim that constant environments favour individuals with low rates of stochastic phenotypic variation. Finally, our model provides a "proof of concept" of the verbal hypothesis that phenotypic stability is a key driver in rescuing the adaptive potential of an asexual lineage and supports the notion that intense selection pressure can, to an extent, offset the deleterious effects of high phenotypic instability and biased epimutations, and steer an asexual population back from the brink of an evolutionary dead end.

Methods for Integrated Air Sampling and DNA Analysis for Detection of Airborne Fungal Spores

PubMed Central

Williams, Roger H.; Ward, Elaine; McCartney, H. Alastair

2001-01-01

Integrated air sampling and PCR-based methods for detecting airborne fungal spores, using Penicillium roqueforti as a model fungus, are described. P. roqueforti spores were collected directly into Eppendorf tubes using a miniature cyclone-type air sampler. They were then suspended in 0.1% Nonidet P-40, and counted using microscopy. Serial dilutions of the spores were made. Three methods were used to produce DNA for PCR tests: adding untreated spores to PCRs, disrupting spores (fracturing of spore walls to release the contents) using Ballotini beads, and disrupting spores followed by DNA purification. Three P. roqueforti-specific assays were tested: single-step PCR, nested PCR, and PCR followed by Southern blotting and probing. Disrupting the spores was found to be essential for achieving maximum sensitivity of the assay. Adding untreated spores to the PCR did allow the detection of P. roqueforti, but this was never achieved when fewer than 1,000 spores were added to the PCR. By disrupting the spores, with or without subsequent DNA purification, it was possible to detect DNA from a single spore. When known quantities of P. roqueforti spores were added to air samples consisting of high concentrations of unidentified fungal spores, pollen, and dust, detection sensitivity was reduced. P. roqueforti DNA could not be detected using untreated or disrupted spore suspensions added to the PCRs. However, using purified DNA, it was possible to detect 10 P. roqueforti spores in a background of 4,500 other spores. For all DNA extraction methods, nested PCR was more sensitive than single-step PCR or PCR followed by Southern blotting. PMID:11375150

Formation and characterization of non-growth states in Clostridium thermocellum: spores and L-forms

PubMed Central

2012-01-01

Background Clostridium thermocellum is an anaerobic thermophilic bacterium that exhibits high levels of cellulose solublization and produces ethanol as an end product of its metabolism. Using cellulosic biomass as a feedstock for fuel production is an attractive prospect, however, growth arrest can negatively impact ethanol production by fermentative microorganisms such as C. thermocellum. Understanding conditions that lead to non-growth states in C. thermocellum can positively influence process design and culturing conditions in order to optimize ethanol production in an industrial setting. Results We report here that Clostridium thermocellum ATCC 27405 enters non-growth states in response to specific growth conditions. Non-growth states include the formation of spores and a L-form-like state in which the cells cease to grow or produce the normal end products of metabolism. Unlike other sporulating organisms, we did not observe sporulation of C. thermocellum in low carbon or nitrogen environments. However, sporulation did occur in response to transfers between soluble and insoluble substrates, resulting in approximately 7% mature spores. Exposure to oxygen caused a similar sporulation response. Starvation conditions during continuous culture did not result in spore formation, but caused the majority of cells to transition to a L-form state. Both spores and L-forms were determined to be viable. Spores exhibited enhanced survival in response to high temperature and prolonged storage compared to L-forms and vegetative cells. However, L-forms exhibited faster recovery compared to both spores and stationary phase cells when cultured in rich media. Conclusions Both spores and L-forms cease to produce ethanol, but provide other advantages for C. thermocellum including enhanced survival for spores and faster recovery for L-forms. Understanding the conditions that give rise to these two different non-growth states, and the implications that each has for enabling or enhancing C. thermocellum survival may promote the efficient cultivation of this organism and aid in its development as an industrial microorganism. PMID:22897981

Architecture and assembly of the Bacillus subtilis spore coat.

PubMed

Plomp, Marco; Carroll, Alicia Monroe; Setlow, Peter; Malkin, Alexander J

2014-01-01

Bacillus spores are encased in a multilayer, proteinaceous self-assembled coat structure that assists in protecting the bacterial genome from stresses and consists of at least 70 proteins. The elucidation of Bacillus spore coat assembly, architecture, and function is critical to determining mechanisms of spore pathogenesis, environmental resistance, immune response, and physicochemical properties. Recently, genetic, biochemical and microscopy methods have provided new insight into spore coat architecture, assembly, structure and function. However, detailed spore coat architecture and assembly, comprehensive understanding of the proteomic composition of coat layers, and specific roles of coat proteins in coat assembly and their precise localization within the coat remain in question. In this study, atomic force microscopy was used to probe the coat structure of Bacillus subtilis wild type and cotA, cotB, safA, cotH, cotO, cotE, gerE, and cotE gerE spores. This approach provided high-resolution visualization of the various spore coat structures, new insight into the function of specific coat proteins, and enabled the development of a detailed model of spore coat architecture. This model is consistent with a recently reported four-layer coat assembly and further adds several coat layers not reported previously. The coat is organized starting from the outside into an outermost amorphous (crust) layer, a rodlet layer, a honeycomb layer, a fibrous layer, a layer of "nanodot" particles, a multilayer assembly, and finally the undercoat/basement layer. We propose that the assembly of the previously unreported fibrous layer, which we link to the darkly stained outer coat seen by electron microscopy, and the nanodot layer are cotH- and cotE- dependent and cotE-specific respectively. We further propose that the inner coat multilayer structure is crystalline with its apparent two-dimensional (2D) nuclei being the first example of a non-mineral 2D nucleation crystallization pattern in a biological organism.

Architecture and Assembly of the Bacillus subtilis Spore Coat

PubMed Central

Plomp, Marco; Carroll, Alicia Monroe; Setlow, Peter; Malkin, Alexander J.

2014-01-01

Bacillus spores are encased in a multilayer, proteinaceous self-assembled coat structure that assists in protecting the bacterial genome from stresses and consists of at least 70 proteins. The elucidation of Bacillus spore coat assembly, architecture, and function is critical to determining mechanisms of spore pathogenesis, environmental resistance, immune response, and physicochemical properties. Recently, genetic, biochemical and microscopy methods have provided new insight into spore coat architecture, assembly, structure and function. However, detailed spore coat architecture and assembly, comprehensive understanding of the proteomic composition of coat layers, and specific roles of coat proteins in coat assembly and their precise localization within the coat remain in question. In this study, atomic force microscopy was used to probe the coat structure of Bacillus subtilis wild type and cotA, cotB, safA, cotH, cotO, cotE, gerE, and cotE gerE spores. This approach provided high-resolution visualization of the various spore coat structures, new insight into the function of specific coat proteins, and enabled the development of a detailed model of spore coat architecture. This model is consistent with a recently reported four-layer coat assembly and further adds several coat layers not reported previously. The coat is organized starting from the outside into an outermost amorphous (crust) layer, a rodlet layer, a honeycomb layer, a fibrous layer, a layer of “nanodot” particles, a multilayer assembly, and finally the undercoat/basement layer. We propose that the assembly of the previously unreported fibrous layer, which we link to the darkly stained outer coat seen by electron microscopy, and the nanodot layer are cotH- and cotE- dependent and cotE-specific respectively. We further propose that the inner coat multilayer structure is crystalline with its apparent two-dimensional (2D) nuclei being the first example of a non-mineral 2D nucleation crystallization pattern in a biological organism. PMID:25259857

Production and processing of Metarhizium anisopliae var. acridum submerged conidia for locust and grasshopper control.

PubMed

Kassa, Adane; Stephan, Dietrich; Vidal, Stefan; Zimmermann, Gisbert

2004-01-01

Currently, mycopesticide development for locust and grasshopper control depends on aerial conidia or submerged spores of entomopathogenic fungi. In our study, the production of submerged conidia of Metarhizium anisopliae var. acridum (IMI 330189) was investigated in a liquid medium containing 3% biomalt and 1% yeast extract (BH-medium). The effects of freeze and spray drying techniques on the quality of submerged conidia were determined. The influence of different additives on the viability of fresh submerged conidia and their suitability for oil flowable concentrate formulation development was assessed. In a BH medium maintained at 180 rev min(-1), at 30 degrees C for 72 h, IMI 330189 produced a green pigmented biomass of submerged conidia whereas in Adámek medium it produced a yellowish biomass of submerged spores. The spore concentration was high in both media; however, the size of the spores produced in the BH medium was significantly lower than those produced in Adámek medium (P < 0.001). Submerged conidia can be effectively dried using either freeze or spray drying techniques. The viability and speed of germination were significantly affected by the drying and pulverizing process (P < 0.001). The initial viability was significantly higher for spray-dried submerged conidia than for freeze-dried spores. Pulverizing of freeze-dried submerged conidia reduced the speed of germination and the viability by 63-95%. Dried submerged conidia can be stored over 45 wk at low temperatures (< 10 degrees) without suffering a significant loss in viability. Furthermore, we have identified carriers that are suitable for oil flowable concentrate formulation development.

PsB multiprotein complex of Dictyostelium discoideum. Demonstration of cellulose binding activity and order of protein subunit assembly.

PubMed

McGuire, V; Alexander, S

1996-06-14

The differentiated spores of Dictyostelium are surrounded by an extracellular matrix, the spore coat, which protects them from environmental factors allowing them to remain viable for extended periods of time. This presumably is a major evolutionary advantage. This unique extracellular matrix is composed of cellulose and glycoproteins. Previous work has shown that some of these spore coat glycoproteins exist as a preassembled multiprotein complex (the PsB multiprotein complex) which is stored in the prespore vesicles (Watson, N., McGuire, V., and Alexander, S (1994) J. Cell Sci. 107, 2567-2579). Later in development, the complex is synchronously secreted from the prespore vesicles and incorporated into the spore coat. We now have shown that the PsB complex has a specific in vitro cellulose binding activity. The analysis of mutants lacking individual subunits of the PsB complex revealed the relative order of assembly of the subunit proteins and demonstrated that the protein subunits must be assembled for cellulose binding activity. These results provide a biochemical explanation for the localization of this multiprotein complex in the spore coat.

Design and characterization of a magnetoelastic sensor for the detection of biological agents

NASA Astrophysics Data System (ADS)

Shen, Wen; Mathison, Leslie C.; Petrenko, Valery A.; Chin, Bryan A.

2010-01-01

This paper presents the design and development of a free-standing, magnetoelastic biosensor. The detection principle is presented and various resonance characteristics of the sensor are discussed. Experimental measurements of the sensor resonance frequencies agree with theoretical predictions. The influence of the external magnetic field on the resonance behaviour of the sensor was studied and the optimum dc magnetic fields for best sensitivity in air and in water solutions for 2000 × 400 × 15 µm (2 mm) sensors and 1000 × 200 × 15 µm (1 mm) size sensors were determined to be 75 Oe and 38 Oe, respectively. Both theoretical prediction and experimental results show that smaller sensors have greater mass sensitivity and can theoretically detect mass as small as one biological spore. The sensor platform was immobilized with JRB7 phages for specific, in vitro detection of B. anthracis spores. Real-time detection of spores suspended in water was demonstrated using a flowing system. The 1 mm and 2 mm sensors were found to have a detection limit of 104 spores ml-1 and 105 spores ml-1, respectively.

Asexual-sexual morph connection in the type species of Berkleasmium.

PubMed

Tanney, Joey; Miller, Andrew N

2017-06-01

Berkleasmium is a polyphyletic genus comprising 37 dematiaceous hyphomycetous species. In this study, independent collections of the type species, B. concinnum , were made from Eastern North America. Nuclear internal transcribed spacer rDNA (ITS) and partial nuc 28S large subunit rDNA (LSU) sequences obtained from collections and subsequent cultures showed that Berkleasmium concinnum is the asexual morph of Neoacanthostigma septoconstrictum ( Tubeufiaceae , Tubeufiales ). Phylogenies inferred from Bayesian inference and maximum likelihood analyses of ITS-LSU sequence data confirmed this asexual-sexual morph connection and a re-examination of fungarium reference specimens also revealed the co-occurrence of N. septoconstrictum ascomata and B. concinnum sporodochia. Neoacanthostigma septoconstrictum is therefore synonymized under B. concinnum on the basis of priority. A specimen identified as N. septoconstrictum from Thailand is described as N. thailandicum sp. nov., based on morphological and genetic distinctiveness.

Autolytic hydrolases affect sexual and asexual development of Aspergillus nidulans.

PubMed

Emri, Tamás; Vékony, Viktória; Gila, Barnabás; Nagy, Flóra; Forgács, Katalin; Pócsi, István

2018-03-30

Radial growth, asexual sporulation, and cleistothecia formation as well as extracellular chitinase and proteinase formation of Aspergillus nidulans were monitored in surface cultures in order to study the physiological role of extracellular hydrolase production in carbon-stressed cultures. We set up carbon-stressed and carbon-overfed experimental conditions by varying the starting glucose concentration within the range of 2.5 and 40 g/L. Glucose starvation induced radial growth and hydrolase production and enhanced the maturation of cleistothecia; meanwhile, glucose-rich conditions enhanced mycelial biomass, conidia, and cleistothecia production. Double deletion of chiB and engA (encoding an extracellular endochitinase and a β-1,3-endoglucanase, respectively) decreased conidia production under carbon-stressed conditions, suggesting that these autolytic hydrolases can support conidia formation by releasing nutrients from the cell wall polysaccharides of dead hyphae. Double deletion of prtA and pepJ (both genes encode extracellular proteases) reduced the number of cleistothecia even under carbon-rich conditions except in the presence of casamino acids, which supports the view that sexual development and amino acid metabolism are tightly connected to each other in this fungus.

In vivo imaging of basement membrane movement: ECM patterning shapes Hydra polyps.

