Yellow Strain Research Articles

Isolation and identification of yellow pigment producing marine fungi strain and characterization of the melanin pigment

Marine fungi can excrete many kinds of pigments with the potential to be applied in food and pharmaceutical industrials.Yellow pigments are widely used in the process of alcohol, beverage, and other food.In the present study, the melanin-producing fungi strains were screened from the mud collected from Haizhou Bay.The yellow pigment-producing fungi strain MP-16 was identified as Penicillium citreonigrum based on morphological characteristics and ITS r DNA sequence analysis.The pigment showed a maximum absorption peak at 330 nm.The pigment showed good resistance to heat, natural sunlight, and UV light.The pigment was stable form p H6 to p H10, and the temperature form 20 to 100 ℃. Metal ions had little effects on its stability.The results laid the foundation for the further study of the pigment.

RELATIVE IMPACTS OF DIFFERENT SEASONS ON THE QUALITIES ERISILK COCOONS AND ERI SILK YARN OF FOUR DIFFERENT MUTANTSTRAINS OF PhilosamiariciniBsd.

The relative impacts of different seasons of ericulture namely May-June,July-Aug., Sep.-Oct. and Nov.-Dec on the qualities of erisilk cocoons and erisilk yarn of four different mutant strains of PhilosamiariciniViz; Yellow plain, Yellow zebra, G.B. plain and G.B. zebra have been evaluated under the reeling parameters. Results obtained indicate that the reeling performances of G.B. plain mutant strain is best in the season May-June (summer)followed by July-August (Rainy) and are relatively better in respect of qualities of eri cocoons and eri silk yarn in comparison to two other seasons namely September-October and November-December. The said variations are probably on account of the fact that May-June followed by July-August seasons provide relatively better and conducive eco-friendly environment to mutant strain than two other seasons of eri culture in respect of cocoon weight, shell weight, shell ratio, filament length of eri silk yarn, rate of eri silk tenacity of fibre and elongation percentage of fibre. The significant variations in relation to four different seasons in the qualities of eri cocoons and eri silk yarn of yellow plain mutant strain of Philosamiaricini (the popular eri silkworm among the four mutant strains) also have been found relatively better in respect of qualities of eri cocoons and eri silk yarn in all the seasons in spite of evident variation on account of genetic variabilities and differences in the genetic make-up of mutant strains.

Phylogenic Analysis and Serotyping of Rice yellow mottle virus Strains in Uganda

Phylogenic Analysis and Serotyping of Rice yellow mottle virus Strains in Uganda

Refractoriness of Indian Aedes aegypti to Oral Infection with Yellow Fever Virus 17D Strain

Yellow fever virus (YFV) is the causative agent of yellow fever. It is one of the most important hemorrhagic arboviral infection of global public health significance. It is categorised under category ‘C’ of potential bioterrorism agent. Effect of geographical variation on vector competence in Ae. aegypti has been well documented for several viruses including YFV. In the present study, the vector competence of Ae. aegypti mosquitoes collected from Gwalior, India for YFV 17D vaccine strain was evaluated to understand the risk of its transmission. Further the risk associated with transmission of YFV 17D vaccine strain from viremic vaccinees to mosquitoes and subsequently to naive individuals was assessed. Ae. aegypti were orally infected with high titer of YFV 17D strain and the infection status was investigated at 7 and 14 day post infection (dpi) using a highly sensitive quantitative RT-PCR assay. None of the Ae. aegypti mosquito orally infected with YFV 17D strain was found to be positive for YFV. The infection rate was found to be zero per cent at both 7 dpi and 14 dpi. These results demonstrated the inability of the YFV 17D strain to cause infection or replication in the midgut of Ae. aegypti. Due to the highly attenuated replication of this strain in Ae. aegypti midgut, there is a minimal risk of its transmission. Further, it is unlikely for a mosquito that feeds on a viremic vaccine to get infected with this vaccine strain. The risk of transmission of YFV 17D strain by Indian Ae. aegypti mosquitoes is negligible. Further vector competence study using epidemic strain of YFV will aid in risk assessment analysis of YFV in India.

