Ustilago maydis populations tracked maize through domestication and cultivation in the Americas

The domestication of crops and the development of agricultural societies not only brought about major changes in human interactions with the environment but also in plants' interactions with the diseases that challenge them. We evaluated the impact of the domestication of maize from teosinte and the widespread cultivation of maize on the historical demography of Ustilago maydis, a fungal pathogen of maize. To determine the evolutionary response of the pathogen's populations, we obtained multilocus genotypes for 1088 U. maydis diploid individuals from two teosinte subspecies in Mexico and from maize in Mexico and throughout the Americas. Results identified five major U. maydis populations: two in Mexico; two in South America; and one in the United States. The two populations in Mexico diverged from the other populations at times comparable to those for the domestication of maize at 6000-10000 years before present. Maize domestication and agriculture enforced sweeping changes in U. maydis populations such that the standing variation in extant pathogen populations reflects evolution only since the time of the crop's domestication.     

In vitro interactions between Fusarium verticillioides and Ustilago maydis through real-time PCR and metabolic profiling

The goal of this research was to determine mechanisms of interaction between endophytic strains of Fusarium verticillioides (Sacc.) Nirenberg and the pathogen, Ustilago maydis (DC) (Corda). Endophytic strains of the fungus F. verticillioides are commonly found in association with maize (Zea mays) and when co-inoculated with U. maydis, often lead to decreased disease severity caused by the pathogen. Here, we developed methods (liquid chromatography-mass spectrometry) to evaluate changes in relative concentration of metabolites produced during in vitro interactions between the endophyte and pathogen. Fungi were grown on two different media, in single and in confronted cultures. We used real-time PCR (qPCR) assays to measure relative changes in fungal biomass, that occurred in confronted cultures compared to single cultures. The results showed that most secondary metabolites are constitutively produced by each species. Metabolite profiles are complex for U. maydis (twenty chromatographic peaks detected) while relatively fewer compounds were detected for F. verticillioides (six chromatographic peaks). In confronted cultures, metabolite ratio (metabolite concentration/biomass) generally increases for U. maydis metabolites while no significant changes were observed for most F. verticillioides metabolites. The results show that F. verticillioides is a strong antagonist of U. maydis as its presence leads to large reductions in U. maydis biomass. We infer that few U. maydis metabolites likely serve antibiotic functions against F. verticillioides. The methods described here are sufficiently sensitive to detect small changes in biomass and metabolite concentration associated with differing genotypes of the interacting species.     

Population genetics of the soybean aphid in North America and East Asia: test for introduction between native and introduced populations

Understanding the population genetic diversity and structure of recently introduced pest species is important for determining appropriate strategies for pest control. In recent years, the rate of introduction of new invasive insect pests between continents has continued to increase. About a decade ago, the soybean aphid, Aphis glycines Matsumura, was introduced from East Asia (EA) into North America (NA) and is now widely established in NA. To compare soybean aphid populations between the native and invasive regions, we examined 689 individuals obtained from 28 different collections in NA and EA. A total of 8 microsatellite loci were used for population genetics statistics. Genetic differentiations among NA populations were very low compared to those among EA populations. Gene diversity and mean number of alleles in NA populations averaged 0.40 and 2.70, respectively, whereas in EA they averaged 0.55 and 4.32, respectively. Structure analysis of all populations revealed two distinct structures in the invaded and in the native regions. NA populations were divided into two distinct structures consistent with their geographic distribution. Among EA populations, certain Korean populations were genetically closest to NA populations, especially those from Ohio and Delaware. An approximate Bayesian computation test also supports an introduction into NA from Korea. These suggest that Korea is most likely to be the origin of soybean aphids in North America, rather than China or Japan.     