PubMed

Aufschnaiter, Roland; Zamir, Evan A; Little, Charles D; Özbek, Suat; Münder, Sandra; David, Charles N; Li, Li; Sarras, Michael P; Zhang, Xiaoming

2011-12-01

Growth and morphogenesis during embryonic development, asexual reproduction and regeneration require extensive remodeling of the extracellular matrix (ECM). We used the simple metazoan Hydra to examine the fate of ECM during tissue morphogenesis and asexual budding. In growing Hydra, epithelial cells constantly move towards the extremities of the animal and into outgrowing buds. It is not known, whether these tissue movements involve epithelial migration relative to the underlying matrix or whether cells and ECM are displaced as a composite structure. Furthermore, it is unclear, how the ECM is remodeled to adapt to the shape of developing buds and tentacles. To address these questions, we used a new in vivo labeling technique for Hydra collagen-1 and laminin, and tracked the fate of ECM in all body regions of the animal. Our results reveal that Hydra 'tissue movements' are largely displacements of epithelial cells together with associated ECM. By contrast, during the evagination of buds and tentacles, extensive movement of epithelial cells relative to the matrix is observed, together with local ECM remodeling. These findings provide new insights into the nature of growth and morphogenesis in epithelial tissues.

Testing a Hypothesis for the Evolution of Sex

NASA Astrophysics Data System (ADS)

Örçal, Bora; Tüzel, Erkan; Sevim, Volkan; Jan, Naeem; Erzan, Ayşe.

An asexual set of primitive bacteria is simulated with a bit-string Penna model with a Fermi function for survival. A recent hypothesis by Jan, Stauffer, and Moseley on the evolution of sex from asexual cells as a strategy for trying to escape the effects of deleterious mutations is checked. This strategy is found to provide a successful scenario for the evolution of a stable macroscopic sexual population.

Differential evolution of asexual and sexual females in a benign culture environment

PubMed Central

Snell, Terry W.

2013-01-01

Here we report one of the first investigations of evolvability of lifespan and reproduction in metazoans, examining both extrinsic and intrinsic factors. We tested effects on senescence of an environmental variable (simulated lake hydroperiod, the length of time an aquatic habitat is inundated), female reproductive physiology (asexual females that reproduce by ameiosis, versus sexual females reproducing by meiosis), and time in a benign culture environment (minimal, if any, external mortality factors). To do this we established chemostat cultures of the rotifer Brachionus plicatilis s.s., and maintained the cultures for 385 d. Hydroperiod alone or in interaction with the effects of time in the benign environment (season) or reproductive physiology had no significant effect on the net reproductive rate, generation time, or rate of aging. Yet combining animals from both ephemeral and permanent hydroperiods revealed a 26% increase in asexual female lifespan across seasons (23% decrease in the rate of aging) and a 56% increase in asexual fecundity, suggesting that maintenance in benign laboratory conditions leads to slower aging. The relative stasis of traits for sexual females implies an impact of reproductive physiology on evolvability. In addition we found a positive correlation between fecundity and lifespan, suggesting an absence of trade-offs in life history traits in the benign laboratory environment. PMID:24795527

Dynamics of asexual reproduction in flatworms

NASA Astrophysics Data System (ADS)

Schoetz, Eva-Maria; Talbot, Jared; Dunkel, Joern

2010-03-01

Planarians (flatworms) are one of the simplest bilaterally symmetric organisms and famous for their extraordinary regenerative capabilities. One can cut a worm in 100 pieces and after a few weeks one obtains 100 new worms that have reconstructed their entire body, including a central nervous system. This amazing regenerative capability is due to a population of stem cells distributed throughout the planarian body. These stem cells do not only allow the worms to heal without scarring after wounding, they also allow for asexual reproduction: Planarians can split themselves in two, and then regenerate the missing body parts within about a week. Naively, one would think that this kind of asexual reproduction could be captured by simple models that describe cell growth in bacteria or other lower organisms. However, we find that there is much more to the story by monitoring >15 generations of many individuals, as well as the long-term behavior (> 9 months) of worm populations under different environmental conditions, such as population density, temperature, and feeding frequency. Surprisingly, we observe that reproduction decreases with increasing food supply, opposite to the relationship between food and reproduction in other asexually reproducing organisms (e.g. bacteria, yeast), and causing obese worms. Finally, our data allows us to address the question of aging in an organism that is thought to be ``forever young''.

The frequency of sex in fungi

PubMed Central

James, Timothy Y.

2016-01-01

Fungi are a diverse group of organisms with a huge variation in reproductive strategy. While almost all species can reproduce sexually, many reproduce asexually most of the time. When sexual reproduction does occur, large variation exists in the amount of in- and out-breeding. While budding yeast is expected to outcross only once every 10 000 generations, other fungi are obligate outcrossers with well-mixed panmictic populations. In this review, we give an overview of the costs and benefits of sexual and asexual reproduction in fungi, and the mechanisms that evolved in fungi to reduce the costs of either mode. The proximate molecular mechanisms potentiating outcrossing and meiosis appear to be present in nearly all fungi, making them of little use for predicting outcrossing rates, but also suggesting the absence of true ancient asexual lineages. We review how population genetic methods can be used to estimate the frequency of sex in fungi and provide empirical data that support a mixed mode of reproduction in many species with rare to frequent sex in between rounds of mitotic reproduction. Finally, we highlight how these estimates might be affected by the fungus-specific mechanisms that evolved to reduce the costs of sexual and asexual reproduction. This article is part of the themed issue ‘Weird sex: the underappreciated diversity of sexual reproduction’. PMID:27619703

The frequency of sex in fungi.

PubMed

Nieuwenhuis, Bart P S; James, Timothy Y

2016-10-19

Fungi are a diverse group of organisms with a huge variation in reproductive strategy. While almost all species can reproduce sexually, many reproduce asexually most of the time. When sexual reproduction does occur, large variation exists in the amount of in- and out-breeding. While budding yeast is expected to outcross only once every 10 000 generations, other fungi are obligate outcrossers with well-mixed panmictic populations. In this review, we give an overview of the costs and benefits of sexual and asexual reproduction in fungi, and the mechanisms that evolved in fungi to reduce the costs of either mode. The proximate molecular mechanisms potentiating outcrossing and meiosis appear to be present in nearly all fungi, making them of little use for predicting outcrossing rates, but also suggesting the absence of true ancient asexual lineages. We review how population genetic methods can be used to estimate the frequency of sex in fungi and provide empirical data that support a mixed mode of reproduction in many species with rare to frequent sex in between rounds of mitotic reproduction. Finally, we highlight how these estimates might be affected by the fungus-specific mechanisms that evolved to reduce the costs of sexual and asexual reproduction.This article is part of the themed issue 'Weird sex: the underappreciated diversity of sexual reproduction'. © 2016 The Author(s).

Evidence of parasexual activity in "asexual amoebae" Cochliopodium spp. (Amoebozoa): extensive cellular and nuclear fusion.

PubMed

Tekle, Yonas I; Anderson, O Roger; Lecky, Ariel F

2014-09-01

The majority of microbial eukaryotes have long been considered asexual, though new evidence indicates sex, or sexual-like (parasexual) behaviors that deviate from the usual union of two gametes, among other variant aspects. Over a dozen amoebozoans are implicated to have sexual stages. However, the exact mechanism by which sex occurs in these lineages remains elusive. This is mainly due to the diverse quality and cryptic nature of their life cycle. In this study we present evidence of some previously unreported aspects of the life cycle of an amoeba, Cochliopodium, that undergoes unusual intraspecific interactions using light microscopy and immunocytochemistry. Similar to other amoebozoans, Cochliopodium, is considered asexual with no published reports of sex or parasexuality. We also investigated environmental conditions that govern the observed intraspecific interactions. Both light microscopic and immunocytochemistry evidence demonstrates Cochliopodium undergoes cellular fusion (plasmogamy) and nuclear fusion (karyogamy). Large plasmodia eventually undergo karyogamy and contain large fused, polyploid, nuclei. These are observed to fragment, subsequently, by karyotomy (nuclear fission) and cytoplasmic fission to yield uninucleated amoebae. This process could lead to a non-meiotic, parasexual exchange of chromosomes in Cochliopodium. These findings strongly suggest that Cochliopodium is involved in parasexual activity and should no longer be considered strictly asexual. Copyright © 2014 Elsevier GmbH. All rights reserved.

In Vitro Activities of Primaquine-Schizonticide Combinations on Asexual Blood Stages and Gametocytes of Plasmodium falciparum.

PubMed

Cabrera, Mynthia; Cui, Liwang

2015-12-01

Currently, the World Health Organization recommends addition of a 0.25-mg base/kg single dose of primaquine (PQ) to artemisinin combination therapies (ACTs) for Plasmodium falciparum malaria as a gametocytocidal agent for reducing transmission. Here, we investigated the potential interactions of PQ with the long-lasting components of the ACT drugs for eliminating the asexual blood stages and gametocytes of in vitro-cultured P. falciparum strains. Using the SYBR green I assay for asexual parasites and a flow cytometry-based assay for gametocytes, we determined the interactions of PQ with the schizonticides chloroquine, mefloquine, piperaquine, lumefantrine, and naphthoquine. With the sums of fractional inhibitory concentrations and isobolograms, we were able to determine mostly synergistic interactions for the various PQ and schizonticide combinations on the blood stages of P. falciparum laboratory strains. The synergism in inhibiting asexual stages and gametocytes was highly evident with PQ-naphthoquine, whereas synergism was moderate for the PQ-piperaquine, PQ-chloroquine, and PQ-mefloquine combinations. We have detected potentially antagonistic interactions between PQ and lumefantrine under certain drug combination ratios, suggesting that precautions might be needed when PQ is added as the gametocytocide to the artemether-lumefantrine ACT (Coartem). Copyright © 2015, American Society for Microbiology. All Rights Reserved.

The Pattern of Genetic Variability in Apomictic Clones of Taraxacum officinale Indicates the Alternation of Asexual and Sexual Histories of Apomicts

PubMed Central

Majeský, Ľuboš; Vašut, Radim J.; Kitner, Miloslav; Trávníček, Bohumil

2012-01-01

Dandelions (genus Taraxacum) comprise a group of sexual diploids and apomictic polyploids with a complicated reticular evolution. Apomixis (clonal reproduction through seeds) in this genus is considered to be obligate, and therefore represent a good model for studying the role of asexual reproduction in microevolutionary processes of apomictic genera. In our study, a total of 187 apomictic individuals composing a set of nine microspecies (sampled across wide geographic area in Europe) were genotyped for six microsatellite loci and for 162 amplified fragment length polymorphism (AFLP) markers. Our results indicated that significant genetic similarity existed within accessions with low numbers of genotypes. Genotypic variability was high among accessions but low within accessions. Clustering methods discriminated individuals into nine groups corresponding to their phenotypes. Furthermore, two groups of apomictic genotypes were observed, which suggests that they had different asexual histories. A matrix compatibility test suggests that most of the variability within accession groups was mutational in origin. However, the presence of recombination was also detected. The accumulation of mutations in asexual clones leads to the establishment of a network of clone mates. However, this study suggests that the clones primarily originated from the hybridisation between sexual and apomicts. PMID:22870257

Characterization of fungal spores in ambient particulate matter: A study from the Himalayan region

NASA Astrophysics Data System (ADS)

Kumar, Ajay; Attri, Arun K.

2016-10-01

Fungal spores as a constituent of ambient particulate matter (PM) is of concern; they not only display the physical traits of a particle, but are also potential allergens and health risk. An investigation over fourteen month was undertaken at a rural site located in the Western Himalayan region, to evaluate the PM associated fungal spores' concentration and diversity. The season-wise change in the fungal spores concentration in the Coarse Particulate Matter (CPM) fraction (aerodynamic diameter > 10 μm) varied from 500 to 3899 spores m-3. Their average concentration over 14 months was 1517 spores m-3. Significant diversity of fungal spores in the CPM samples was observed; 27 individual genera of fungal spores were identified, of which many were known allergens. Presence of Ascomycota and Basidiomycota fungal spores was dominant in the samples; ∼20% of the spores were un-characterized. The season-wise variability in fungal spores showed a statistically significant high correlation with CPM load. Maximum number concentration of the spores in CPM was recorded in the summer, while minimum in the winter. The high diversity of spores occurred during monsoon and post monsoon months. The meteorological factors played an important role in the fungal spores' distribution profile. The temporal profile of the spores showed significant correlation with the ambient temperature (T), relative humidity (RH), wind speed (WS) and planetary boundary layer (PBL) height. Strong correlation of WS with fungal spores and CPM, and wind back trajectories suggest that re-suspension and wind assisted transport of PM contributes to ambient CPM associated fungal spores.

Viability and infectivity of fresh and cryopreserved Nosema ceranae spores.

PubMed

McGowan, Janine; De la Mora, Alvaro; Goodwin, Paul H; Habash, Marc; Hamiduzzaman, Mollah Md; Kelly, Paul G; Guzman-Novoa, Ernesto

2016-12-01

The microsporidium fungus Nosema ceranae is an intracellular parasite that infects the midgut of the honey bee, Apis mellifera. A major limitation of research on N. ceranae is that the fungus is non-culturable and thus studying it depends on the seasonal availability of Nosema spores. Also, spore viability and infectivity can vary considerably, and thus there is a need for reliable methods for determining those traits. This study examined different conditions for N. ceranae spore cryopreservation at -70°C, assessing spore viability and infectivity. Viability was determined by a staining procedure counting total spores numbers with bright field microscopy and un-viable spore numbers with the fluorescent dye, propidium iodide. Spore infectivity was determined with a dilution inoculation assay. Infectivity was dependent on the inoculum dose for the proportion of bees with detectable Nosema infections based on the number of spores per bee at 18days after inoculation; 4000 spores per bee or higher were needed to get approx. 100% of the inoculated bees infected. The median infective dose (ID 50 ) was 149 spores per bee, and the minimum dose capable of causing a detectable infection was 1.28 spores. The proportion of N. ceranae infected bees correlated significantly with the number of spores per bee (r=0.98, P<0.0001). N. ceranae spores cryopreserved in water or 10% glycerol did not differ in viability compared to fresh spores, but lost infectivity when inoculated into bees. This study shows that while cryopreservation of N. ceranae spores can preserve viability, the spores can have reduced infectivity. Copyright © 2016 Elsevier B.V. All rights reserved.