Mucilaginibacter antarcticus sp. nov., isolated from tundra soil.

The novel, pale yellow bacterial strain, designated S14-88T, was isolated from a tundra soil near Antarctic Peninsula, South Shetland Islands, and its taxonomic position was investigated by a genotypic and phenotypic analysis. Cells were facultatively anaerobic, Gram-stain-negative, non-motile and rod-shaped. Growth occurred at 4-28 °C (optimum at 15 °C), at pH 7.0-8.0 (optimum at 7.0) and with 0-0.6 % (w/v) NaCl (optimum, no NaCl). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain S14-88T formed a lineage within the genus Mucilaginibacter. The 16S rRNA gene sequence similarity between strain S14-88T and the type strains of related species ranged from 92.2 to 96.5 %, and the 16S rRNA gene sequence of S14-88T showed highest similarity of 96.5 % to Mucilaginibacter soyangensis HME6664T. The major cellular fatty acids of strain S14-88T were iso-C15 : 0 and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c). The major respiratory quinone was menaquinone MK-7, and the main polar lipid was phosphatidylethanolamine. The DNA G+C content of strain S14-88T was 42.3 mol%. On the basis of the evidence presented in this study, strain S14-88T is considered to represent a novel species of the genus Mucilaginibacter, for which the name Mucilaginibacter antarcticus sp. nov. is proposed. The type strain is S14-88T (=CCTCC AB 2015321T=KCTC 52232T).

Description of Novosphingobiumflavum sp. nov., isolated from soil.

A novel yellow bacterial strain, designated UCM-28T, was isolated from forest soil in Gyeonggi-Do, South Korea. The isolated strain was Gram-stain-negative, aerobic, non-spore-forming, non-motile and rod-shaped, and grew at 10-37 °C, pH 5.5-9 and with 0-1 % NaCl. It could reduce nitrate to nitrite and hydrolyse aesculin. We determined the taxonomic position of strain UCM-28T; based on the 16S rRNA gene sequence, the strain belongs to the genus Novosphingobium. The bacterium showed the highest similarity to Novosphingobiumpiscinae SLH-16T (98.9 %), Novosphingobium rhizosphaerae JM-1T (97.7 %), Novosphingobium taihuense T3-B9T (97.2 %), Novosphingobium subterraneum DSM 12447T (97.1 %), Novosphingobium aromaticivorans DSM 12444T (97.1 %) and Novosphingobium capsulatum GIFU 11526T (96.7 %). Phylogenic trees also confirmed that strain UCM-28T is most closely related to Novosphingobiumpiscinae SLH-16T and others, and is positioned within the genus Novosphingobium. The DNA relatedness of strain UCM-28T with its references was in the range of 20.9-35.2 %. The polar lipid profile revealed diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, sphingoglycolipid, phosphatidylcholine, phosphatidylmonomethylethanolamine, six unidentified polar lipids and two unknown glycolipids. The major quinone was ubiquinone Q-10, and the major polyamine was spermidine. The DNA G+C content was 63.5 mol%. The major fatty acids included (>10 %) summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) (46.3 %), summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) (24.9 %) and C14 : 0 2-OH (11.8 %). Based on the phylogenetic and phenotypic data, strain UCM-28T should be classified within the genus Novosphingobium as a representative of a novel species, named Novosphingobium flavum sp. nov. The type strain is UCM-28T (=KACC 18571T=NBRC 111647T).

Flavihumibacter sediminis sp. nov., isolated from tidal flat sediment.