Sipha maydis: distribution and host range of a new aphid pest of winter cereals in Argentina

Sipha maydis (Passerini) is a new aphid pest of cereals and cultivated and wild grasses in Argentina. This species was recently introduced into America, and nothing is known of its distribution or host range in South America. A better understanding of its biology is likely to facilitate control. This article records 1) the distribution and 2) the host range of S. maydis in Argentina. Over the period 2004-2006 samples were collected from 32 populations at several localities in Argentina. The number of S. maydis, accompanying aphid species, and the host from which they were collected were recorded. The distribution of S. maydis ranged from 32 degrees 52' to 42 degrees 03' S, and from 57 degrees 41' to 71 degrees 24' W, bounded by isothermals 18 and 10 degrees C and isohyets 200-400 and <1,200 mm. No S. maydis were found in the subtropical region, even in winter. In the field, the different populations showed very different host preferences. S. maydis was found mainly on cultivated barley and wheat and on wild Bromus spp. and Sorghum halepense (L.) Pers. No aphids were found on maize, Zea mays L. Most of the damage to winter cereal crops occurred at the seedling stage in early autumn and of adult plants when infestations occurred in late spring. In the 4 yr after the first record of S. maydis in Argentina, it colonized a huge area similar to that colonized by Diuraphis noxia (Mordvilko) in 10 yr. The wide range of regions, hosts and climatic conditions this species is adapted to is likely to make the control of this pest very difficult.     

Variation in interferon sensitivity and induction among strains of eastern equine encephalitis virus

Eastern equine encephalitis virus (EEEV) causes human encephalitis in North America (NA), but in South America (SA) it has rarely been associated with human disease, suggesting that SA strains are less virulent. To evaluate the hypothesis that this virulence difference is due to a greater ability of NA strains to evade innate immunity, we compared replication of NA and SA strains in Vero cells pretreated with interferon (IFN). Human IFN-alpha, -beta, and -gamma generally exhibited less effect on replication of NA than SA strains, supporting this hypothesis. In the murine model, no consistent difference in IFN induction was observed between NA and SA strains. After infection with most EEEV strains, higher viremia levels and shorter survival times were observed in mice deficient in IFN-alpha/beta receptors than in wild-type mice, suggesting that IFN-alpha/beta is important in controlling replication. In contrast, IFN-gamma receptor-deficient mice infected with NA and SA strains had similar viremia levels and mortality rates to those of wild-type mice, suggesting that IFN-gamma does not play a major role in murine protection. Mice pretreated with poly(I-C), a nonspecific IFN inducer, exhibited dose-dependent protection against fatal eastern equine encephalitis, further evidence that IFN is important in controlling disease. Overall, our in vivo results did not support the hypothesis that NA strains are more virulent in humans due to their greater ability to counteract the IFN response. However, further studies using a better model of human disease are needed to confirm the results of differential human IFN sensitivity obtained in our in vitro experiments.     

Absence of genetic divergence between western corn rootworms (Coleoptera: Chrysomelidae) resistant and susceptible to control by crop rotation

The western corn rootworm, Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae), is a major pest of corn, Zea mays L., in North America that has recently invaded Europe. A loss of ovipositional fidelity to cornfields has allowed the species to circumvent crop rotation as a means of control in part of its range in the United States. Analyses of variation at eight microsatellite loci provided no evidence for general genetic differentiation between samples of western corn rootworm collected in soybean, Glycine max (L.) Merr., fields and those collected in cornfields both inside and outside the rotation-resistance problem area. This result suggests that few or no barriers to gene flow exist between rotation-resistant and -susceptible rootworm populations. The implications of this result for the management of western corn rootworm in North America and Europe are discussed.     

Interaction between founder effect and selection during biological invasion in an aquatic plant

