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We have isolated a Penicillium species belonging to section Lanata-Divaricata from crop production soils in south central Colorado. The fungus was rigorously identified using morphological methods, metabolite analysis, scanning electron microscopy, and multi-locus sequencing phylogeny and was found to be Penicillium raperi G. Sm. In culture, the fungus bears smooth-walled conidia born terminally in chains on divaricate conidiophores with complex branching patterns. Optimal growth in culture was achieved on CYA at 28 °C, but the fungus also grew on MEA and CYA at temperatures ranging from 25 to 37 °C. A concatenated ITS, β-tubulin, and calmodulin tree showed that the isolated fungus was nested within the P. raperi clade. Various classes of metabolites (e.g., benzenoids, lipids, nucleosides, and organic acids) were produced in culture on different media. The fungus colonized Colletotrichum coccodes and inhibited growth of the agriculturally significant pathogen, P. expansum, in culture but was not pathogenic on wound-inoculated potato or apple fruit. We propose that Penicillium raperi can be utilized to make small molecules of industrial interest and optimized as a biological control agent for potato and apple postharvest diseases. [ABSTRACT FROM AUTHOR]

Rice grain quality is a multifaceted quantitative trait that impacts crop value and is influenced by multiple genetic and environmental factors. Chemical, physical, and visual analyses are the standard methods for measuring grain quality. In this study, we evaluated high-throughput hyperspectral imaging for quantification of rice grain quality and classification of grain samples by genetic sub-population and production environment. Whole grain rice samples from the USDA mini-core collection grown in multiple locations were evaluated using hyperspectral imaging and compared with results from standard phenotyping. Loci associated with hyperspectral values were mapped in the mini-core with 3.2 million SNPs in a genome-wide association study (GWAS). Our results show that visible and near infra-red (Vis/NIR) spectroscopy can classify rice according to sub-population and production environment based on differences in physicochemical grain properties. The 702–900 nm range of the NIR spectrum was associated with the chalky grain trait. GWAS revealed that grain chalk and hyperspectral variation share genomic regions containing several plausible candidate genes for grain chalkiness. Hyperspectral quantification of grain chalk was validated using a segregating bi-parental mapping population. These results indicate that Vis/NIR can be used for non-destructive high throughput phenotyping of grain chalk and potentially other grain quality properties. [ABSTRACT FROM AUTHOR]

The Mythimna unipuncta nucleopolyhedrovirus isolate KY310 (MyunNPV-KY310) is an alphabaculovirus isolated from a true armyworm (Mythimna unipuncta) population in Kentucky, USA. Occlusion bodies of this virus were examined by electron microscopy and the genome sequence was determined by 454 pyrosequencing. MyunNPV-KY310 occlusion bodies consisted of irregular polyhedra measuring 0.8-1.8 µm in diameter and containing multiple virions, with one to six nucleocapsids per virion. The genome sequence was determined to be 156,647 bp with a nucleotide distribution of 43.9% G+C. 152 ORFs and six homologous repeat (hr) regions were annotated for the sequence, including the 38 core genes of family Baculoviridae and an additional group of 26 conserved alphabaculovirus genes. BLAST queries and phylogenetic inference confirmed that MyunNPV-KY310 is most closely related to the alphabaculovirus Leucania separata nucleopolyhedrovirus isolate AH1, which infects Mythimna separata. In contrast, MyunNPV-KY310 did not exhibit a close relationship with Mythimna unipuncta nucleopolyhedrovirus isolate #7, an alphabaculovirus from the same host species. MyunNPV-KY310 lacks the gp64 envelope glycoprotein, which is a characteristic of group II alphabaculoviruses. However, this virus and five other alphabaculoviruses lacking gp64 are placed outside the group I and group II clades in core gene phylogenies, further demonstrating that viruses of genus Alphabaculovirus do not occur in two monophyletic clades. Potential instances of MyunNPV-KY310 ORFs arising by horizontal transfer were detected. Although there are now genome sequences of four different baculoviruses from M. unipuncta, comparison of their genome sequences provides little insight into the genetic basis for their host specificity. [ABSTRACT FROM AUTHOR]