A cryptic pigment biosynthetic pathway uncovered by heterologous expression is essential for conidial development in Pestalotiopsis fici.

PubMed

Zhang, Peng; Wang, Xiuna; Fan, Aili; Zheng, Yanjing; Liu, Xingzhong; Wang, Shihua; Zou, Huixi; Oakley, Berl R; Keller, Nancy P; Yin, Wen-Bing

2017-08-01

Spore pigmentation is very common in the fungal kingdom. The best studied pigment in fungi is melanin which coats the surface of single cell spores. What and how pigments function in a fungal species with multiple cell conidia is poorly understood. Here, we identified and deleted a polyketide synthase (PKS) gene PfmaE and showed that it is essential for multicellular conidial pigmentation and development in a plant endophytic fungus, Pestalotiopsis fici. To further characterize the melanin pathway, we utilized an advanced Aspergillus nidulans heterologous system for the expression of the PKS PfmaE and the Pfma gene cluster. By structural elucidation of the pathway metabolite scytalone in A. nidulans, we provided chemical evidence that the Pfma cluster synthesizes DHN melanin. Combining genetic deletion and combinatorial gene expression of Pfma cluster genes, we determined that the putative reductase PfmaG and the PKS are sufficient for the synthesis of scytalone. Feeding scytalone back to the P. fici ΔPfmaE mutant restored pigmentation and multicellular adherence of the conidia. These results cement a growing understanding that pigments are essential not simply for protection of spores from biotic and abiotic stresses but also for spore structural development. © 2017 John Wiley & Sons Ltd.

REVERSIBLE ACTIVATION FOR GERMINATION AND SUBSEQUENT CHANGES IN BACTERIAL SPORES1

PubMed Central

Lee, W. H.; Ordal, Z. John

1963-01-01

Lee, W. H. (University of Illinois, Urbana) and Z. John Ordal. Reversible activation for germination and subsequent changes in bacterial spores. J. Bacteriol. 85:207–217. 1963.—It was possible to isolate refractile spores of Bacillus megaterium, from a calcium dipicolinate germination solution, that were activated and would germinate spontaneously in distilled water. Some of the characteristics of the initial phases of bacterial spore germination were determined by studying these unstable activated spores. Activated spores of B. megaterium were resistant to stains and possessed a heat resistance intermediate between that of dormant and of germinated spores. The spontaneous germination of activated spores was inhibited by copper, iron, silver, or mercury salts, saturated o-phenanthroline, or solutions having a low pH value, but not by many common inhibitors. These inhibitions could be partially or completely reversed by the addition of sodium dipicolinate. The activated spores could be deactivated and made similar to dormant spores by treatment with acid. Analyses of the exudates from the variously treated spore suspensions revealed that whatever inhibited the germination of activated spores also inhibited the release of spore material. The composition of the germination exudates was different than that of extracts of dormant spores. Although heavy suspensions of activated spores gradually became swollen and dark when suspended in solutions of o-phenanthroline or at pH 4, the materials released resembled those found in extracts of dormant spores rather than those of normal germination exudates. Images PMID:16561987

Spore production in Paecilomyces lilacinus (Thom.) samson strains on agro-industrial residues

PubMed Central

Robl, Diogo; Sung, Letizia B.; Novakovich, João Henrique; Marangoni, Paulo R.D.; Zawadneak, Maria Aparecida C.; Dalzoto, Patricia R.; Gabardo, Juarez; Pimentel, Ida Chapaval

2009-01-01

Paecilomyces lilacinus has potential for pests control. We aimed to analyze mycelial growth and spore production in P. lilacinus strains in several agro-industrial residues and commercial media. This study suggests alternative nutrient sources for fungi production and that the biotechnological potential of agro-industrial refuses could be employed in byproducts development. PMID:24031361

Predictive model for Clostridium perfringens growth in roast beef during cooling and inhibition of spore germination and outgrowth by organic acid salts.

PubMed

Sánchez-Plata, Marcos X; Amézquita, Alejandro; Blankenship, Erin; Burson, Dennis E; Juneja, Vijay; Thippareddi, Harshavardhan

2005-12-01

Spores of foodborne pathogens can survive traditional thermal processing schedules used in the manufacturing of processed meat products. Heat-activated spores can germinate and grow to hazardous levels when these products are improperly chilled. Germination and outgrowth of Clostridium perfringens spores in roast beef during chilling was studied following simulated cooling schedules normally used in the processed-meat industry. Inhibitory effects of organic acid salts on germination and outgrowth of C. perfringens spores during chilling and the survival of vegetative cells and spores under abusive refrigerated storage was also evaluated. Beef top rounds were formulated to contain a marinade (finished product concentrations: 1% salt, 0.2% potassium tetrapyrophosphate, and 0.2% starch) and then ground and mixed with antimicrobials (sodium lactate and sodium lactate plus 2.5% sodium diacetate and buffered sodium citrate and buffered sodium citrate plus 1.3% sodium diacetate). The ground product was inoculated with a three-strain cocktail of C. perfringens spores (NCTC 8238, NCTC 8239, and ATCC 10388), mixed, vacuum packaged, heat shocked for 20 min at 75 degrees C, and chilled exponentially from 54.5 to 7.2 degrees C in 9, 12, 15, 18, or 21 h. C. perfringens populations (total and spore) were enumerated after heat shock, during chilling, and during storage for up to 60 days at 10 degrees C using tryptose-sulfite-cycloserine agar. C. perfringens spores were able to germinate and grow in roast beef (control, without any antimicrobials) from an initial population of ca. 3.1 log CFU/g by 2.00, 3.44, 4.04, 4.86, and 5.72 log CFU/g after 9, 12, 15, 18, and 21 h of exponential chilling. A predictive model was developed to describe sigmoidal C. perfringens growth curves during cooling of roast beef from 54.5 to 7.2 degrees C within 9, 12, 15, 18, and 21 h. Addition of antimicrobials prevented germination and outgrowth of C. perfringens regardless of the chill times. C. perfringens spores could be recovered from samples containing organic acid salts that were stored up to 60 days at 10 degrees C. Extension of chilling time to > or =9 h resulted in >1 log CFU/g growth of C. perfringens under anaerobic conditions in roast beef. Organic acid salts inhibited outgrowth of C. perfringens spores during chilling of roast beef when extended chill rates were followed. Although C. perfringens spore germination is inhibited by the antimicrobials, this inhibition may represent a hazard when such products are incorporated into new products, such as soups and chili, that do not contain these antimicrobials, thus allowing spore germination and outgrowth under conditions of temperature abuse.

The differential effects of heat-shocking on the viability of spores from Bacillus anthracis, Bacillus subtilis, and Clostridium sporogenes after treatment with peracetic acid- and glutaraldehyde-based disinfectants

PubMed Central

March, Jordon K; Pratt, Michael D; Lowe, Chinn-Woan; Cohen, Marissa N; Satterfield, Benjamin A; Schaalje, Bruce; O'Neill, Kim L; Robison, Richard A

2015-01-01

This study investigated (1) the susceptibility of Bacillus anthracis (Ames strain), Bacillus subtilis (ATCC 19659), and Clostridium sporogenes (ATCC 3584) spores to commercially available peracetic acid (PAA)- and glutaraldehyde (GA)-based disinfectants, (2) the effects that heat-shocking spores after treatment with these disinfectants has on spore recovery, and (3) the timing of heat-shocking after disinfectant treatment that promotes the optimal recovery of spores deposited on carriers. Suspension tests were used to obtain inactivation kinetics for the disinfectants against three spore types. The effects of heat-shocking spores after disinfectant treatment were also determined. Generalized linear mixed models were used to estimate 6-log reduction times for each spore type, disinfectant, and heat treatment combination. Reduction times were compared statistically using the delta method. Carrier tests were performed according to AOAC Official Method 966.04 and a modified version that employed immediate heat-shocking after disinfectant treatment. Carrier test results were analyzed using Fisher's exact test. PAA-based disinfectants had significantly shorter 6-log reduction times than the GA-based disinfectant. Heat-shocking B. anthracis spores after PAA treatment resulted in significantly shorter 6-log reduction times. Conversely, heat-shocking B. subtilis spores after PAA treatment resulted in significantly longer 6-log reduction times. Significant interactions were also observed between spore type, disinfectant, and heat treatment combinations. Immediately heat-shocking spore carriers after disinfectant treatment produced greater spore recovery. Sporicidal activities of disinfectants were not consistent across spore species. The effects of heat-shocking spores after disinfectant treatment were dependent on both disinfectant and spore species. Caution must be used when extrapolating sporicidal data of disinfectants from one spore species to another. Heat-shocking provides a more accurate picture of spore survival for only some disinfectant/spore combinations. Collaborative studies should be conducted to further examine a revision of AOAC Official Method 966.04 relative to heat-shocking. PMID:26185111

Vulnerability of Bacillus spores and of related genera to physical impaction injury with particular reference to spread-plating.

PubMed

Thomas, P; Sekhar, A C; Mujawar, M M

2014-11-01

To examine whether bacterial spores are vulnerable to impaction injury during standard spread-plating or to other modes of physical impaction. Employing heat-challenged spores of Bacillus pumilus, Bacillus subtilis, Bacillus thuringiensis, Lysinibacillus, Paenibacillus and Brevibacillus spp. from day-4 to day-10 nutrient agar (NA) plates in 50% ethanol, plating the spore suspension to the extent of just drying the agar surface on fresh NA (50-60 s; SP-B) was tested in comparison with the spreader-independent approach of spotting-and-tilt-spreading (SATS), or a brief plating (<10 s; SP-A). Spore CFU was significantly reduced with SP-B in different organisms (23-40%) over SATS independent of the spore size. Comparing 4-, 7- and 10-day-old B. pumilus spores, the former two displayed significant CFU reduction in SP-B indicating a spore age-related effect. Continuous plating for 2-5 min showed a reduction in spore CFU in all organisms depending on plating duration. CFU reduction effect with SP-B was less manifest on refrigerated plates where no friction was experienced but acute on prewarmed and surface-dried plates. Spreader movement over agar surface subsequent to the exhaustion of free moisture proved highly detrimental to spores. A simulated plating study by plating the spores over a plastic film till drying showed a significant reduction in spore CFU. DAPI staining and glass bead-vortexing studies confirmed spore disruption through physical impaction. Bacterial spores are vulnerable to injury during spread-plating or with other forms of physical impaction with variable effects on different genotypes independent of the spore size but altered by spore age. Implications during spore CFU estimations employing spread-plating and during spore surveillance, and the recommendation of SATS as an easier and safer alternative for spore CFU enumeration. © 2014 The Society for Applied Microbiology.

Multifactorial Resistance of Bacillus subtilis Spores to High-Energy Proton Radiation: Role of Spore Structural Components and the Homologous Recombination and Non-Homologous End Joining DNA Repair Pathways

PubMed Central

Reitz, Günther; Li, Zuofeng; Klein, Stuart; Nicholson, Wayne L.

2012-01-01

Abstract The space environment contains high-energy charged particles (e.g., protons, neutrons, electrons, α-particles, heavy ions) emitted by the Sun and galactic sources or trapped in the radiation belts. Protons constitute the majority (87%) of high-energy charged particles. Spores of Bacillus species are one of the model systems used for astro- and radiobiological studies. In this study, spores of different Bacillus subtilis strains were used to study the effects of high energetic proton irradiation on spore survival. Spores of the wild-type B. subtilis strain [mutants deficient in the homologous recombination (HR) and non-homologous end joining (NHEJ) DNA repair pathways and mutants deficient in various spore structural components such as dipicolinic acid (DPA), α/β-type small, acid-soluble spore protein (SASP) formation, spore coats, pigmentation, or spore core water content] were irradiated as air-dried multilayers on spacecraft-qualified aluminum coupons with 218 MeV protons [with a linear energy transfer (LET) of 0.4 keV/μm] to various final doses up to 2500 Gy. Spores deficient in NHEJ- and HR-mediated DNA repair were significantly more sensitive to proton radiation than wild-type spores, indicating that both HR and NHEJ DNA repair pathways are needed for spore survival. Spores lacking DPA, α/β-type SASP, or with increased core water content were also significantly more sensitive to proton radiation, whereas the resistance of spores lacking pigmentation or spore coats was essentially identical to that of the wild-type spores. Our results indicate that α/β-type SASP, core water content, and DPA play an important role in spore resistance to high-energy proton irradiation, suggesting their essential function as radioprotectants of the spore interior. Key Words: Bacillus—Spores—DNA repair—Protection—High-energy proton radiation. Astrobiology 12, 1069–1077. PMID:23088412

Species Specific Bacterial Spore Detection Using Lateral-Flow Immunoassay with DPA-Triggered Tb Luminescence

NASA Technical Reports Server (NTRS)

Ponce, Adrian

2003-01-01

A method of detecting bacterial spores incorporates (1) A method of lateral-flow immunoassay in combination with (2) A method based on the luminescence of Tb3+ ions to which molecules of dipicolinic acid (DPA) released from the spores have become bound. The present combination of lateral-flow immunoassay and DPA-triggered Tb luminescence was developed as a superior alternative to a prior lateral-flow immunoassay method in which detection involves the visual observation and/or measurement of red light scattered from colloidal gold nanoparticles. The advantage of the present combination method is that it affords both (1) High selectivity for spores of the species of bacteria that one seeks to detect (a characteristic of lateral-flow immunoassay in general) and (2) Detection sensitivity much greater (by virtue of the use of DPA-triggered Tb luminescence instead of gold nanoparticles) than that of the prior lateral-flow immunoassay method

Quantitative inactivation-mechanisms of P. digitatum and A. niger spores based on atomic oxygen dose

NASA Astrophysics Data System (ADS)

Ito, Masafumi; Hashizume, Hiroshi; Ohta, Takayuki; Hori, Masaru

2014-10-01

We have investigated inactivation mechanisms of Penicillium digitatum and Asperguills niger spores using atmospheric-pressure radical source quantitatively. The radical source was specially developed for supplying only neutral radicals without charged species and UV-light emissions. Reactive oxygen radical densities such as grand-state oxygen atoms, excited-state oxygen molecules and ozone were measured using VUV and UV absorption spectroscopies. The measurements and the treatments of spores were carried out in an Ar-purged chamber for eliminating the influences of OH, NOx and so on. The results revealed that the inactivation of spores can be explained by atomic-oxygen dose under the conditions employing neutral ROS irradiations. On the basis of the dose, we have observed the changes of intracellular organelles and membrane functions using TEM, SEM and confocal- laser fluorescent microscopy. From these results, we discuss the detail inactivation-mechanisms quantitatively based on atomic-oxygen dose.