A yellow, Gram-staining-positive, aerobic, rod-shaped and non-motile bacterial strain, designated CJ663T, was isolated from the tidal flat sediment in Ganghwa-do, South Korea. Strain CJ663T grew optimally on R2A at 30 °C and pH 7.0 and did not require NaCl for growth. Phylogenetic analysis based on 16S rRNA gene sequence similarity showed that strain CJ663T belonged to the genus Flavihumibacter within the family Chitinophagaceae and was most closely related to Flavihumibacter cheonanensis KACC 17467T (98.3 % similarity), followed by Flavihumibacter solisilvae KACC 17917T (97.4 %). DNA-DNA relatedness levels of strain CJ663T were 42.9 % to F. cheonanensis KACC 17467T and 48.6 % to F. solisilvae KACC 17917T. The major isoprenoid quinone was menaquinone 7 (MK-7). The predominant polar lipids were phosphatidylethanolamine, aminophospholipid and two unidentified lipids. The major cellular fatty acids of strain CJ663T were iso-C15 : 0, anteiso-C15 : 0 and iso-C17 : 0 3-OH. The G+C content of the genomic DNA was 47.7 mol%. On the basis of data from this polyphasic taxonomic study, strain CJ663T represents a novel species in the genus Flavihumibacter, for which name Flavihumibacter sediminis sp. nov. is proposed; the type strain is CJ663T (=KACC 18874T=JCM 31431T).

Genetic diversity and breeding history of Winter Mushroom (Flammulina velutipes) in China uncovered by genomic SSR markers

Flammulina velutipes is one of the most widely cultivated mushroom species in China. However, its genetic background remains poorly understood due to the limited sampling and poor molecular markers used. In this study, 124 F. velutipes strains were employed, including 110 cultivars and 14 wild strains, and 25 new SSR markers were developed based on the genome of F. velutipes. A total of 153 alleles were detected in 124 strains to investigate the improper cultivar naming, genetic diversity and breeding history of F. velutipes in China. Our fingerprinting analyses indicated that 65 strains can be differentiated from the total of 124 strains, and over 53% of the strains are labeled with improper commercial names. The genetic diversities of wild strains are higher than those of the cultivars, suggesting that wild strains may harbor a large “arsenal” gene pool in nature available for strain breeding. The white cultivars in China were originally introduced from Japan, while the yellow cultivars were directly domesticated from wild strains isolated from southeastern China or hybridized between the white cultivars and yellow strains.

Association of AMPK subunit gene polymorphisms with growth, feed intake, and feed efficiency in meat-type chickens

Investigations on regulatory genes of feed intake will provide a rational scientific basis to improve future selection indices for more efficient chickens. In the present study, we investigated the association of 13 previously reported SNPs in the chicken adenosine monophosphate activated protein kinase (AMPK) subunits PRKAB1, PRKAG2, and PRKAG3 genes with body weight (BW), body weight gain (BWG), feed intake (FI), and feed conversion ratio (FCR) in two distinct yellow meat-type strains. Six SNPs with a very low minor allele frequency were removed by genotype quality control and data filtering. The experimental population comprised 796 pedigreed males from two strains with different genetic backgrounds, 335 chickens from N202 and 461 chickens from N301. BW at 49 (BW49) and 70 days of age (BW70) and FI (from 49 to 70 days of age) were determined individually. BWG and FCR were computed based on BW and FI in the interval between 49 to 70 days. The results indicated that PRKAB1 SNPs rs14094358 and rs14094362 were significantly associated with BW70, BWG, and FI in the N202 strain, and rs14094361 and rs14094363 were significantly associated with FI and FCR in the N301 strain (P < 0.05). In addition, the PRKAG2 SNP rs14133282 showed significant association with FI in N202, and rs13535812 was significantly associated with BW70 in N202 (P < 0.05). Moreover, the PRKAG3 SNP rs13595570 was significantly associated with BW in N202 (P < 0.05), and significantly associated with FI and FCR in N301 (P < 0.05). Additionally, a two-SNP haplotype comprising rs14094361 and rs14094362 in PRKAB1 was significantly associated with BWG in N202 (P < 0.05). Meanwhile, haplotypes based on two SNPs, rs14133282, and rs13535812, showed significant effects on FI in N202 (P < 0.05). Our findings therefore provide important evidence for association of AMPK subunits polymorphisms with body weight, feed intake, and feed efficiency that may be applied in meat-type chicken breeding programs.