Lilies are grown in the field for bulb production where they are susceptible to infection by Pratylenchus penetrans, the root lesion nematode. These migratory nematodes feed on roots and often physically disrupt the root tissues when many nematodes enter and move throughout the root to feed. The goal of this study was to find a root-specific or root-preferred promoter because of our interest in genetic engineering of lilies for nematode resistance. Several promoters, the Agrobacterium rolD and mas2, CaMV 35S, and rice RPC1 were examined in transgenic Lilium longiflorum, Easter lily, plants that contained the bar-uidA fusion gene under each promoter. Histochemical staining showed that the rolD, mas2, and CaMV 35S promoters directed gus gene expression in the cortex and stele of transformed roots. Gus expression was limited to the stele of roots with the rice RPC1 promoter making it unsuitable for engineering nematode resistance. The mas2 promoter was found to be expressed primarily in roots rather than shoots whereas the CaMV 35S promoter expressed well in both roots and shoots. Levels of gus specific activity were relatively high for both the mas2 and CaMV 35S promoter in roots but low with the rolD promoter. Because the mas2 promoter had relatively high levels of gus specific activity in roots and not shoots, and expression was throughout cortex and stele tissues of the roots, mas2 appears to be a promising promoter for engineering resistance to root lesion nematodes in Lilium longiflorum. [ABSTRACT FROM AUTHOR]

A baculovirus isolate from a USDA Forest Service collection was characterized by electron microscopy and analysis of its genome sequence. The isolate, formerly referred to as Pseudoletia (Mythimna) sp. nucleopolyhedrovirus #7 (MyspNPV#7), was determined by barcoding PCR to derive from the host species Mythimna unipuncta (true armyworm) and was renamed Mythimna unipuncta nucleopolyhedrovirus #7 (MyunNPV#7). The occlusion bodies (OBs) and virions exhibited a size and morphology typical for OBs produced by the species of genus Alphabaculovirus, with occlusion-derived virions consisting of 2-5 nucleocapsids within a single envelope. The MyunNPV#7 genome was determined to be 148,482 bp with a 48.58% G+C nucleotide distribution. A total of 159 ORFs of 150 bp or larger were annotated in the genome sequence, including the 38 core genes of family Baculoviridae. The genome contained six homologous repeat regions (hrs) consisting of multiple copies of a 34-bp imperfect palindrome. Phylogenetic inference from concatenated baculovirus core gene amino acid sequence alignments placed MyunNPV#7 with group II alphabaculoviruses isolated from other armyworm and cutworm host species of lepidopteran family Noctuidae. MyunNPV#7 could be distinguished from other viruses in this group on the basis of differences in gene content and order. Pairwise nucleotide distances suggested that MyunNPV#7 represents a distinct species in Alphabaculovirus. The MyunNPV#7 genome was found to contain two copies of the late expression factor-7 (lef-7) gene, a feature not reported for any other baculovirus genome to date. Both copies of lef-7 encoded an F-box domain, which is required for the function of LEF-7 in baculovirus DNA replication. [ABSTRACT FROM AUTHOR]

Tuckerella japonica Ehara (Acari: Tetranychoidea: Tuckerellidae) was found on stems of seedling plants of Camellia sinensis (L.) O. Kuntze (Theaceae) in the continental USA. This mite is able to pierce exposed green periderm tissue with its paired stylets on 1- to 3- or 4-year-old stems where the outer bark had split longitudinally. The mite was not found on branches older than 3 or 4 years, where splitting diminished and, eventually, a uniform covering of bark was formed. The mouthparts of T. japonica were examined under scanning electron microscopy and their external morphology was compared with known Tetranychoidea. There were usually one or two feeding holes in an area where the female subsequently deposited one or more eggs. Females were observed defending the areas where their eggs and/or young occurred. This behavior along with potentially limited access to exposed periderm or availability of shoots in the rows of plants may restrict higher populations of T. japonica from becoming established. The potential of T. japonica as an economic pest remains unknown at this time. [ABSTRACT FROM AUTHOR]