Avirulent Bacillus anthracis Strain with Molecular Assay Targets as Surrogate for Irradiation-Inactivated Virulent Spores.

PubMed

Plaut, Roger D; Staab, Andrea B; Munson, Mark A; Gebhardt, Joan S; Klimko, Christopher P; Quirk, Avery V; Cote, Christopher K; Buhr, Tony L; Rossmaier, Rebecca D; Bernhards, Robert C; Love, Courtney E; Berk, Kimberly L; Abshire, Teresa G; Rozak, David A; Beck, Linda C; Stibitz, Scott; Goodwin, Bruce G; Smith, Michael A; Sozhamannan, Shanmuga

2018-04-01

The revelation in May 2015 of the shipment of γ irradiation-inactivated wild-type Bacillus anthracis spore preparations containing a small number of live spores raised concern about the safety and security of these materials. The finding also raised doubts about the validity of the protocols and procedures used to prepare them. Such inactivated reference materials were used as positive controls in assays to detect suspected B. anthracis in samples because live agent cannot be shipped for use in field settings, in improvement of currently deployed detection methods or development of new methods, or for quality assurance and training activities. Hence, risk-mitigated B. anthracis strains are needed to fulfill these requirements. We constructed a genetically inactivated or attenuated strain containing relevant molecular assay targets and tested to compare assay performance using this strain to the historical data obtained using irradiation-inactivated virulent spores.

A quasispecies approach to the evolution of sexual replication in unicellular organisms.

PubMed

Tannenbaum, Emmanuel; Fontanari, José F

2008-03-01

This study develops a simplified model describing the evolutionary dynamics of a population composed of obligate sexually and asexually reproducing, unicellular organisms. The model assumes that the organisms have diploid genomes consisting of two chromosomes, and that the sexual organisms replicate by first dividing into haploid intermediates, which then combine with other haploids, followed by the normal mitotic division of the resulting diploid into two new daughter cells. We assume that the fitness landscape of the diploids is analogous to the single-fitness-peak approach often used in single-chromosome studies. That is, we assume a master chromosome that becomes defective with just one point mutation. The diploid fitness then depends on whether the genome has zero, one, or two copies of the master chromosome. We also assume that only pairs of haploids with a master chromosome are capable of combining so as to produce sexual diploid cells, and that this process is described by second-order kinetics. We find that, in a range of intermediate values of the replication fidelity, sexually reproducing cells can outcompete asexual ones, provided the initial abundance of sexual cells is above some threshold value. The range of values where sexual reproduction outcompetes asexual reproduction increases with decreasing replication rate and increasing population density. We critically evaluate a common approach, based on a group selection perspective, used to study the competition between populations and show its flaws in addressing the evolution of sex problem.

Sponge budding is a spatiotemporal morphological patterning process: Insights from synchrotron radiation-based x-ray microtomography into the asexual reproduction of Tethya wilhelma.

PubMed

Hammel, Jörg U; Herzen, Julia; Beckmann, Felix; Nickel, Michael

2009-09-08

Primary agametic-asexual reproduction mechanisms such as budding and fission are present in all non-bilaterian and many bilaterian animal taxa and are likely to be metazoan ground pattern characters. Cnidarians display highly organized and regulated budding processes. In contrast, budding in poriferans was thought to be less specific and related to the general ability of this group to reorganize their tissues. Here we test the hypothesis of morphological pattern formation during sponge budding. We investigated the budding process in Tethya wilhelma (Demospongiae) by applying 3D morphometrics to high resolution synchrotron radiation-based x-ray microtomography (SR-muCT) image data. We followed the morphogenesis of characteristic body structures and identified distinct morphological states which indeed reveal characteristic spatiotemporal morphological patterns in sponge bud development. We discovered the distribution of skeletal elements, canal system and sponge tissue to be based on a sequential series of distinct morphological states. Based on morphometric data we defined four typical bud stages. Once they have reached the final stage buds are released as fully functional juvenile sponges which are morphologically and functionally equivalent to adult specimens. Our results demonstrate that budding in demosponges is considerably more highly organized and regulated than previously assumed. Morphological pattern formation in asexual reproduction with underlying genetic regulation seems to have evolved early in metazoans and was likely part of the developmental program of the last common ancestor of all Metazoa (LCAM).

Sponge budding is a spatiotemporal morphological patterning process: Insights from synchrotron radiation-based x-ray microtomography into the asexual reproduction of Tethya wilhelma

PubMed Central

Hammel, Jörg U; Herzen, Julia; Beckmann, Felix; Nickel, Michael

2009-01-01

Background Primary agametic-asexual reproduction mechanisms such as budding and fission are present in all non-bilaterian and many bilaterian animal taxa and are likely to be metazoan ground pattern characters. Cnidarians display highly organized and regulated budding processes. In contrast, budding in poriferans was thought to be less specific and related to the general ability of this group to reorganize their tissues. Here we test the hypothesis of morphological pattern formation during sponge budding. Results We investigated the budding process in Tethya wilhelma (Demospongiae) by applying 3D morphometrics to high resolution synchrotron radiation-based x-ray microtomography (SR-μCT) image data. We followed the morphogenesis of characteristic body structures and identified distinct morphological states which indeed reveal characteristic spatiotemporal morphological patterns in sponge bud development. We discovered the distribution of skeletal elements, canal system and sponge tissue to be based on a sequential series of distinct morphological states. Based on morphometric data we defined four typical bud stages. Once they have reached the final stage buds are released as fully functional juvenile sponges which are morphologically and functionally equivalent to adult specimens. Conclusion Our results demonstrate that budding in demosponges is considerably more highly organized and regulated than previously assumed. Morphological pattern formation in asexual reproduction with underlying genetic regulation seems to have evolved early in metazoans and was likely part of the developmental program of the last common ancestor of all Metazoa (LCAM). PMID:19737392

Sporulation environment influences spore properties in Bacillus: evidence and insights on underlying molecular and physiological mechanisms.

PubMed

Bressuire-Isoard, Christelle; Broussolle, Véronique; Carlin, Frédéric

2018-05-17

Bacterial spores are resistant to physical and chemical insults, which make them a major concern for public health and for industry. Spores help bacteria to survive extreme environmental conditions that vegetative cells cannot tolerate. Spore resistance and dormancy are important properties for applications in medicine, veterinary health, food safety, crop protection, and other domains. The resistance of bacterial spores results from a protective multilayered structure and from the unique composition of the spore core. The mechanisms of sporulation and germination, the first stage after breaking of dormancy, and organization of spore structure have been extensively studied in Bacillus species. This review aims to illustrate how far the structure, composition and properties of spores are shaped by the environmental conditions in which spores form. We look at the physiological and molecular mechanisms underpinning how sporulation media and environment deeply affect spore yield, spore properties like resistance to wet heat and physical and chemical agents, germination, and further growth. For example, spore core water content decreases as sporulation temperature increases, and resistance to wet heat increases. Controlling the fate of Bacillus spores is pivotal to controlling bacterial risks and process efficiencies in, for example, the food industry, and better control hinges on better understanding how sporulation conditions influence spore properties.

Bacillus anthracis spore movement does not require a carrier cell and is not affected by lethal toxin in human lung models.

PubMed

Booth, J Leland; Duggan, Elizabeth S; Patel, Vineet I; Langer, Marybeth; Wu, Wenxin; Braun, Armin; Coggeshall, K Mark; Metcalf, Jordan P

2016-10-01

The lung is the entry site for Bacillus anthracis in inhalation anthrax, the most deadly form of the disease. Spores escape from the alveolus to regional lymph nodes, germinate and enter the circulatory system to cause disease. The roles of carrier cells and the effects of B. anthracis toxins in this process are unclear. We used a human lung organ culture model to measure spore uptake by antigen presenting cells (APC) and alveolar epithelial cells (AEC), spore partitioning between these cells, and the effects of B. anthracis lethal toxin and protective antigen. We repeated the study in a human A549 alveolar epithelial cell model. Most spores remained unassociated with cells, but the majority of cell-associated spores were in AEC, not in APC. Spore movement was not dependent on internalization, although the location of internalized spores changed in both cell types. Spores also internalized in a non-uniform pattern. Toxins affected neither transit of the spores nor the partitioning of spores into AEC and APC. Our results support a model of spore escape from the alveolus that involves spore clustering with transient passage through intact AEC. However, subsequent transport of spores by APC from the lung to the lymph nodes may occur. Published by Elsevier Masson SAS.

Inactivation of chemical and heat-resistant spores of Bacillus and Geobacillus by nitrogen cold atmospheric plasma evokes distinct changes in morphology and integrity of spores.

PubMed

van Bokhorst-van de Veen, Hermien; Xie, Houyu; Esveld, Erik; Abee, Tjakko; Mastwijk, Hennie; Nierop Groot, Masja

2015-02-01

Bacterial spores are resistant to severe conditions and form a challenge to eradicate from food or food packaging material. Cold atmospheric plasma (CAP) treatment is receiving more attention as potential sterilization method at relatively mild conditions but the exact mechanism of inactivation is still not fully understood. In this study, the biocidal effect by nitrogen CAP was determined for chemical (hypochlorite and hydrogen peroxide), physical (UV) and heat-resistant spores. The three different sporeformers used are Bacillus cereus a food-borne pathogen, and Bacillus atrophaeus and Geobacillus stearothermophilus that are used as biological indicators for validation of chemical sterilization and thermal processes, respectively. The different spores showed variation in their degree of inactivation by applied heat, hypochlorite, hydrogen peroxide, and UV treatments, whereas similar inactivation results were obtained with the different spores treated with nitrogen CAP. G. stearothermophilus spores displayed high resistance to heat, hypochlorite, hydrogen peroxide, while for UV treatment B. atrophaeus spores are most tolerant. Scanning electron microscopy analysis revealed distinct morphological changes for nitrogen CAP-treated B. cereus spores including etching effects and the appearance of rough spore surfaces, whereas morphology of spores treated with heat or disinfectants showed no such changes. Moreover, microscopy analysis revealed CAP-exposed B. cereus spores to turn phase grey conceivably because of water influx indicating damage of the spores, a phenomenon that was not observed for non-treated spores. In addition, data are supplied that exclude UV radiation as determinant of antimicrobial activity of nitrogen CAP. Overall, this study shows that nitrogen CAP treatment has a biocidal effect on selected Bacillus and Geobacillus spores associated with alterations in spore surface morphology and loss of spore integrity. Copyright © 2014 Elsevier Ltd. All rights reserved.

Understanding of the importance of the spore coat structure and pigmentation in the Bacillus subtilis spore resistance to low-pressure plasma sterilization

NASA Astrophysics Data System (ADS)

Raguse, Marina; Fiebrandt, Marcel; Denis, Benjamin; Stapelmann, Katharina; Eichenberger, Patrick; Driks, Adam; Eaton, Peter; Awakowicz, Peter; Moeller, Ralf

2016-07-01

Low-pressure plasmas have been evaluated for their potential in biomedical and defense purposes. The sterilizing effect of plasma can be attributed to several active agents, including (V)UV radiation, charged particles, radical species, neutral and excited atoms and molecules, and the electric field. Spores of Bacillus subtilis were used as a bioindicator and a genetic model system to study the sporicidal effects of low-pressure plasma decontamination. Wild-type spores, spores lacking the major protective coat layers (inner, outer, and crust), pigmentation-deficient spores or spore impaired in encasement (a late step in coat assembly) were systematically tested for their resistance to low-pressure argon, hydrogen, and oxygen plasmas with and without admixtures. We demonstrate that low-pressure plasma discharges of argon and oxygen discharges cause significant physical damage to spore surface structures as visualized by atomic force microscopy. Spore resistance to low-pressure plasma was primarily dependent on the presence of the inner, and outer spore coat layers as well as spore encasement, with minor or less importance of the crust and spore pigmentation, whereas spore inactivation itself was strongly influenced by the gas composition and operational settings.

A new method to evaluate the biocontrol potential of single spore isolates of fungal entomopathogens

PubMed Central

Posada, Francisco J.; Vega, Fernando E.

2005-01-01

Fifty Beauveria bassiana (Balsamo) Vuillemin (Ascomycota: Hypocreales) strains isolated from the coffee berry borer were used to develop a novel screening method aimed at selecting strains with the highest biocontrol potential. The screening method is based on percent insect mortality, average survival time, mortality distribution, percent spore germination, fungal life cycle duration, and spore production on the insect. Based on these parameters, only 11 strains merited further study. The use of a sound scientific protocol for the selection of promising fungal entomopathogens should lead to more efficient use of time, labor, and financial resources in biological control programs. PMID:17119619

Transcriptional regulation of fksA, a β-1,3-glucan synthase gene, by the APSES protein StuA during Aspergillus nidulans development.

PubMed

Park, Bum-Chan; Park, Yun-Hee; Yi, Soohyun; Choi, Yu Kyung; Kang, Eun-Hye; Park, Hee-Moon

2014-11-01

The temporal and spatial regulation of β-1,3-glucan synthesis plays an important role in morphogenesis during fungal growth and development. Northern blot analysis showed that the transcription of fksA, the gene encoding β-1,3-glucan synthase in Aspergillus nidulans, was cell-cycle-dependent and increased steadily over the duration of the vegetative period, but its overall expression during the asexual and sexual stages was fairly constant up until the time of transcription cessation. In an A. nidulans strain mutated in the eukaryotic bHLH-like APSES transcription factor stuA1, the transcriptional level of fksA, and consequently the content of alkali-insoluble cell wall β-glucan, significantly increased at the conidial chain formation and maturation stage. Electrophoretic mobility shift assays revealed that StuA was bound to StREs (StuA Response Elements) on the fksA promoter region. Promoter analysis with sGFP-fusion constructs also indicated the negative regulation of fksA expression by StuA, especially during asexual development. Taken together, these data suggest that StuA plays an important role in cell wall biogenesis during the development of A. nidulans, by controlling the transcription level of fksA.