First International Workshop on Zika Virus Held by Oswaldo Cruz Foundation FIOCRUZ in Northeast Brazil March 2016 - A Meeting Report.

First International Workshop on Zika Virus Held by Oswaldo Cruz Foundation FIOCRUZ in Northeast Brazil March 2016 - A Meeting Report.

A humanized monoclonal antibody neutralizes yellow fever virus strain 17D-204 in vitro but does not protect a mouse model from disease

The yellow fever virus (YFV) vaccine 17D-204 is considered safe and effective, yet rare severe adverse events (SAEs), some resulting in death, have been documented following vaccination. Individuals exhibiting post-vaccinal SAEs are ideal candidates for antiviral monoclonal antibody (MAb) therapy; the time until appearance of clinical signs post-exposure is usually short and patients are quickly hospitalized. We previously developed a murine-human chimeric monoclonal antibody (cMAb), 2C9-cIgG, reactive with both virulent YFV and 17D-204, and demonstrated its ability to prevent and treat YF disease in both AG129 mouse and hamster models of infection. To counteract possible selection of 17D-204 variants that escape neutralization by treatment with a single MAb (2C9-cIgG), we developed a second cMAb, 864-cIgG, for use in combination with 2C9-cIgG in post-vaccinal therapy. MAb 864-cIgG recognizes/neutralizes only YFV 17D-204 vaccine substrain and binds to domain III (DIII) of the viral envelope protein, which is different from the YFV type-specific binding site of 2C9-cIgG in DII. Although it neutralized 17D-204 in vitro, administration of 864-cIgG had no protective capacity in the interferon receptor-deficient AG129 mouse model of 17D-204 infection. The data presented here show that although DIII-specific 864-cIgG neutralizes virus infectivity in vitro, it does not have the ability to abrogate disease in vivo. Therefore, combination of 864-cIgG with 2C9-cIgG for treatment of YF vaccination SAEs does not appear to provide an improvement on 2C9-cIgG therapy alone.

Nocardioides ginkgobilobae sp. nov., an endophytic actinobacterium isolated from the root of the living fossil Ginkgo biloba L.

A Gram-stain-positive, aerobic and yellow actinobacterial strain, designated SYP-A7303T, was isolated from the root of Ginkgo biloba L. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain SYP-A7303T belongs to the genus Nocardioides. The 16S rRNA gene sequence of strain SYP-A7303T showed highest similarity to Nocardioides marinus CL-DD14T ( = JCM 15615T) (98.3 %) and Nocardioides aquiterrae GW-9T ( = JCM 11813T) (97.1 %), and less than 96.9 % to the type strains of other species of the genus Nocardioides. Strain SYP-A7303T grew optimally at 28 °C, pH 7.0 and in the absence of NaCl. It contained ll-2,6-diaminopimelic acid in the cell-wall peptidoglycan, with mannose, ribose, rhamnose, glucose and galactose as whole-cell sugars. The polar lipids consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and an unknown lipid. The menaquinone was MK-8(H4) and the predominant cellular fatty acids were iso-C16 : 0, C18 : 1ω9c and C17 : 1ω8c. The DNA G+C content was 72 mol%. Mean DNA-DNA relatedness values between strain SYP-A7303T and the closely related strains N. marinus JCM 15615T and N. aquiterrae JCM 11813T were 62.5 ± 2.4 and 56.5 ± 3.5 %, respectively. Based on the morphological, physiological, biochemical and chemotaxonomic characteristics presented in this study, strain SYP-A7303T represents a novel species of the genus Nocardioides, for which the name Nocardioides ginkgobilobae sp. nov. is proposed. The type strain is SYP-A7303T ( = DSM 100492T = KCTC 39594T).