Long-distance colonization and rapid range expansion associated with biological invasion may have major evolutionary consequences via both stochastic processes and selection. Using large-scale population genetic surveys, we demonstrate a major shift in the relative frequency of sexually fertile diploid versus sexually sterile triploid populations associated with the invasion of North America by a clonal aquatic plant, Butomus umbellatus. Most populations across the native European range were triploid (84% of 108), whereas most introduced populations were diploid (71% of 136). We evaluated the roles of stochastic processes versus natural selection in causing this shift by surveying predominantly neutral genetic variation at 28 RAPD loci. In Europe (EU) we detected 47 distinct genotypes among 142 plants sampled from 71 populations, whereas in North America (NA) we detected only six genotypes among 138 plants from 69 populations. Of the six NA genotypes, a set of four closely related genotypes were found only in triploid populations and a pair of closely related genotypes were found only in diploid populations, and these were genetically divergent from the triploid genotypes. This result is consistent with severe founder effect. Because sex creates genotypic variation and produces offspring with greater dispersal potential than those produced clonally, we tested the hypothesis that sexual reproduction characteristic of diploids has given them a colonization advantage that accounts for their high frequency in NA. However, we found little or no evidence of sexual recruitment in introduced diploids. One very widespread heterozygous genotype occurred in 95% of 38 introduced diploid populations (i.e., 72 of 76 plants surveyed) suggesting predominant clonal reproduction. Moreover genotypic diversity was not higher within or among diploid than triploid populations in either the native or introduced range. Low genetic diversity in diploid populations was also supported by a comparison of within-population quantitative variation for plant size under a common greenhouse environment. Thus, diploids have not been favored during colonization owing to their sexual fertility. However, concurrent studies have shown that NA diploids exhibit a much higher capacity for clonal reproduction, via small vegetative bulbils, than NA triploids, which almost never produce bulbils. The same difference in clonal capacity is not a consistent feature of the native EU populations. Taken together, these results suggest that strong founder effect has set the stage for a major increase in diploid frequency due to the particular, and possibly idiosyncratic, features of the diploid and triploid lineages introduced to North America.     

Common marmosets (Callithrix jacchus) as a nonhuman primate model to assess the virulence of eastern equine encephalitis virus strains

Eastern equine encephalitis virus (EEEV) produces the most severe human arboviral disease in North America (NA) and is a potential biological weapon. However, genetically and antigenically distinct strains from South America (SA) have seldom been associated with human disease or mortality despite serological evidence of infection. Because mice and other small rodents do not respond differently to the NA versus SA viruses like humans, we tested common marmosets (Callithrix jacchus) by using intranasal infection and monitoring for weight loss, fever, anorexia, depression, and neurologic signs. The NA EEEV-infected animals either died or were euthanized on day 4 or 5 after infection due to anorexia and neurologic signs, but the SA EEEV-infected animals remained healthy and survived. The SA EEEV-infected animals developed peak viremia titers of 2.8 to 3.1 log(10) PFU/ml on day 2 or 4 after infection, but there was no detectable viremia in the NA EEEV-infected animals. In contrast, virus was detected in the brain, liver, and muscle of the NA EEEV-infected animals at the time of euthanasia or death. Similar to the brain lesions described for human EEE, the NA EEEV-infected animals developed meningoencephalitis in the cerebral cortex with some perivascular hemorrhages. The findings of this study identify the common marmoset as a useful model of human EEE for testing antiviral drugs and vaccine candidates and highlight their potential for corroborating epidemiological evidence that some, if not all, SA EEEV strains are attenuated for humans.     

Gene flow in the exotic 14-spotted ladybird beetle, Propylea quatuordecimpunctata.

Propylea quatuordecimpunctata is a Palearctic ladybird beetle that has recently become established fortuitously in northeastern North America. This predator has also been cultured by the United States Department of Agriculture from geographically diverse Old World populations and propagated for release in the U.S.A. Massive releases of P. quatuordecimpunctata have not resulted in any recognized established populations, raising a number of questions about its population genetics. We sampled beetles for electrophoretic variation in three feral populations in the U.S.A., one feral population in Europe, one cultured population from Canada, and six cultured populations from the Old World. Of the 31 putative genetic loci resolved, 26 were polymorphic (84%). Mean heterozygosity was 18.32 +/- 2.84% among all loci and was 21.84 +/- 2.89% among only polymorphic loci. Cultured and feral populations showed similar levels of heterozygosity at the 11 loci scored. New World and Old World beetles showed no significant differences in heterozygosities. Wright's fixation index FST was 0.034 +/- 0.021 among feral populations and 0.331 +/- 0.101 among cultured populations. Drift was the major force driving differentiation of cultured beetle populations. Our data do not suggest that the failure of P. quatuordecimpunctata to become established in areas where it was deliberately released was related to a paucity of genie diversity. Key words : allozymes, breeding structure, biological control, gene flow, colonization.     