Ploidy, sex and crossing over in an evolutionary aging model

NASA Astrophysics Data System (ADS)

Lobo, Matheus P.; Onody, Roberto N.

2006-02-01

Nowadays, many forms of reproduction coexist in nature: Asexual, sexual, apomictic and meiotic parthenogenesis, hermaphroditism and parasex. The mechanisms of their evolution and what made them successful reproductive alternatives are very challenging and debated questions. Here, using a simple evolutionary aging model, we give a possible scenario. By studying the performance of populations where individuals may have diverse characteristics-different ploidies, sex with or without crossing over, as well as the absence of sex-we find an evolution sequence that may explain why there are actually two major or leading groups: Sexual and asexual. We also investigate the dependence of these characteristics on different conditions of fertility and deleterious mutations. Finally, if the primeval organisms on Earth were, in fact, asexual individuals we conjecture that the sexual form of reproduction could have more easily been set and found its niche during a period of low-intensity mutations.

Transposable elements in sexual and ancient asexual taxa

PubMed Central

Arkhipova, Irina; Meselson, Matthew

2000-01-01

Sexual reproduction allows deleterious transposable elements to proliferate in populations, whereas the loss of sex, by preventing their spread, has been predicted eventually to result in a population free of such elements [Hickey, D. A. (1982) Genetics 101, 519–531]. We tested this expectation by screening representatives of a majority of animal phyla for LINE-like and gypsy-like reverse transcriptases and mariner/Tc1-like transposases. All species tested positive for reverse transcriptases except rotifers of the class Bdelloidea, the largest eukaryotic taxon in which males, hermaphrodites, and meiosis are unknown and for which ancient asexuality is supported by molecular genetic evidence. Mariner-like transposases are distributed sporadically among species and are present in bdelloid rotifers. The remarkable lack of LINE-like and gypsy-like retrotransposons in bdelloids and their ubiquitous presence in other taxa support the view that eukaryotic retrotransposons are sexually transmitted nuclear parasites and that bdelloid rotifers evolved asexually. PMID:11121049

Bacteria, mould and yeast spore inactivation studies by scanning electron microscope observations.

PubMed

Rozali, Siti N M; Milani, Elham A; Deed, Rebecca C; Silva, Filipa V M

2017-12-18

Spores are the most resistant form of microbial cells, thus difficult to inactivate. The pathogenic or food spoilage effects of certain spore-forming microorganisms have been the primary basis of sterilization and pasteurization processes. Thermal sterilization is the most common method to inactivate spores present on medical equipment and foods. High pressure processing (HPP) is an emerging and commercial non-thermal food pasteurization technique. Although previous studies demonstrated the effectiveness of thermal and non-thermal spore inactivation, the in-depth mechanisms of spore inactivation are as yet unclear. Live and dead forms of two food spoilage bacteria, a mould and a yeast were examined using scanning electron microscopy before and after the inactivation treatment. Alicyclobacillus acidoterrestris and Geobacillus stearothermophilus bacteria are indicators of acidic foods pasteurization and sterilization processes, respectively. Neosartorya fischeri is a phyto-pathogenic mould attacking fruits. Saccharomyces cerevisiae is a yeast with various applications for winemaking, brewing, baking and the production of biofuel from crops (e.g. sugar cane). Spores of the four microbial species were thermally inactivated. Spores of S. cerevisiae were observed in the ascus and free form after thermal and HPP treatments. Different forms of damage and cell destruction were observed for each microbial spore. Thermal treatment inactivated bacterial spores of A. acidoterrestris and G. stearothermophilus by attacking the inner core of the spore. The heat first altered the membrane permeability allowing the release of intracellular components. Subsequently, hydration of spores, physicochemical modifications of proteins, flattening and formation of indentations occurred, with subsequent spore death. Regarding N. fischeri, thermal inactivation caused cell destruction and leakage of intracellular components. Both thermal and HPP treatments of S. cerevisiae free spores attacked the inner membrane, altering its permeability, and allowing in final stages the transfer of intracellular components to the outside. The spore destruction caused by thermal treatment was more severe than HPP, as HPP had less effect on the spore core. All injured spores have undergone irreversible volume and shape changes. While some of the leakage of spore contents is visible around the deformed but fully shaped spore, other spores exhibited large indentations and were completely deformed, apparently without any contents inside. This current study contributed to the understanding of spore inactivation by thermal and non-thermal processes. Copyright © 2017 Elsevier B.V. All rights reserved.

Monitoring airborne fungal spores in an experimental indoor environment to evaluate sampling methods and the effects of human activity on air sampling.

PubMed Central

Buttner, M P; Stetzenbach, L D

1993-01-01

Aerobiological monitoring was conducted in an experimental room to aid in the development of standardized sampling protocols for airborne microorganisms in the indoor environment. The objectives of this research were to evaluate the relative efficiencies of selected sampling methods for the retrieval of airborne fungal spores and to determine the effect of human activity on air sampling. Dry aerosols containing known concentrations of Penicillium chrysogenum spores were generated, and air samples were taken by using Andersen six-stage, Surface Air System, Burkard, and depositional samplers. The Andersen and Burkard samplers retrieved the highest numbers of spores compared with the measurement standard, an aerodynamic particle sizer located inside the room. Data from paired samplers demonstrated that the Andersen sampler had the highest levels of sensitivity and repeatability. With a carpet as the source of P. chrysogenum spores, the effects of human activity (walking or vacuuming near the sampling site) on air sampling were also examined. Air samples were taken under undisturbed conditions and after human activity in the room. Human activity resulted in retrieval of significantly higher concentrations of airborne spores. Surface sampling of the carpet revealed moderate to heavy contamination despite relatively low airborne counts. Therefore, in certain situations, air sampling without concomitant surface sampling may not adequately reflect the level of microbial contamination in indoor environments. PMID:8439150

Asynchronous spore germination in isogenic natural isolates of Saccharomyces paradoxus.

PubMed

Stelkens, Rike B; Miller, Eric L; Greig, Duncan

2016-05-01

Spores from wild yeast isolates often show great variation in the size of colonies they produce, for largely unknown reasons. Here we measure the colonies produced from single spores from six different wild Saccharomyces paradoxus strains. We found remarkable variation in spore colony sizes, even among spores that were genetically identical. Different strains had different amounts of variation in spore colony sizes, and variation was not affected by the number of preceding meioses, or by spore maturation time. We used time-lapse photography to show that wild strains also have high variation in spore germination timing, providing a likely mechanism for the variation in spore colony sizes. When some spores from a laboratory strain make small colonies, or no colonies, it usually indicates a genetic or meiotic fault. Here, we demonstrate that in wild strains spore colony size variation is normal. We discuss and assess potential adaptive and non-adaptive explanations for this variation. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Role of Visible Light-Activated Photocatalyst on the Reduction of Anthrax Spore-Induced Mortality in Mice

PubMed Central

Huang, Hsin-Hsien; Wong, Ming-Show; Lin, Hung-Chi; Chang, Hsin-Hou

2009-01-01

Background Photocatalysis of titanium dioxide (TiO2) substrates is primarily induced by ultraviolet light irradiation. Anion-doped TiO2 substrates were shown to exhibit photocatalytic activities under visible-light illumination, relative environmentally-friendly materials. Their anti-spore activity against Bacillus anthracis, however, remains to be investigated. We evaluated these visible-light activated photocatalysts on the reduction of anthrax spore-induced pathogenesis. Methodology/Principal Findings Standard plating method was used to determine the inactivation of anthrax spore by visible light-induced photocatalysis. Mouse models were further employed to investigate the suppressive effects of the photocatalysis on anthrax toxin- and spore-mediated mortality. We found that anti-spore activities of visible light illuminated nitrogen- or carbon-doped titania thin films significantly reduced viability of anthrax spores. Even though the spore-killing efficiency is only approximately 25%, our data indicate that spores from photocatalyzed groups but not untreated groups have a less survival rate after macrophage clearance. In addition, the photocatalysis could directly inactivate lethal toxin, the major virulence factor of B. anthracis. In agreement with these results, we found that the photocatalyzed spores have tenfold less potency to induce mortality in mice. These data suggest that the photocatalysis might injury the spores through inactivating spore components. Conclusion/Significance Photocatalysis induced injuries of the spores might be more important than direct killing of spores to reduce pathogenicity in the host. PMID:19132100

Optical and biological properties of plasma-treated Neurospora crassa spores as studied by absorption, circular dichroism, and Raman spectroscopy

NASA Astrophysics Data System (ADS)

Lee, Geon Joon; Park, Gyungsoon; Choi, Eun Ha

2017-11-01

We studied the effect of plasma treatment on the optical, structural and biological properties of Neurospora crassa ( N. crassa) spores. An atmospheric-pressure plasma jet (APPJ) was used to generate reactive oxygen and nitrogen species in aqueous solution. The APPJ treatment of N. crassa spores in water significantly reduced the viability of spores. The reduction in the spore viability can be attributed to the reactive species from the plasma itself and those derived from the reaction of plasma radicals with aqueous solution. These structural modifications were contingent on the medium in which N. crassa spores were suspended; plasma treatment of N. crassa spores in PBS did not significantly affect the viability of spores as compared with N. crassa spores in water. Scanning electron microscopy images and circular dichroism spectra indicated that the spore cell wall was damaged by plasma treatment. The optical absorption spectrum of untreated N. crassa spores exhibited two resonance absorption bands at approximately λ1 ≈ 260 nm and λ2 ≈ 472 nm, originating from deoxyribonucleic acid (DNA) and β-carotene. The Raman spectrum of untreated N. crassa spores exhibited three main peaks at 1519, 1157 and 1006 cm -1, attributed to β-carotene inside the cell wall. The Raman spectra showed that the APPJ treatment of N. crassa spores in water caused degradation of β-carotene, affecting the viability of spores.

Characterization of a commercialized SERS-active substrate and its application to the identification of intact Bacillus endospores

NASA Astrophysics Data System (ADS)

Alexander, Troy A.; Le, Dianna M.

2007-06-01

Surface-enhanced-Raman-spectroscopy (SERS) can be made an attractive approach for the identification of Raman-active compounds and biological materials (i.e., toxins, viruses, or intact bacterial cells or spores) through development of reproducible, spatially uniform SERS-active substrates. Recently, reproducible (from substrate to substrate), spatially homogeneous (over large areas) SERS-active substrates have been commercialized and are now available in the marketplace. Scanning electron microscopy and high-resolution, tapping-mode atomic force microscopy have been used to analyze these novel plasmonic surfaces for topographical consistency. Additionally, we have assessed, by wavelength-tunable microreflectance spectrometry, the spatial distribution of the localized surface plasmon resonance (LSPR) across a single substrate surface as well as the LSPR λMAX variance from substrate to substrate. These analyses reveal that these surfaces are topologically uniform with small LSPR variance from substrate to substrate. Further, we have utilized these patterned surfaces to acquire SERS spectral signatures of four intact, genetically distinct Bacillus spore species cultivated under identical growth conditions. Salient spectral signature features make it possible to discriminate among these genetically distinct spores. Additionally, partial least squares, a multivariate calibration method, has been used to develop personal-computer-borne algorithms useful for classification of unknown spore samples based solely on SERS spectral signatures. To our knowledge, this is the first report detailing application of these commercially available SERS-active substrates to identification of intact Bacillus spores.

Sensing Red, White & Blue: Using Spores of the Sensitive Fern to Introduce Important Concepts in Biology

ERIC Educational Resources Information Center

Kiss, Helen G.; Kiss, John Z.

2005-01-01

Contrary to popular belief, plants are very much in tune and in time with their immediate environment. The most important environmental cues for plants are light and gravity. In this article, the authors discuss the effects of light on plant development and use the spores of the sensitive fern (Onoclea sensibilis) in laboratory exercises to…

The differential effects of heat-shocking on the viability of spores from Bacillus anthracis, Bacillus subtilis, and Clostridium sporogenes after treatment with peracetic acid- and glutaraldehyde-based disinfectants.

PubMed

March, Jordon K; Pratt, Michael D; Lowe, Chinn-Woan; Cohen, Marissa N; Satterfield, Benjamin A; Schaalje, Bruce; O'Neill, Kim L; Robison, Richard A

2015-10-01

This study investigated (1) the susceptibility of Bacillus anthracis (Ames strain), Bacillus subtilis (ATCC 19659), and Clostridium sporogenes (ATCC 3584) spores to commercially available peracetic acid (PAA)- and glutaraldehyde (GA)-based disinfectants, (2) the effects that heat-shocking spores after treatment with these disinfectants has on spore recovery, and (3) the timing of heat-shocking after disinfectant treatment that promotes the optimal recovery of spores deposited on carriers. Suspension tests were used to obtain inactivation kinetics for the disinfectants against three spore types. The effects of heat-shocking spores after disinfectant treatment were also determined. Generalized linear mixed models were used to estimate 6-log reduction times for each spore type, disinfectant, and heat treatment combination. Reduction times were compared statistically using the delta method. Carrier tests were performed according to AOAC Official Method 966.04 and a modified version that employed immediate heat-shocking after disinfectant treatment. Carrier test results were analyzed using Fisher's exact test. PAA-based disinfectants had significantly shorter 6-log reduction times than the GA-based disinfectant. Heat-shocking B. anthracis spores after PAA treatment resulted in significantly shorter 6-log reduction times. Conversely, heat-shocking B. subtilis spores after PAA treatment resulted in significantly longer 6-log reduction times. Significant interactions were also observed between spore type, disinfectant, and heat treatment combinations. Immediately heat-shocking spore carriers after disinfectant treatment produced greater spore recovery. Sporicidal activities of disinfectants were not consistent across spore species. The effects of heat-shocking spores after disinfectant treatment were dependent on both disinfectant and spore species. Caution must be used when extrapolating sporicidal data of disinfectants from one spore species to another. Heat-shocking provides a more accurate picture of spore survival for only some disinfectant/spore combinations. Collaborative studies should be conducted to further examine a revision of AOAC Official Method 966.04 relative to heat-shocking. © 2015 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

New amino acid germinants for spores of the enterotoxigenic Clostridium perfringens type A isolates.