Luteimonas tolerans sp. nov., isolated from hexachlorocyclohexane-contaminated soil.

A Gram-stain-negative, aerobic, rod-shaped, non-spore-forming, yellow pigmented bacterial strain (UM1T) was isolated from the hexachlorocyclohexane (HCH)-contaminated dumpsite located at Ummari village in Lucknow, India. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain UM1T belongs to the genus Luteimonas with Luteimonas aestuarii B9T as the closest neighbour (97.2% 16S rRNA gene sequence similarity). The DNA G+C content of strain UM1T was 64.3 mol%. The major polar lipids were diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE) and phosphatidylglycerol (PG). Main fatty acids were iso-C15:0, iso-C11:0, iso-C11:0 3-OH, iso-C17:0 and summed feature 9 (C16:0 10-methyl and/or iso-C17:1ω9c). Ubiquinone (Q-8) was the only respiratory quinone. Spermidine was detected as the major polyamine. The DNA-DNA relatedness value of strain UM1T with respect to its closest neighbour Luteimonas aestuarii B9T was well below 70 % (∼49%). Thus, data obtained from phylogenetic analysis, DNA-DNA hybridization, and chemotaxonomical and biochemical analyses supports classification of strain UM1T as representative of a novel species of the genus Luteimonas, for which the name Luteimonas tolerans sp. nov. is proposed. The type strain is UM1T (=DSM 28473T=MCC 2572T=KCTC 42936T).

Enhancement of yellow pigment production by intraspecific protoplast fusion of Monascus spp. yellow mutant (ade−) and white mutant (prototroph)

To breed industrially useful strains of a slow-growing, yellow pigment producing strain of Monascus sp., protoplasts of Monascus purpureus yellow mutant (ade−) and rapid-growing M. purpureus white mutant (prototroph) were fused and fusants were selected on minimal medium (MM). Preliminary conventional protoplast fusion of the two strains was performed and the result showed that only white colonies were detected on MM. It was not able to differentiate the fusants from the white parental prototroph. To solve this problem, the white parental prototroph was thus pretreated with 20mM iodoacetamide (IOA) for cytoplasm inactivation and subsequently taken into protoplast fusion with slow-growing Monascus yellow mutant. Under this development technique, only the fusants, with viable cytoplasm from Monascus yellow mutant (ade−), could thus grow on MM, whereas neither IOA pretreated white parental prototroph nor yellow auxotroph (ade−) could survive. Fifty-three fusants isolated from yellow colonies obtained through this developed technique were subsequently inoculated on complete medium (MY agar). Fifteen distinguished yellow colonies from their parental yellow mutant were then selected for biochemical, morphological and fermentative properties in cassava starch and soybean flour (SS) broth. Finally, three most stable fusants (F7, F10 and F43) were then selected and compared in rice solid culture. Enhancement of yellow pigment production over the parental yellow auxotroph was found in F7 and F10, while enhanced glucoamylase activity was found in F43. The formation of fusants was further confirmed by monacolin K content, which was intermediate between the two parents (monacolin K-producing yellow auxotroph and non-monacolin K producing white prototroph).

Role of the major antigenic membrane protein in phytoplasma transmission by two insect vector species.