Similar mating and sperm displacement patterns in two highly divergent D. simulans populations from Africa and Europe

The frequency of remating in Drosophila melanogaster is affected by both genetic and ecological factors. We studied the remating behaviour in one European (Italy) and one African (Uganda) Drosophila simulans population using six highly polymorphic microsatellite markers. Despite that the populations were genetically distinct (F(ST) = 0.18) and originated from very dissimilar ecological settings with different population densities, we inferred a very similar mating pattern. The remating parameter alpha was similar in both populations (a = 1.3-1.4). No more than two distinct paternal genotypes per family were detected in each population.     

SUBCELLULAR DISTRIBUTION OF NORADRENALINE IN GUINEA-PIG CEREBRAL CORTEX AND SPLEEN

Abstract— The distribution of noradrenaline (NA) in subcellular fractions of guinea-pig cerebral cortex and spleen was determined by differential and density gradient centrifugation. Of the primary fractions, the microsomal fraction from both tissues was enriched in NA, that of the spleen having the higher specific activity. Microsomal fractions were therefore placed on gradients and NA determined in the subfractions since these fractions appeared suitable preparations in which to search for discrete populations of vesicles. So that the non-occluded micro-particulate bound noradrenaline (MPBNA) content of gradient subfractions could be measured, [³H]NA was used to control for the diffusion and or adsorption of free NA, and occluded lactate dehydrogenase was used to estimate the amount of entrapped MPBNA and soluble NA. Non-occluded MPBNA on gradients from microsomal fractions of cerebral cortex formed a single peak mainly in subfraction F (0.6-0.8 m -sucrose). Spleen microsomal fractions, however yielded two peaks of MPBNA. one in sub-fractions D to G (0.4-1.0 m -sucrose) and the other in sub-fraction J (1.4 m -sucrosc); electron microscopy showed that the latter subfraction contained large vesicles.
Since there were unexpectedly small amounts of MPBNA in microsomal subfractions D and E of cerebral cortex, the synaptosome fraction was investigated. Following water treatment of synaptosomes. MPBNA formed a peak in subfraction E (0.4-0.6 m -sucrose) with smaller amounts in subfractions D and F (0.4 and 0.6 0.8 m -sucrose).     

Genetic variation in Bromus tectorum (Poaceae): differentiation in the eastern United States

Bromus tectorum, a devastating plant invader in western North America, had entered Pennsylvania by 1790. Although rare, or extirpated, in the east until the 1850s, it was collected with increasing frequency after 1859 from Vermont to Virginia. Using enzyme electrophoresis, we analyzed 38 populations of this grass in the eastern U.S. to determine their genetic variation and structure as well as assess their relatedness to populations in the west. Genetic variation among eastern U.S. populations is low: mean number of alleles per locus (A), percent polymorphic loci per population (%P), and expected heterozygosity (H(exp)) are 1.01, 1.05%, and 0.002, respectively. No heterozygotes were detected. The eastern populations are genetically similar: mean genetic identity for all populations was 0.990 with values among population pairs ranged from 0.913 to 1.000. Thirteen populations in eastern and western North America shared Pgm-1a and Pgm-2a, while eight populations shared Mdh-2b and Mdh-3b. Other alleles detected in western North America (Got-4c, Got-4d, and Pgi-2b) were not, however, found in eastern U.S. populations. The invasion of North America by B. tectorum occurred through multiple introductions on both coasts; results from historical and genetic evidence suggest that eastern populations stem from a minimum of two introductions. The 19th century westward spread of B. tectorum from the East appears to be plausible.     

Conservation of the b mating-type gene complex among bipolar and tetrapolar smut fungi.