PubMed

Udompijitkul, Pathima; Alnoman, Maryam; Banawas, Saeed; Paredes-Sabja, Daniel; Sarker, Mahfuzur R

2014-12-01

Clostridium perfringens spore germination plays a critical role in the pathogenesis of C. perfringens-associated food poisoning (FP) and non-food-borne (NFB) gastrointestinal diseases. Germination is initiated when bacterial spores sense specific nutrient germinants (such as amino acids) through germinant receptors (GRs). In this study, we aimed to identify and characterize amino acid germinants for spores of enterotoxigenic C. perfringens type A. The polar, uncharged amino acids at pH 6.0 efficiently induced germination of C. perfringens spores; L-asparagine, L-cysteine, L-serine, and L-threonine triggered germination of spores of most FP and NFB isolates; whereas, L-glutamine was a unique germinant for FP spores. For cysteine- or glutamine-induced germination, gerKC spores (spores of a gerKC mutant derivative of FP strain SM101) germinated to a significantly lower extent and released less DPA than wild type spores; however, a less defective germination phenotype was observed in gerAA or gerKB spores. The germination defects in gerKC spores were partially restored by complementing the gerKC mutant with a recombinant plasmid carrying wild-type gerKA-KC, indicating that GerKC is an essential GR protein. The gerKA, gerKC, and gerKB spores germinated significantly slower with L-serine and L-threonine than their parental strain, suggesting the requirement for these GR proteins for normal germination of C. perfringens spores. In summary, these results indicate that the polar, uncharged amino acids at pH 6.0 are effective germinants for spores of C. perfringens type A and that GerKC is the main GR protein for germination of spores of FP strain SM101 with L-cysteine, L-glutamine, and L-asparagine. Copyright © 2014 Elsevier Ltd. All rights reserved.

Influence of spore moisture content on the dry-heat resistance of Bacillus subtilis var. niger.

PubMed

Angelotti, R; Maryanski, J H; Butler, T F; Peeler, J T; Campbell, J E

1968-05-01

The dry-heat resistance of Bacillus subtilis var. niger spores located in or on various materials was determined as D and z values in the range of 105 through 160 C. The systems tested included spores located on steel and paper strips, spores located between stainless-steel washers mated together under 150 inch-lb and 12 inch-lb of torque, and spores encapsulated in methylmethacrylate and epoxy plastics. D values for a given temperature varied with the test system. High D values were observed for the systems in which spores were encapsulated or under heavy torque, whereas lower D values were observed for the steel and paper strip systems and the lightly torqued system. Similar z values were obtained for the plastic and steel strip systems (z(D) = 21 C), but an unusually low z for spores on paper (z(D) = 12.9 C) and an unusually high z for spores on steel washers mated at 150 inch-lb of torque (z(D) = 32 C) were observed. The effect of spore moisture content on the D value of spores encapsulated in water-impermeable plastic was determined, and maximal resistance was observed for spores with a water activity (a(w)) of 0.2 to 0.4. Significantly decreased D values were observed for spores with moisture contents below a(w) 0.2 or above a(w) 0.4. The data indicate that the important factors to be considered when measuring the dry heat resistance of spores are (i) the initial moisture content of the spore, (ii) the rate of spore desiccation during heating, (iii) the water retention capacity of the material in or on which spores are located, and (iv) the relative humidity of the system at the test temperature.

Imaging bacterial spores by soft-x-ray microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stead, A.D.; Ford, T.W.; Judge, J.

1997-04-01

Bacterial spores are able to survive dehydration, but neither the physiological nor structural basis of this have been fully elucidated. Furthermore, once hydrated, spores often require activation before they will germinate. Several treatments can be used to activate spores, but in the case of Bacillus subtlis the most effective is heat treatment. The physiological mechanism associated with activation is also not understood, but some workers suggest that the loss of calcium from the spores may be critical. However, just prior to germination, the spores change from being phase bright to phase dark when viewed by light microscopy. Imaging spores bymore » soft x-ray microscopy is possible without fixation. Thus, in contrast to electron microscopy, it is possible to compare the structure of dehydrated and hydrated spores in a manner not possible previously. A further advantage is that it is possible to monitor individual spores by phase contrast light microscopy immediately prior to imaging with soft x-rays; whereas, with both electron microscopy and biochemical studies, it is a population of spores being studied without knowledge of the phase characteristics of individual spores. This study has therefore tried to compare dehydrated and hydrated spores and to determine if there is a mass loss from individual spores as they pass the transition from being phase bright to phase dark.« less

Morphogenetic Pathway of Spore Wall Assembly in Saccharomyces cerevisiae

PubMed Central

Coluccio, Alison; Bogengruber, Edith; Conrad, Michael N.; Dresser, Michael E.; Briza, Peter; Neiman, Aaron M.

2004-01-01

The Saccharomyces cerevisiae spore is protected from environmental damage by a multilaminar extracellular matrix, the spore wall, which is assembled de novo during spore formation. A set of mutants defective in spore wall assembly were identified in a screen for mutations causing sensitivity of spores to ether vapor. The spore wall defects in 10 of these mutants have been characterized in a variety of cytological and biochemical assays. Many of the individual mutants are defective in the assembly of specific layers within the spore wall, leading to arrests at discrete stages of assembly. The localization of several of these gene products has been determined and distinguishes between proteins that likely are involved directly in spore wall assembly and probable regulatory proteins. The results demonstrate that spore wall construction involves a series of dependent steps and provide the outline of a morphogenetic pathway for assembly of a complex extracellular structure. PMID:15590821

Inactivation of Bacillus subtilis spores by high pressure CO2 with high temperature.

PubMed

Rao, Lei; Xu, Zhenzhen; Wang, Yongtao; Zhao, Feng; Hu, Xiaosong; Liao, Xiaojun

2015-07-16

The objective of this study was to investigate the inactivation of the Bacillus subtilis spores by high pressure CO2 combined with high temperature (HPCD+HT) and to analyze the clumping effect of the spores on their HPCD+HT resistance. The spores of B. subtilis were subjected to heat at 0.1 MPa and HPCD at 6.5-25 MPa, and 82 °C, 86 °C, and 91 °C for 0-120 min. The spores were effectively inactivated by HPCD+HT, but a protective effect on the spores was also found, which was closely correlated to the pressure, temperature and time. The spores treated by HPCD+HT at 6.5 and 10 MPa exhibited a two-stage inactivation curve of shoulder and log-linear regions whereas the spores at 15-25 MPa exhibited a three-stage inactivation curve of shoulder, log-linear and tailing regions, and these curves were well fitted to the Geeraerd model. Approximately 90% of pyridine-2,6-dicarboxylic acid (DPA) was released after HPCD+HT and the 90% DPA release time depend on the pressure and temperature. Moreover, the spore clumping in suspensions was examined by dynamic light scattering. The particle size of the spore suspensions increased with the increase of pressure, temperature and time, indicating the spore clumping. 0.1% Tween 80 as a surfactant inhibited the spore clumping and increased the inactivation ratio of the spores by HPCD+HT. These results indicated that the spore clumping enhanced the spores' resistance to HPCD+HT and induced a protective effect. Copyright © 2015 Elsevier B.V. All rights reserved.

Monitoring of Commitment, Blocking, and Continuation of Nutrient Germination of Individual Bacillus subtilis Spores

PubMed Central

Zhang, Pengfei; Liang, Jintao; Yi, Xuan; Setlow, Peter

2014-01-01

Short exposures of Bacillus spores to nutrient germinants can commit spores to germinate when germinants are removed or their binding to the spores' nutrient germinant receptors (GRs) is inhibited. Bacillus subtilis spores were exposed to germinants for various periods, followed by germinant removal to prevent further commitment. Release of spore dipicolinic acid (DPA) was then measured by differential interference contrast microscopy to monitor germination of multiple individual spores, and spores did not release DPA after 1 to 2 min of germinant exposure until ∼7 min after germinant removal. With longer germinant exposures, percentages of committed spores with times for completion of DPA release (Trelease) greater than the time of germinant removal (Tb) increased, while the time Tlag − Tb, where Tlag represents the time when rapid DPA release began, was decreased but rapid DPA release times (ΔTrelease = Trelease − Tlag) were increased; Factors affecting average Trelease values and the percentages of committed spores were germinant exposure time, germinant concentration, sporulation conditions, and spore heat activation, as previously shown for commitment of spore populations. Surprisingly, germination of spores given a 2nd short germinant exposure 30 to 45 min after a 1st exposure of the same duration was significantly higher than after the 1st exposure, but the number of spores that germinated in the 2nd germinant exposure decreased as the interval between germinant exposures increased up to 12 h. The latter results indicate that spores have some memory, albeit transient, of their previous exposure to nutrient germinants. PMID:24769693

Spore formation in Myxococcus xanthus is tied to cytoskeleton functions and polysaccharide spore coat deposition

PubMed Central

Müller, Frank D.; Schink, Christian W.; Hoiczyk, Egbert; Cserti, Emöke; Higgs, Penelope I.

2011-01-01

Summary Myxococcus xanthus is a Gram-negative bacterium that differentiates into environmentally resistant spores. Spore differentiation involves septation-independent remodelling of the rod-shaped vegetative cell into a spherical spore and deposition of a thick and compact spore coat outside of the outer membrane. Our analyses suggest that spore coat polysaccharides are exported to the cell surface by the Exo outer membrane polysaccharide export/polysaccharide co-polymerase 2a (OPX/PCP-2a) machinery. Conversion of the capsule-like polysaccharide layer into a compact spore coat layer requires the Nfs proteins which likely form a complex in the cell envelope. Mutants in either nfs, exo, or two other genetic loci encoding homologs of polysaccharide synthesis enzymes, fail to complete morphogenesis from rods to spherical spores and instead produce a transient state of deformed cell morphology before reversion into typical rods. We additionally provide evidence that the cell cytoskeletal protein, MreB, plays an important role in rod to spore morphogenesis and for spore outgrowth. These studies provide evidence that this novel gram-negative differentiation process is tied to cytoskeleton functions and polysaccharide spore coat deposition. PMID:22188356

Spore ornamentation of Haplosporidium pickfordi Barrow, 1961 (Haplosporidia), a parasite of freshwater snails in Michigan, USA.

PubMed

Burreson, E M

2001-01-01

Spore ornamentation is increasingly recognized as a key character for species differentiation and genus assignment in the phylum Haplosporidia. Unfortunately, spore ornamentation is known for only a small number of described species so it is difficult to assign most species to genera with any confidence. Scanning and transmission electron microscopy were used to determine the presence and morphology of spore ornamentation of Haplosporidium pickfordi collected from the digestive gland of the snail Physella parkeri in Douglas Lake, Michigan. Spores possess filaments that are derived from the spore wall and originate from two separate areas at the posterior end of the spore. When spores are first isolated from host tissue, filaments are fused into a sheet that wraps around the spore, passing under the opercular lid. These filaments gradually unravel when spores are held in water and after about 14 d most filaments project freely from the posterior end of the spore. The number of filaments could not be determined with certainty, but appears to be approximately nine. Filaments are 100 nm in diam. and up to 50 microm in length. The presence of spore wall-derived filaments confirms the placement of the parasite in the genus Haplosporidium.

Multigeneration Cross-Contamination of Mail with Bacillus anthracis Spores

PubMed Central

Edmonds, Jason; Lindquist, H. D. Alan; Sabol, Jonathan; Martinez, Kenneth; Shadomy, Sean; Cymet, Tyler; Emanuel, Peter

2016-01-01

The release of biological agents, including those which could be used in biowarfare or bioterrorism in large urban areas, has been a concern for governments for nearly three decades. Previous incidents from Sverdlosk and the postal anthrax attack of 2001 have raised questions on the mechanism of spread of Bacillus anthracis spores as an aerosol or contaminant. Prior studies have demonstrated that Bacillus atrophaeus is easily transferred through simulated mail handing, but no reports have demonstrated this ability with Bacillus anthracis spores, which have morphological differences that may affect adhesion properties between spore and formite. In this study, equipment developed to simulate interactions across three generations of envelopes subjected to tumbling and mixing was used to evaluate the potential for cross-contamination of B. anthracis spores in simulated mail handling. In these experiments, we found that the potential for cross-contamination through letter tumbling from one generation to the next varied between generations while the presence of a fluidizer had no statistical impact on the transfer of material. Likewise, the presence or absence of a fluidizer had no statistically significant impact on cross-contamination levels or reaerosolization from letter opening. PMID:27123934

UV-Resistant Non-Spore-Forming Bacteria From Spacecraft-Assembly Facilities

NASA Technical Reports Server (NTRS)

Venkateswaran, Kasthuri

2008-01-01

Four species of non-spore-forming bacteria collected from clean-room surfaces in spacecraft-assembly facilities could survive doses of ultraviolet (UV) radiation that would suffice to kill most known cultivable bacterial species. In a previous study, high UV resistance was found in spores of the SAFR-032 strain of Bacillus pumilus, as reported in "Ultraviolet- Resistant Bacterial Spores," NASA Tech Briefs, Vol. 31, No. 9 (September 2007), page 94. These studies are parts of a continuing effort to understand the survival of hardy species of bacteria under harsh conditions, and develop means of sterilizing spacecraft to prevent biocontamination of Mars that could in turn interfere with future life detection missions. The four species investigated were Arthrobacter sp. KSC_Ak2i, Microbacterium schleiferi LMA_AkK1, Brevundimonas diminuta KSC_Ak3a, and Sphingomonas trueperi JSC_Ak7-3. In the study, cells of these species were mixed into Atacama Desert soil (to elucidate the shadowing effect of soil particles) and the resulting mixtures were tested both in solution and in a desiccated state under simulated Martian atmospheric and UV conditions. The UV-survival indices of Arthrobacter sp. and Microbacterium schleiferi were found to be comparable to those of Bacillus pumilus spores.