BackgroundPhytoplasmas are bacterial plant pathogens (class Mollicutes), transmitted by phloem feeding leafhoppers, planthoppers and psyllids in a persistent/propagative manner. Transmission of phytoplasmas is under the control of behavioral, environmental and geographical factors, but molecular interactions between membrane proteins of phytoplasma and vectors may also be involved. The aim of the work was to provide experimental evidence that in vivo interaction between phytoplasma antigenic membrane protein (Amp) and vector proteins has a role in the transmission process. In doing so, we also investigated the topology of the interaction at the gut epithelium and at the salivary glands, the two barriers encountered by the phytoplasma during vector colonization.MethodsExperiments were performed on the ‘Candidatus Phytoplasma asteris’ chrysanthemum yellows strain (CYP), and the two leafhopper vectors Macrosteles quadripunctulatus Kirschbaum and Euscelidius variegatus Kirschbaum.To specifically address the interaction of CYP Amp at the gut epithelium barrier, insects were artificially fed with media containing either the recombinant phytoplasma protein Amp, or the antibody (A416) or both, and transmission, acquisition and inoculation efficiencies were measured.An abdominal microinjection protocol was employed to specifically address the interaction of CYP Amp at the salivary gland barrier. Phytoplasma suspension was added with Amp or A416 or both, injected into healthy E. variegatus adults and then infection and inoculation efficiencies were measured.An internalization assay was developed, consisting of dissected salivary glands from healthy E. variegatus exposed to phytoplasma suspension alone or together with A416 antibody. The organs were then either observed in confocal microscopy or subjected to DNA extraction and phytoplasma quantification by qPCR, to visualize and quantify possible differences among treatments in localization/presence/number of CYP cells.ResultsArtificial feeding and abdominal microinjection protocols were developed to address the two barriers separately. The in vivo interactions between Amp of ‘Candidatus Phytoplasma asteris’ Chrysanthemum yellows strain (CYP) and vector proteins were studied by evaluating their effects on phytoplasma transmission by Euscelidius variegatus and Macrosteles quadripunctulatus leafhoppers. An internalization assay was developed, consisting of dissected salivary glands from healthy E. variegatus exposed to phytoplasma suspension alone or together with anti-Amp antibody. To visualize possible differences among treatments in localization/presence of CYP cells, the organs were observed in confocal microscopy. Pre-feeding of E. variegatus and M. quadripunctulatus on anti-Amp antibody resulted in a significant decrease of acquisition efficiencies in both species. Inoculation efficiency of microinjected E. variegatus with CYP suspension and anti-Amp antibody was significantly reduced compared to that of the control with phytoplasma suspension only. The possibility that this was due to reduced infection efficiency or antibody-mediated inhibition of phytoplasma multiplication was ruled out. These results provided the first indirect proof of the role of Amp in the transmission process.ConclusionProtocols were developed to assess the in vivo role of the phytoplasma native major antigenic membrane protein in two phases of the vector transmission process: movement through the midgut epithelium and colonization of the salivary glands. These methods will be useful also to characterize other phytoplasma-vector combinations. Results indicated for the first time that native CYP Amp is involved in vivo in specific crossing of the gut epithelium and salivary gland colonization during early phases of vector infection.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-015-0522-5) contains supplementary material, which is available to authorized users.

Development of monoclonal antibodies and serological assays specific for Barley yellow dwarf virus GAV strain