In the phytopathogenic fungus Ustilago hordei, one locus with two alternate alleles, MAT-1 and MAT-2, controls mating and the establishment of the infectious dikaryon (bipolar mating). In contrast, for U. maydis, these functions are associated with two different gene complexes, called a and b (tetrapolar mating); the a complex has two alternate specificities, and the b gene complex is multiallelic. We have found homologs for the b gene complex in U. hordei and have cloned one from each mating type using sequences from one bEast allele of U. maydis as a probe. Sequence analysis revealed two divergent open reading frames in each b complex, which we called bW (bWest) and bE (bEast) in analogy with the b gene complex of U. maydis. The predicted bW and bE gene products from the two different mating types showed approximately 75% identity when homologous polypeptides were compared. All of the characterized bW and bE gene products have variable amino-terminal regions, conserved carboxy-terminal regions, and similar homeodomain motifs. Sequence comparisons with the bW1 and bE1 genes of U. maydis showed conservation in organization and structure. Transformation of the U. hordei b gene complex into a U. hordei strain of opposite mating type showed that the b genes from the two mating types are functional alleles. The U. hordei b genes, when introduced into U. maydis, rendered the haploid transformants weakly pathogenic on maize. These results indicate that structurally and functionally conserved b genes are present in U. hordei.     

Processing and secretion of a virally encoded antifungal toxin in transgenic tobacco plants: evidence for a Kex2p pathway in plants.

Ustilago maydis is a fungal pathogen of maize. Some strains of U. maydis encode secreted polypeptide toxins capable of killing other susceptible strains of U. maydis. We show here that one of these toxins, the KP6 killer toxin, is synthesized by transgenic tobacco plants containing the viral toxin cDNA under the control of a cauliflower mosaic virus promoter. The two components of the KP6 toxin, designated alpha and beta, with activity and specificity identical to those found in toxin secreted by U. maydis cells, were isolated from the intercellular fluid of the transgenic tobacco plants. The beta polypeptide from tobacco was identical in size and N-terminal sequence to the U. maydis KP6 beta polypeptide. Processing of the KP6 preprotoxin in U. maydis requires a subtilisin-like processing protease, Kex2p, which is present in both animal and fungal cells and is required for processing of (among other things) small secreted polypeptide hormones and secreted toxins. Our findings present evidence for Kex2p-like processing activity in plants. The systemic production of this viral killer toxin in crop plants may provide a new method of engineering biological control of fungal pathogens in crop plants.     

玉米田蚜虫种群的空间动态

【目的】明确我国黄淮海地区玉米蚜虫种类及其空间分布,掌握玉米蚜虫田间发生规律。【方法】采用系统调查法于2009—2010年在河北廊坊对春玉米、夏玉米上玉米蚜虫种群的发生动态进行了研究。【结果】结果表明,该地区取食为害玉米的蚜虫有5种;玉米蚜Rhopalosiphum maidis(Fitch)和禾谷缢管蚜R.padi(L.)混合发生,为玉米田蚜虫的优势种群。【结论】几种蚜虫的混合种群在玉米田间的动态分布始终呈聚集分布;二项分布k和聚集型指标m~*/x判断表明,随着玉米的生长发育,玉米蚜虫表现扩散-聚集-再扩散-再聚集的趋势。     

Pathways of introduction of the invasive aquatic plant Cabomba caroliniana

The pathway and frequency of species' introductions can affect the extent, impact, and management of biological invasions. Here, we examine the pathway of introduction of the aquatic plant Cabomba caroliniana (fanwort) into Canada and the northern United States using plastid DNA sequence (intergenic spacers atpF‐atpH, trnH‐psbA, and trnL‐trnF) and DNA content analyses. We test the hypothesis that the spread of fanwort is a result of commercial trade by comparing a Canadian population (Kasshabog Lake, ON) to native populations from southern U.S., introduced populations in northern U.S., and plants from commercial retailers. Thirteen plastid haplotypes were identified throughout North America, including one dominant haplotype, which was present in all C. caroliniana populations. Several rare haplotypes were used to infer shared colonization history. In particular, the Canadian population shared two rare alleles with a population from Massachusetts, suggesting range expansion of C. caroliniana from the northern U.S. However, the possibility of a commercial introduction cannot be excluded, as common alleles were shared between the Canadian population and both commercial and southern U.S. sources. Variation in C. caroliniana genome size was bimodal and populations were classified into “high” and “low” categories. The Canadian population had DNA contents similar to several northern U.S. populations (low DNA content). This may provide additional support for range expansion from these introduced populations rather than from commercial sources or populations in the southern U.S., which had high DNA content.     