The effects of salinity and temperature shock on Kappaphycus alvarezii seaweed spores release

NASA Astrophysics Data System (ADS)

Harwinda, F. K.; Satyantini, W. H.; Masithah, E. W.

2018-04-01

One of the reproductive aspects of development step that is considered as the solution of this issue is seaweed sporulation technique through which is induced through salinity and temperature shock. This study aims to determine the effect of combination and interaction of salinity and temperature shock on the release of K. alvarezii spores in order to produce superior seeds. This research was conducted using Complete Randomized Design Factorial which consists of nine combinations of treatments and three replications. The used treatment in this study is the combination of different environmental factors such as salinity shock and temperature shock. The data were analyzed using ANOVA (Analysis of Variance) followed by Duncan Multiple Range Test. The results showed that salinity (31 ppt, 33 ppt, and 35 ppt) and temperature (30°C, 32°C, and 34°C). shock affected the osmoregulation system and the release of K. alvarezii spores. The salinity shock and temperature shock had interaction with K. alvarezii spore release on the sixth and seventh day with the best treatment at 32°C temperature and 31 ppt salinity and released 5413 cells/ml spores on the seventh day.

Reestablishment of endogonaceae on Mount St. Helens: survival of residuals

DOE Office of Scientific and Technical Information (OSTI.GOV)

Allen, M.F.; MacMahon, J.A.; Andersen, D.C.

1984-01-01

The 18 May 1980 eruption of Mount St. Helens resulted in the burial of relatively well developed soils under variable depths of sterile tephra and ash. During summer 1982, we examined a series of sites and estimated the numbers of spores of Endogonaceae that had been transported from the buried soil to the new ground surface by either abiotic or biotic vectors. There was no difference between spore counts of Endogone spp. or Glomus spp. in the buried soils of forests and clear-cuts; spores were rare in the tephra at any site. In areas featuring less than or equal tomore » 50 cm of tephra, spores were transported to the surface by gophers (in previous clear-cut areas) and by ants (in previous forest and clear-cut habitats). In the Pumice Plain, an area devoid of gophers and ants, erosion exposed spores to the surface. We found no evidence to suggest that endogonaceous fungi grow back up root systems from buried horizons. We hypothesize that small-scale perturbations (erosion, gopher and ant mounds) following the major volcanic disturbance may drive succession by exposing buried mycorrhizal and decomposer fungi. 26 references, 2 figures, 3 tables.« less

Aspergillosis in waterfowl

USGS Publications Warehouse

Herman, Carlton M.; Sladen, William J. L.

1958-01-01

Aspergillosis, a respiratory disease most commonly caused by the fungus Aspergillus fumigatus, although frequently the cause of losses in captive birds, has been little studied in wild waterfowl and other avian species. Evidence indicates this to be of importance in the wild, and studies were conducted to determine factors relating to its epizoology. Field collections from corn and other plants have yielded infective spores of Aspergillus which were inoculated into experimental chickens and ducklings and then re-isolated from characteristic lesions. A technique was developed for inoculating suspensions of known numbers of spores directly into one of the posterior thoracic airsacs. It was demonstrated that less than one million spores of A. fumigatus killed less than one-half of the experimental chickens, 10 million spores killed over 80 per cent and 50 million killed all inoculated chickens as well as ducklings. Older birds were able to survive as many as 500 million spores except when in a weakened condition. Chickens usually started dying within two days after inoculation while those that survived as long as 11 days usually fully recovered by three weeks. Pathological involvement usually was confined to lungs and airsacs. The procedures and techniques involved in these studies were illustrated on a color motion picture.

ENZYMES OF GLUCOSE AND PYRUVATE CATABOLISM IN CELLS, SPORES, AND GERMINATED SPORES OF CLOSTRIDIUM BOTULINUM1

PubMed Central

Simmons, Richard J.; Costilow, Ralph N.

1962-01-01

Simmons, R. J. (Michigan State University, East Lansing), and R. N. Costilow. Enzymes of glucose and pyruvate catabolism in cells, spores, and germinated spores of Clostridium botulinum. J. Bacteriol. 84:1274–1281. 1962.—An investigation was made of the enzymes of vegetative cells, spores, and germinated spores of Clostridium botulinum 62-A to elucidate a pathway of glucose metabolism. Manometric studies were conducted with intact cells, and various enzymes and enzyme systems were assayed in cell-free and spore-free extracts by use of spectrophotometric and colorimetric procedures. Glucose fermentation was found to be inducible; glucokinase was the controlling enzyme. All other enzymes of the Embden-Meyerhof-Parnas (EMP) pathway were found in both induced and non-induced cells, but they were in relatively low concentrations in the latter. This, plus the fact that no glucose-6-phosphate dehydrogenase was detected, led to the conclusion that glucose is catabolized primarily by the EMP system. A number of glycolytic enzymes were also found in extracts of spores and germinated spores of this organism, but the activities were extremely low as compared with activities in cell extracts. A phosphoroclastic-type reaction was readily demonstrated in both glucose-adapted and non-adapted cells, but not in spores and germinated spores. However, both acetokinase and phosphotransacetylase, as well as coenzyme A transphorase, were detected in spores and germinated-spore extracts, although at very low activity levels as compared with cell extracts. The specific activity of diaphorase in spore extracts was about one-half that of corresponding cell extracts, and the activity of reduced diphosphopyridine nucleotide (DPNH) oxidase was actually higher in the spore extracts. In addition, the DPNH oxidase in spore extracts was considerably more heat-stable than that in extracts of cells or germinated spores. PMID:13977433

Meteorological factors associated with abundance of airborne fungal spores over natural vegetation

NASA Astrophysics Data System (ADS)

Crandall, Sharifa G.; Gilbert, Gregory S.

2017-08-01

The abundance of airborne fungal spores in agricultural and urban settings increases with greater air temperature, relative humidity, or precipitation. The same meteorological factors that affect temporal patterns in spore abundance in managed environments also vary spatially across natural habitats in association with differences in vegetation structure. Here we investigated how temporal and spatial variation in aerial spore abundance is affected by abiotic (weather) and biotic (vegetation) factors as a foundation for predicting how fungi may respond to changes in weather and land-use patterns. We measured the phenology of airborne fungal spores across a mosaic of naturally occurring vegetation types at different time scales to describe (1) how spore abundance changes over time, (2) which local meteorological variables are good predictors for airborne spore density, and (3) whether spore abundance differs across vegetation types. Using an air volumetric vacuum sampler, we collected spore samples at 3-h intervals over a 120-h period in a mixed-evergreen forest and coastal prairie to measure diurnal, nocturnal, and total airborne spore abundance across vegetation types. Spore samples were also collected at weekly and monthly intervals in mixed-evergreen forest, redwood forest, and maritime chaparral vegetation types from 12 field sites across two years. We found greater airborne spore densities during the wetter winter months compared to the drier summer months. Mean total spore abundance in the mixed-evergreen forest was twice than in the coastal prairie, but there were no significant differences in total airborne spore abundance among mixed-evergreen forest, redwood forest, and maritime chaparral vegetation types. Weekly and monthly peaks in airborne spore abundance corresponded with rain events and peaks in soil moisture. Overall, temporal patterns in meteorological factors were much more important in determining airborne fungal spore abundance than the vegetation type. This suggests that overall patterns of fungal spore dynamics may be predictable across heterogeneous landscapes based on local weather patterns.

The distribution of Paenibacillus larvae spores in adult bees and honey and larval mortality, following the addition of American foulbrood diseased brood or spore-contaminated honey in honey bee (Apis mellifera) colonies.

PubMed

Lindström, Anders; Korpela, Seppo; Fries, Ingemar

2008-09-01

Within colony transmission of Paenibacillus larvae spores was studied by giving spore-contaminated honey comb or comb containing 100 larvae killed by American foulbrood to five experimental colonies respectively. We registered the impact of the two treatments on P. larvae spore loads in adult bees and honey and on larval mortality by culturing for spores in samples of adult bees and honey, respectively, and by measuring larval survival. The results demonstrate a direct effect of treatment on spore levels in adult bees and honey as well as on larval mortality. Colonies treated with dead larvae showed immediate high spore levels in adult bee samples, while the colonies treated with contaminated honey showed a comparable spore load but the effect was delayed until the bees started to utilize the honey at the end of the flight season. During the winter there was a build up of spores in the adult bees, which may increase the risk for infection in spring. The results confirm that contaminated honey can act as an environmental reservoir of P. larvae spores and suggest that less spores may be needed in honey, compared to in diseased brood, to produce clinically diseased colonies. The spore load in adult bee samples was significantly related to larval mortality but the spore load of honey samples was not.

Storage and persistence of a candidate fungal biopesticide for use against adult malaria vectors

PubMed Central

2012-01-01

Background New products aimed at augmenting or replacing chemical insecticides must have operational profiles that include both high efficacy in reducing vector numbers and/or blocking parasite transmission and be long lasting following application. Research aimed at developing fungal spores as a biopesticide for vector control have shown considerable potential yet have not been directly assessed for their viability after long-term storage or following application in the field. Methods Spores from a single production run of the entomopathogenic fungi Beauveria bassiana were dried and then stored under refrigeration at 7°C. After 585 days these spores were sub-sampled and placed at either 22°C, 26°C or 32°C still sealed in packaging (closed storage) or in open beakers and exposed to the 80% relative humidity of the incubator they were kept in. Samples were subsequently taken from these treatments over a further 165 days to assess viability. Spores from the same production run were also used to test their persistence following application to three different substrates, clay, cement and wood, using a hand held sprayer. The experiments were conducted at two different institutes with one using adult female Anopheles stephensi and the other adult female Anopheles gambiae. Mosquitoes were exposed to the treated substrates for one hour before being removed and their survival monitored for the next 14 days. Assays were performed at monthly intervals over a maximum seven months. Results Spore storage under refrigeration resulted in no loss of spore viability over more than two years. Spore viability of those samples kept under open and closed storage was highly dependent on the incubation temperature with higher temperatures decreasing viability more rapidly than cooler temperatures. Mosquito survival following exposure was dependent on substrate type. Spore persistence on the clay substrate was greatest achieving 80% population reduction for four months against An. stephensi and for at least five months against Anopheles gambiae. Cement and wood substrates had more variable mortality with the highest spore persistence being two to three months for the two substrates respectively. Conclusions Spore shelf-life under refrigeration surpassed the standard two year shelf-life expected of a mosquito control product. Removal to a variety of temperatures under either closed or open storage indicated that samples sent out from refrigeration should be deployed rapidly in control operations to avoid loss of viability. Spore persistence following application onto clay surfaces was comparable to a number of chemical insecticides in common use. Persistence on cement and wood was shorter but in one assay still comparable to some organophosphate and pyrethroid insecticides. Optimized formulations could be expected to improve spore persistence still further. PMID:23098323

Storage and persistence of a candidate fungal biopesticide for use against adult malaria vectors.

PubMed

Blanford, Simon; Jenkins, Nina E; Christian, Riann; Chan, Brian H K; Nardini, Luisa; Osae, Michael; Koekemoer, Lizette; Coetzee, Maureen; Read, Andrew F; Thomas, Matthew B

2012-10-25

New products aimed at augmenting or replacing chemical insecticides must have operational profiles that include both high efficacy in reducing vector numbers and/or blocking parasite transmission and be long lasting following application. Research aimed at developing fungal spores as a biopesticide for vector control have shown considerable potential yet have not been directly assessed for their viability after long-term storage or following application in the field. Spores from a single production run of the entomopathogenic fungi Beauveria bassiana were dried and then stored under refrigeration at 7°C. After 585 days these spores were sub-sampled and placed at either 22°C, 26°C or 32°C still sealed in packaging (closed storage) or in open beakers and exposed to the 80% relative humidity of the incubator they were kept in. Samples were subsequently taken from these treatments over a further 165 days to assess viability. Spores from the same production run were also used to test their persistence following application to three different substrates, clay, cement and wood, using a hand held sprayer. The experiments were conducted at two different institutes with one using adult female Anopheles stephensi and the other adult female Anopheles gambiae. Mosquitoes were exposed to the treated substrates for one hour before being removed and their survival monitored for the next 14 days. Assays were performed at monthly intervals over a maximum seven months. Spore storage under refrigeration resulted in no loss of spore viability over more than two years. Spore viability of those samples kept under open and closed storage was highly dependent on the incubation temperature with higher temperatures decreasing viability more rapidly than cooler temperatures. Mosquito survival following exposure was dependent on substrate type. Spore persistence on the clay substrate was greatest achieving 80% population reduction for four months against An. stephensi and for at least five months against Anopheles gambiae. Cement and wood substrates had more variable mortality with the highest spore persistence being two to three months for the two substrates respectively. Spore shelf-life under refrigeration surpassed the standard two year shelf-life expected of a mosquito control product. Removal to a variety of temperatures under either closed or open storage indicated that samples sent out from refrigeration should be deployed rapidly in control operations to avoid loss of viability. Spore persistence following application onto clay surfaces was comparable to a number of chemical insecticides in common use. Persistence on cement and wood was shorter but in one assay still comparable to some organophosphate and pyrethroid insecticides. Optimized formulations could be expected to improve spore persistence still further.

What does the geography of parthenogenesis teach us about sex?

PubMed

Tilquin, Anaïs; Kokko, Hanna

2016-10-19

Theory predicts that sexual reproduction is difficult to maintain if asexuality is an option, yet sex is very common. To understand why, it is important to pay attention to repeatably occurring conditions that favour transitions to, or persistence of, asexuality. Geographic parthenogenesis is a term that has been applied to describe a large variety of patterns where sexual and related asexual forms differ in their geographic distribution. Often asexuality is stated to occur in a habitat that is, in some sense, marginal, but the interpretation differs across studies: parthenogens might not only predominate near the margin of the sexuals' distribution, but might also extend far beyond the sexual range; they may be disproportionately found in newly colonizable areas (e.g. areas previously glaciated), or in habitats where abiotic selection pressures are relatively stronger than biotic ones (e.g. cold, dry). Here, we review the various patterns proposed in the literature, the hypotheses put forward to explain them, and the assumptions they rely on. Surprisingly, few mathematical models consider geographic parthenogenesis as their focal question, but all models for the evolution of sex could be evaluated in this framework if the (often ecological) causal factors vary predictably with geography. We also recommend broadening the taxa studied beyond the traditional favourites.This article is part of the themed issue 'Weird sex: the underappreciated diversity of sexual reproduction'. © 2016 The Author(s).