In 2014, we performed a nationwide survey in Korean radish fields to investigate the distribution and variability of Turnip mosaic virus (TuMV). Brassica rapa ssp. pekinensis sap-inoculated with three isolates of TuMV from infected radish tissue showed different symptom severities, whereas symptoms in Raphanus sativus were similar for each isolate. The helper component-protease (HC-Pro) genes of each isolate were sequenced, and phylogenetic analysis showed that the three Korean isolates were clustered into the basal-BR group. The HC-Pro proteins of these isolates were tested for their RNA silencing suppressor (VSR) activity and subcellular localization in Nicotiana benthamiana. A VSR assay by co-agroinfiltration of HC-Pro with soluble-modified GFP (smGFP) showed that HC-Pro of isolate R007 and R041 showed stronger VSR activity than R065. The HC-Pros showed 98.25 % amino acid identity, and weak VSR isolate (R065) has a single variant residue in the C-terminal domain associated with protease activity and self-interaction compared to isolates with strong VSR activity. Formation of large subcellular aggregates of GFP:HC-Pro fusion proteins in N. benthamiana was only observed for HC-Pro from isolates with strong VSR activity, suggesting that R065 'weak' HC-Pro may have diminished self-association; substitution of the variant C-terminal residue largely reversed the HC-Pro aggregation and silencing suppressor characteristics. The lack of correlation between VSR efficiency and induction of systemic necrosis (SN) suggests that differences in viral accumulation due to HC-Pro are not responsible for SN. [ABSTRACT FROM AUTHOR]

The main pathogen of Gladiolus plants is Fusarium oxysporum, a soilborne fungus that infects roots and corms resulting in death of the plant. Purified D4E1, a synthetic antimicrobial peptide, was previously reported (De Lucca et al. ) to inhibit F. oxysporum spores from forming mycelial colonies in vitro at a concentration of 3 μM making it a candidate gene for genetic engineering of Gladiolus for resistance to F. oxysporum. Gladiolus cv. Peter Pears plants were transformed by particle bombardment with plasmid DNA containing a 90 bp D4E1 gene that was under the control of the duplicated CaMV 35S promoter. Five of the 14 independently transformed plant lines were evaluated for resistance to F. oxysporum. Transgenic plants were tested in vitro for resistance to F. oxysporum, and several lines appeared to be more resistant than the control plants that lacked D4E1. Cell extracts of transgenic Gladiolus lines 6(1) and 7(1) inhibited germinated spores of F. oxysporum f. sp. gladioli from forming mycelial colonies by 34 and 38 %, respectively, in vitro. F. oxysporum f. sp . gladioli was transformed with the ECFP (cyan) gene allowing us to follow the growth of F. oxysporum during infection of D4E1-transformed and non-transformed roots. Fluorescence observations using confocal laser scanning microscopy showed that 3-10 days after infection, F. oxysporum covered the surface of the root and formed pseudo-appressoria, but hyphae were never observed to penetrate cells of the root. Ten days after infection with F. oxysporum, non-transformed roots had completely disintegrated whereas transgenic roots of line 7(1) were just beginning to lose their cellular integrity. Cell extracts from the five transgenic lines showed either an inhibition of F. oxysporum mycelial colony formation or less fungal hyphae were observed to infect their roots as compared to non-transformed Gladiolus plants. [ABSTRACT FROM AUTHOR]

Previous research on the locomotion of the Nematalycidae has only been undertaken on Gordialycus, which is by far the most elongated genus of the family. Gordialycus is dependent on an unusual form of peristalsis to move around. It was not known whether the genera of Nematalycidae with shorter bodies also used this mode of locomotion. Our videographic recordings of Osperalycus did not reveal peristalsis. Instead, this mite appears to move around the mineral regolith via the expansion and constriction of the metapodosomal and genital region, allowing greater versatility in the way that the annular regions contract and extend. This type of locomotion would enable relatively short bodied nematalycids to anchor themselves into secure positions before extending their anterior regions through tight spaces. Low-temperature scanning electron micrographs show that the short bodied genera have integumental features that appear to be associated with this mode of locomotion. Peristalsis is almost certainly a more derived form of locomotion that is an adaptation to the unusually long body form of Gordialycus. [ABSTRACT FROM AUTHOR]