Genetic diversity and clonal vs. sexual reproduction in Fallopia spp. (Polygonaceae)

Although fundamental to the study of invasion mechanisms, the relationship between mode of reproduction and plant invasion is not well understood. Fallopia japonica (Japanese knotweed), a highly aggressive invasive plant in both Europe and North America, serves as a model species for examining this relationship. In Britain, F. japonica var. japonica is a single female clone reproducing solely through vegetative growth or obligate hybridization with other Fallopia spp. In the U.S., however, there is more evidence for sexual reproduction. Here, simple sequence repeat (SSR) markers were developed, and three Massachusetts populations were sampled at regular intervals. The amount of sexual and clonal reproduction in each population was determined based on within-population genetic diversity. Clonal growth was apparent, but the populations together contained 26 genotypes and had evidence of sexual reproduction. One genotype that was present in all populations matched the single aggressive British clone of F. japonica var. japonica. Also, a potentially diagnostic marker for the F. sachalinensis genome provided evidence of inter- and intraspecific sexual reproduction and introgression. These differences observed in U.S. populations compared to European populations have significant implications for management of Fallopia spp. in the U.S. and underscore the importance of regional studies of invasive species.     

Genetic variability and population differentiation in captive baird's Tapirs (Tapirus bairdii)

The objectives of this study were to assess the level of genetic variability and population differentiation within captive populations of an endangered large mammal, Baird's tapir (Tapirus bairdii). We genotyped 37 captive animals from North American (NA) and Central American (CA) zoos and conservation ranches using six polymorphic microsatellite loci. Standard indices of genetic variability (allelic richness and diversity, and heterozygosity) were estimated and compared between captive populations, and between captive and wild population samples. In addition, we evaluated levels of population differentiation using Weir and Cockerham's version of Wright's F-statistics. The results indicate that the NA and CA captive populations of Baird's tapirs have retained levels of genetic variability similar to that measured in a wild population. However, inbreeding coefficients estimated from the molecular data indicate that the CA captive population is at increased risk of losing genetic variability due to inbreeding. Despite this, estimated levels of population differentiation indicate limited divergence of the CA captive population from the wild population. Careful management appears to have kept inbreeding coefficients low in the NA captive population; however, population differentiation levels indicate that the NA population has experienced increased divergence from wild populations due to a founder effect and isolation. Based on these results, we conclude that intermittent exchanges of Baird's tapirs between the NA and CA captive populations will benefit both populations by increasing genetic variability and effective population size, while reducing inbreeding and divergence from wild populations. Zoo Biol 23:521–531, 2004. © 2004 Wiley-Liss, Inc.     

The a and b loci of Ustilago maydis hybridize with DNA sequences from other smut fungi.

The smut fungi are obligately parasitic during the sexual phase of their life cycle, and the mating-type genes of these fungi play key roles in both sexual development and pathogenicity. Among species of smut fungi it is common to find a bipolar mating system in which one locus with two alternate alleles is believed to control cell fusion and establishment of the infectious cell type. Alternatively, several species have a tetrapolar mating system in which two different genetic loci, one of which has multiple alleles, control fusion and subsequent development of the infection hyphae. Cloned sequences from the a and b mating-type loci of the tetrapolar smut fungus Ustilago maydis were used as hybridization probes to DNAs from 23 different fungal strains, including smut fungi with both tetrapolar and bipolar mating systems. In general, all of the smut fungi hybridized with the mating-type genes from U. maydis, suggesting conservation of the sequences involved in mating interactions. A selection of DNAs from other ascomycete and basidiomycete fungi failed to hybridize with the U. maydis mating-type sequences. Exceptions to this finding include hybridization of DNA from the a1 idiomorph of U. maydis to DNA from one strain of U. violacea and hybridization of both a idiomorphs to DNA from Saccharomyces cerevisiae.