Comparative Transcriptomic and Proteomic Analyses Reveal a FluG-Mediated Signaling Pathway Relating to Asexual Sporulation of Antrodia camphorata.

PubMed

Li, Hua-Xiang; Lu, Zhen-Ming; Zhu, Qing; Gong, Jin-Song; Geng, Yan; Shi, Jin-Song; Xu, Zheng-Hong; Ma, Yan-He

2017-09-01

Medicinal mushroom Antrodia camphorata sporulate large numbers of arthroconidia in submerged fermentation, which is rarely reported in basidiomycetous fungi. Nevertheless, the molecular mechanisms underlying this asexual sporulation (conidiation) remain unclear. Here, we used comparative transcriptomic and proteomic approaches to elucidate possible signaling pathway relating to the asexual sporulation of A. camphorata. First, 104 differentially expressed proteins and 2586 differential cDNA sequences during the culture process of A. camphorata were identified by 2DE and RNA-seq, respectively. By applying bioinformatics analysis, a total of 67 genes which might play roles in the sporulation were obtained, and 18 of these genes, including fluG, sfgA, SfaD, flbA, flbB, flbC, flbD, nsdD, brlA, abaA, wetA, ganB, fadA, PkaA, veA, velB, vosA, and stuA might be involved in a potential FluG-mediated signaling pathway. Furthermore, the mRNA expression levels of the 18 genes in the proposed FluG-mediated signaling pathway were analyzed by quantitative real-time PCR. In summary, our study helps elucidate the molecular mechanisms underlying the asexual sporulation of A. camphorata, and provides also useful transcripts and proteome for further bioinformatics study of this valuable medicinal mushroom. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Aspergillus, its sexual states and the new International Code of Nomenclature.

PubMed

Pitt, John I; Taylor, John W

2014-01-01

The newly adopted International Code of Nomenclature for algae, fungi and plants (ICN) demands that dimorphic fungi, in particular those with both sexual and asexual names, now bear a single name. Although priority is no longer associated with the mode of reproduction, the ICN requires justification for choosing an asexual name over an existing sexual one. The phylogenetic approach that made dual nomenclature for fungi obsolete can be used to help choose names for large groups of fungi that are best known by asexual names. Here we apply this approach to one of the largest and most diverse asexual genera, the genus Aspergillus. We find that existing sexual names may be given to well supported clades of fungi with distinct phenotypes, which include sexual morphology as well as physiological attributes associated with xerophily, thermophily and mycotoxin production. One group of species important to food production and food safety, Aspergillus subgen. Circumdati, lacks a well supported clade; here we propose that the name Aspergillus be retained for this group. Recognizing that nomenclature has economic and social implications, particularly for old, important genera, we discuss the consequences of various scenarios to implement the new "one name for one fungus" article in the ICN, showing that our approach requires the fewest appeals to the ICN while retaining the name Aspergillus for many of the most economically and socially important species. © 2014 by The Mycological Society of America.

Robust inducible Cre recombinase activity in the human malaria parasite Plasmodium falciparum enables efficient gene deletion within a single asexual erythrocytic growth cycle.

PubMed

Collins, Christine R; Das, Sujaan; Wong, Eleanor H; Andenmatten, Nicole; Stallmach, Robert; Hackett, Fiona; Herman, Jean-Paul; Müller, Sylke; Meissner, Markus; Blackman, Michael J

2013-05-01

Asexual blood stages of the malaria parasite, which cause all the pathology associated with malaria, can readily be genetically modified by homologous recombination, enabling the functional study of parasite genes that are not essential in this part of the life cycle. However, no widely applicable method for conditional mutagenesis of essential asexual blood-stage malarial genes is available, hindering their functional analysis. We report the application of the DiCre conditional recombinase system to Plasmodium falciparum, the causative agent of the most dangerous form of malaria. We show that DiCre can be used to obtain rapid, highly regulated site-specific recombination in P. falciparum, capable of excising loxP-flanked sequences from a genomic locus with close to 100% efficiency within the time-span of a single erythrocytic growth cycle. DiCre-mediated deletion of the SERA5 3' UTR failed to reduce expression of the gene due to the existence of alternative cryptic polyadenylation sites within the modified locus. However, we successfully used the system to recycle the most widely used drug resistance marker for P. falciparum, human dihydrofolate reductase, in the process producing constitutively DiCre-expressing P. falciparum clones that have broad utility for the functional analysis of essential asexual blood-stage parasite genes. © 2013 John Wiley & Sons Ltd.

What does the geography of parthenogenesis teach us about sex?

PubMed Central

Kokko, Hanna

2016-01-01

Theory predicts that sexual reproduction is difficult to maintain if asexuality is an option, yet sex is very common. To understand why, it is important to pay attention to repeatably occurring conditions that favour transitions to, or persistence of, asexuality. Geographic parthenogenesis is a term that has been applied to describe a large variety of patterns where sexual and related asexual forms differ in their geographic distribution. Often asexuality is stated to occur in a habitat that is, in some sense, marginal, but the interpretation differs across studies: parthenogens might not only predominate near the margin of the sexuals' distribution, but might also extend far beyond the sexual range; they may be disproportionately found in newly colonizable areas (e.g. areas previously glaciated), or in habitats where abiotic selection pressures are relatively stronger than biotic ones (e.g. cold, dry). Here, we review the various patterns proposed in the literature, the hypotheses put forward to explain them, and the assumptions they rely on. Surprisingly, few mathematical models consider geographic parthenogenesis as their focal question, but all models for the evolution of sex could be evaluated in this framework if the (often ecological) causal factors vary predictably with geography. We also recommend broadening the taxa studied beyond the traditional favourites. This article is part of the themed issue ‘Weird sex: the underappreciated diversity of sexual reproduction’. PMID:27619701

Associations Between Helminth Infections, Plasmodium falciparum Parasite Carriage and Antibody Responses to Sexual and Asexual Stage Malarial Antigens.

PubMed

Ateba-Ngoa, Ulysse; Jones, Sophie; Zinsou, Jeannot Fréjus; Honkpehedji, Josiane; Adegnika, Ayola Akim; Agobe, Jean-Claude Dejon; Massinga-Loembe, Marguerite; Mordmüller, Benjamin; Bousema, Teun; Yazdanbakhsh, Maria

2016-08-03

Infections with helminths and Plasmodium spp. overlap in their geographical distribution. It has been postulated that helminth infections may influence malarial transmission by altering Plasmodium falciparum gametocytogenesis. This cross-sectional study assessed the effect of helminth infections on P. falciparum gametocyte carriage and on humoral immune responses to sexual stage antigens in Gabon. Schistosoma haematobium and filarial infections as well as P. falciparum asexual forms and gametocyte carriage were determined. The antibody responses measured were to sexual (Pfs230, Pfs48/45) and asexual P. falciparum antigens (AMA1, MSP1, and GLURP). A total of 287 subjects were included. The prevalence of microscopically detectable P. falciparum asexual parasites was higher in S. haematobium-infected subjects in comparison to their uninfected counterparts (47% versus 26%, P = 0.003), but this was not different when filarial infections were considered. Plasmodium falciparum gametocyte carriage was similar between Schistosoma- or filaria-infected and uninfected subjects. We observed a significant decrease of Pfs48/45 immunoglobulin G titer in S. haematobium-infected subjects (P = 0.037), whereas no difference was seen for Pfs230 antibody titer, nor for antibodies to AMA1, MSP1, or GLURP. Our findings suggest an effect of S. haematobium on antibody responses to some P. falciparum gametocyte antigens that may have consequences for transmission-blocking immunity. © The American Society of Tropical Medicine and Hygiene.

WhiD and WhiB, homologous proteins required for different stages of sporulation in Streptomyces coelicolor A3(2).

PubMed

Molle, V; Palframan, W J; Findlay, K C; Buttner, M J

2000-03-01

The whiD locus, which is required for the differentiation of Streptomyces coelicolor aerial hyphae into mature spore chains, was localized by map-based cloning to the overlap between cosmids 6G4 and D63 of the minimal ordered library of Redenbach et al. (M. Redenbach et al., Mol. Microbiol. 21:77-96, 1996). Subcloning and sequencing showed that whiD encodes a homologue of WhiB, a protein required for the initiation of sporulation septation in S. coelicolor. WhiD and WhiB belong to a growing family of small (76- to 112-residue) proteins of unknown biochemical function in which four cysteines are absolutely conserved; all known members of this family are found in the actinomycetes. A constructed whiD null mutant showed reduced levels of sporulation, and those spores that did form were heat sensitive, lysed extensively, and were highly irregular in size, arising at least in part from irregularity in septum placement. The whiD null mutant showed extreme variation in spore cell wall deposition; most spores had uniformly thin (20- to 30-nm) walls, but spore chains were frequently observed in which there was irregular but very pronounced (up to 170 nm) cell wall thickening at the junctions between spores. whiD null mutant spores were frequently partitioned into irregular smaller units through the deposition of additional septa, which were often laid down in several different planes, very close to the spore poles. These "minicompartments" appeared to be devoid of chromosomal DNA. Two whiD promoters, whiDp1 and whiDp2, were identified, and their activities were analyzed during development of wild-type S. coelicolor on solid medium. Both promoters were developmentally regulated; whiDp1 and whiDp2 transcripts were detected transiently, approximately at the time when sporulation septa were observed in the aerial hyphae.

Use of yeast spores for microencapsulation of enzymes.

PubMed

Shi, Libing; Li, Zijie; Tachikawa, Hiroyuki; Gao, Xiao-Dong; Nakanishi, Hideki

2014-08-01

Here, we report a novel method to produce microencapsulated enzymes using Saccharomyces cerevisiae spores. In sporulating cells, soluble secreted proteins are transported to the spore wall. Previous work has shown that the spore wall is capable of retaining soluble proteins because its outer layers work as a diffusion barrier. Accordingly, a red fluorescent protein (RFP) fusion of the α-galactosidase, Mel1, expressed in spores was observed in the spore wall even after spores were subjected to a high-salt wash in the presence of detergent. In vegetative cells, however, the cell wall cannot retain the RFP fusion. Although the spore wall prevents diffusion of proteins, it is likely that smaller molecules, such as sugars, pass through it. In fact, spores can contain much higher α-galactosidase activity to digest melibiose than vegetative cells. When present in the spore wall, the enzyme acquires resistance to environmental stresses including enzymatic digestion and high temperatures. The outer layers of the spore wall are required to retain enzymes but also decrease accessibility of the substrates. However, mutants with mild spore wall defects can retain and stabilize the enzyme while still permitting access to the substrate. In addition to Mel1, we also show that spores can retain the invertase. Interestingly the encapsulated invertase has significantly lower activity toward raffinose than toward sucrose.This suggests that substrate selectivity could be altered by the encapsulation.

Ceratopteris richardii: a productive model for revealing secrets of signaling and development

NASA Technical Reports Server (NTRS)

Chatterjee, A.; Roux, S. J.

2000-01-01

Ceratopteris richardii is an aquatic fern grown in tropical and subtropical regions of the world. It is proven to be a productive model system for studies in the genetics, biochemistry, and cell biology of basic biologic processes that occur in early gametophytic development. It provides several advantages to biologists, especially those interested in gravitational biology, polarity development, and in the genetics of sexual development. It is easy to culture, has a relatively short life cycle, and offers an array of attractive features that facilitate genetic studies. The germination and early development of large populations of genetically identical spores are easy to synchronize, and both the direction of polarity development and cell-level gravity responses can be measured and readily manipulated within the first 24 h of spore development. Although there is no reliable transformation system available yet in Ceratopteris, recent studies suggest that the technique of RNA interference can be used to block translation of specific genes in a related fern, Marsilea, and current studies will soon reveal the applicability of this approach, as well as of other transformation approaches, in Ceratopteris. A recently completed expressed sequence tag (EST) sequencing project makes available the partial sequence of more than 2000 cDNAs, representing a significant percentage of the genes being expressed during the first 24 h of spore germination, when many developmentally interesting processes are occurring. A microarray of these ESTs is being constructed, so especially for those scientists interested in basic cellular phenomena that occur early in spore germination, the availability of the ESTs and of the microarray will make Ceratopteris an even more attractive model system.

Ceratopteris richardii: a productive model for revealing secrets of signaling and development.

PubMed

Chatterjee, A; Roux, S J

2000-09-01

Ceratopteris richardii is an aquatic fern grown in tropical and subtropical regions of the world. It is proven to be a productive model system for studies in the genetics, biochemistry, and cell biology of basic biologic processes that occur in early gametophytic development. It provides several advantages to biologists, especially those interested in gravitational biology, polarity development, and in the genetics of sexual development. It is easy to culture, has a relatively short life cycle, and offers an array of attractive features that facilitate genetic studies. The germination and early development of large populations of genetically identical spores are easy to synchronize, and both the direction of polarity development and cell-level gravity responses can be measured and readily manipulated within the first 24 h of spore development. Although there is no reliable transformation system available yet in Ceratopteris, recent studies suggest that the technique of RNA interference can be used to block translation of specific genes in a related fern, Marsilea, and current studies will soon reveal the applicability of this approach, as well as of other transformation approaches, in Ceratopteris. A recently completed expressed sequence tag (EST) sequencing project makes available the partial sequence of more than 2000 cDNAs, representing a significant percentage of the genes being expressed during the first 24 h of spore germination, when many developmentally interesting processes are occurring. A microarray of these ESTs is being constructed, so especially for those scientists interested in basic cellular phenomena that occur early in spore germination, the availability of the ESTs and of the microarray will make Ceratopteris an even more attractive model system.