A proof of concept study was conducted to determine if transparent double-sided adhesive tape could be used to recover and detect [by polymerase chain reaction (PCR) and immunofluorescence microscopy (IFA)] Cryptosporidium parvum oocysts on fresh produce and on a food preparation surface. Oocysts were applied on the surface of ten apples, ten peaches, eight cucumbers, and eight tomatoes within circles drawn with a permanent marker. Approximately 18 h later, skin excised from three uncontaminated and three contaminated circles from each piece of produce was subjected to PCR. Pieces of transparent double-sided adhesive tape were lightly pressed onto the surface of three other contaminated circles and examined by PCR. Other pieces of adhesive tape were pressed against the surfaces of three other circles and examined by IFA. At concentrations of 100 and 50 oocysts per circle, every produce item examined by PCR of contaminated excised skin was found positive, and every item examined by adhesive tape subjected to PCR and IFA was found positive, except one. At ten oocysts per circle, every produce item was found positive by PCR of contaminated excised skin, and all apples, cucumbers, and tomatoes were found positive by adhesive tape subjected to IFA. Detection of low numbers of oocysts on peaches by IFA examination of adhesive tape was problematic because trichomes that cover peaches and impart the fuzzy surface partially restrict the tape from reaching some areas where oocysts adhere. Tape combined with IFA was successful in recovering and identifying oocysts from six areas of laminate countertop where the oocysts had been applied and allowed to dry for 30-60 min. These are the first findings to demonstrate that adhesive tape can be used to recover and identify a protozoan parasite from fresh produce and from a laminate food preparation surface. [ABSTRACT FROM AUTHOR]

Three different injection molded bioplastic (BP) pots, BP 29, BP 32, and BP 45 pots, containing processed feather fibers, were evaluated. Growth and flowering of Lilium hybrid 'Triumphator' and Begonia 'Bonfire' were evaluated in new and reused BP pots. Net photosynthesis was measured and macro- and micro-elements were analyzed in 'Triumphator' lily. Ultrastructure of epidermis cells of 'Triumphator' lily were analyzed using a low temperaturescanning electron microscope (LT-SEM). Surface changes, the weight, and macro- and micro-elements of the BP pots were also analyzed. When lilies were grown in BP 32 pots, root and shoot growth were inhibited, flowering was delayed and the net photosynthesis was negative, grana development, and starch granules were not observed as compared to those in BP 45 pots. Changes in the surface of BP 32 pots after forcing lilies (used once) showed a distinct space between feather fibers and other constituents which could be due to the chemical changes resulting from absorption of water. When BP pots were analyzed for pH and macro- and micro-elements, the pH and the concentration of ammonium nitrogen, phosphorus, potassium, calcium, magnesium, sulfur, and iron were significantly higher in the new BP 32 than those of BP 29 and BP 45 pots. Based on the growth and development of lily and begonia and net photosynthesis and grana development of lily, and foliar analysis of begonia grown in three different BP pots, it is concluded that BP 45 is recommended to grow lily and begonia when proper levels of nutrition are maintained during culture. Poor root growth and development of 'Triumphator' lily grown in new BP 32 pots could be associated with leaching of toxic substances once the pots absorb water during the culture. [ABSTRACT FROM AUTHOR]

Immunolocalization of β- and δ-giardin in Giardia duodenalis trophozoites revealed that both giardins are strictly associated with the ventral disk (VD). Optical sectioning of the immunolabeled VD, together with quantitative colocalization of δ- and β-giardin immunoreactivity, demonstrated that δ-giardin is primarily localized to the ventral side, and β-giardin is localized to the dorsal side of the VD. [ABSTRACT FROM AUTHOR]

Fusarium rot caused by Fusarium oxysporum f.sp. gladioli ( Fog) is one of the most serious diseases of Gladiolus, both in the field and in bulbs in storage. In order to study the mechanisms of pathogenesis of this fungus, we have transformed Fog with Agrobacterium tumefaciens binary vectors containing the hygromycin B phosphotransferase ( hph) gene and fluorescence reporter genes EGFP (green), EYFP (yellow) or ECFP (cyan) using the AGL-1 strain of A. tumefaciens. Hygromycin B (100 μg/ml) resistant colonies were observed only when acetosyringone was added to the co-cultivation medium. Transformed colonies are more clearly visible when co-cultivated on cellophane membrane than on Hybond -N membrane. Transformed lines were stably maintained through four serial passages on medium containing hygromycin B, and they expressed green, yellow or cyano fluorescence. PCR with hph-specific primers and Southern blotting with an hph-specific probe were positive for Hyg lines but not for the untransformed isolate. The cyano fluorescence of the ECFP-transformed isolate was clearly distinguishable from the green autofluorescence of Gladiolus roots, signifying the potential of these lines for further histopathological investigations. Transformed lines will be useful for identifying pathogenicity related genes, screening transgenic resistance, and in studies of host-pathogen interactions. [ABSTRACT FROM AUTHOR]

This study was conducted to evaluate the growth and flowering of Begonia boliviensis A. DC. 'Bonfire' when grown in the growing medium mixed with pellets from biobased plastic resins containing chicken feather fibers, and analyze macro- and micro-elements in soil and leaf tissues during different developmental stages of begonia. Growth, flowering, and foliar analysis of plants grown in 10 cm pots containing 95 grams medium mixed with 6 grams each of pellet 29 (P 29), 32 (P 32), 37 (P 37), or 40 (P 40), 3 grams of Na (P NaS), or 9 grams of 45 (P 45) were similar to the control plants. Extra nitrogen was available from P 32 and P 37 based on the growing medium and foliar analysis of plants grown in P 32 (P 32 plant) and P 37 plant during the first 59 days after planting (DAP). Slow growth, low flowering percentage, or even the death of B. boliviensis 'Bonefire' grown in P NaS medium is attributed to high pH and high sulfur concentration. Based on the pH, soluble salts level, and the concentrations of macro- and micro-elements in the medium and leaves, P 45 pellet is the suitable source, if nitrogen is supplied constantly to the optimum level. Pellets prepared following mixing > 30 feather fibers without using glycerol should be further tested. [ABSTRACT FROM AUTHOR]

This study was conducted to (a) characterize bio-plastic pellets containing feather fibers by low temperature-scanning electron microscopy (LT-SEM) and X-Ray diffraction analysis, (b) evaluate growth and flowering of Begonia boliviensis A. DC. 'Bonfire' when grown in a medium amended with pellets, and (c) analyze macro- and micro-elements in the medium and plant tissues of begonia. Based on physical and chemical analyses of six types of pellets mixed in the medium, pellets 32 (P 32), 37 (P 37), 40 (P 40), and P NaS (P NaS), P NaS containing sodium sulfite and > 30% glycerol were not suitable and not recommended for future evaluations. Pellets containing glycerol at 10% were not suitable for the use as an ingredient for commercial growing media and P 45 is considered suitable pellets to prepare pots. [ABSTRACT FROM AUTHOR]

Polyploidy in blackberries and ploidy differences between Rubus species are obstacles to the efficient introduction of valuable germplasm, both intra- and interspecific, into blackberry breeding programs. Expansion of the germplasm base would be facilitated by reducing the ploidy level of blackberry cultivars to the diploid level. In this report selection of twin seed, interspecific hybridization, and pollen irradiation were compared as methods in the recovery of dihaploids from tetraploid blackberry cultivars. One dihaploid was obtained through selection of twin seed and several were obtained following interspecific hybridization. The infrequency of twinning and difficulty in detecting twin seed in Rubus reduced the efficiency of this method. The efficiency with which dihaploids could be obtained following interspecific hybridization varied with the pollen parent. Reduced seed set and seed quality following pollinations with respectively R. parvifolius and R. hirsutus could be used to advantage in the recovery of dihaploids from blackberries. Ploidy reduction in several tetraploid blackberry cultivars was obtained following pollinations with 100 and 150 kR gamma irradiated pollen. Most of the seedlings obtained at the 50 kR dosage were aneuploid. Pollen irradiation at 150 kR was the most efficient method of obtaining dihaploids from tetraploid blackberries. Twenty percent of the seedlings obtained following this treatment were dihaploid. [ABSTRACT FROM AUTHOR]