BackgroundBarley yellow dwarf virus (BYDV) is one of the most devastating plant viruses and belongs to a ubiquitous plant virus group. In China, four BYDV strains (GPV, GAV, PAV and RMV) have been identified based on their specific aphid vectors and serological properties. Among the four identified strains, the GAV is the most common BYDV strain in China. To diagnose, forecast of BYDV GAV, two reliable serological assays for BYDV GAV detection were established.MethodsWe purified virion from a confirmed BYDV GAV source and used it as the immunogen to produce monoclonal antibodies against the virus. Using the hybridoma technology, three highly specific murine monoclonal antibodies were produced and two serological assays [antigen-coated-plate enzyme-linked immunosorbent assay (ACP-ELISA) and dot enzyme-linked immunosorbent assay (dot-ELISA)] were established for the BYDV GAV detection.ResultsAll three monoclonal antibodies reacted strongly and specifically with the BYDV GAV strain in crude leaf extracts. Titers of the monoclonal antibodies in ascitic fluids were up to 10−7 by indirect-ELISA. These three monoclonal antibodies (18A1, 18A9 and 12A11) all belonged to the isotype IgG1, kappa light chain. The highest dilution points for the three antibodies during the ACP-ELISA using infected crude leaf extracts were 1:163,840, 1:81,920 and 1:81,920 (w/v, g · mL−1), respectively. Result of dot-ELISA showed a successful detection of BYDV GAV strain in 1:5,120 (w/v, g · mL−1) diluted wheat leaf crude extracts. Analysis of 22 field wheat leaf samples and 33 aphid samples from the Shaanxi Province in China, using the two newly developed assays confirmed the presence of BYDV GAV in about 80 % of the wheat samples and 18 % of the aphid samples.ConclusionsAll three monoclonal antibodies are highly sensitive and specific to the BYDV GAV. The two newly developed serological assays are simple and effective. These two assays, particularly the dot-ELISA, are useful for high throughput detection of BYDV GAV in host plants and aphid vectors.

Expressing p20 hairpin RNA of Citrus tristeza virus confers Citrus aurantium with tolerance/resistance against stem pitting and seedling yellow CTV strains

The Citrus tristeza virus (CTV) uses 3 silencing suppressor genes, p20, p23 and p25, to resist the attacks from its Citrus hosts. Inactivating these genes is therefore obviously a potential defensive option in addition to the current control strategies including aphid management and the use of mild strain cross protection. In this study, we cloned partial DNA fragments from the three genes, and used them to construct vectors for expressing hairpin RNAs (hpRNAs). To facilitate the formation of hpRNAs, the constructs were introduced in a loop structure. Following transformation of sour orange (Citrus aurantium) with these constructs, 8 p20 hpRNA (hp20) and 1 p25 hpRNA (hp25) expressing lines were obtained. The 7 hp20 transgenic lines were further characterized. Their reactions to CTV were tested following inoculation with CT14A and/or TR-L514, both of which are severe strains. Results showed that 3 lines (hp20-5, hp20-6 and hp20-8) were completely resistant to TR-L514 under greenhouse conditions for no detectable viral load was found in their leaves by PCR. However, they exhibited only partial suppression of TR-L514 under screen house conditions since the virus was detected in their leaves, though 2 months later compared to non-transgenic controls. Further tests showed that hp20-5 was tolerant also to CT14A under screen house conditions. The growth of hp20-5 was much better than others including the controls that were concurrently challenged with CT14A. These results showed that expressing p20 hpRNA was sufficient to confer sour orange with CTV resistance/tolerance.

Sphingomonas morindae sp. nov., isolated from Noni (Morinda citrifolia L.) branch.

Two yellow bacterial strains, designated NBD5(T) and NBD8, isolated from Noni (Morinda citrifolia L.) branch were investigated using a polyphasic taxonomic approach. Cells were Gram-stain-negative, aerobic, non-spore-forming, non-motile and short rod-shaped. Phylogenetic analysis based on 16S rRNA gene sequences suggested that the strains were members of a novel species of the genus Sphingomonas, the seven closest neighbours being Sphingomonas oligoaromativorans SY-6(T) (96.9% similarity), Sphingomonas polyaromaticivorans B2-7(T) (95.8%), Sphingomonas yantingensis 1007(T) (94.9%), Sphingomonas sanguinis IFO 13937(T) (94.7%), Sphingomonas ginsenosidimutans Gsoil 1429(T) (94.6%), Sphingomonas wittichii RW1(T) (94.6%) and Sphingomonas formosensis CC-Nfb-2(T) (94.5%). Strains NBD5T and NBD8 had sphingoglycolipid, phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol and phosphatidylcholine as the major polar lipids, ubiquinone 10 as the predominant respiratory quinone, and sym-homospermidine as the major polyamine. Strains NBD5(T) and NBD8 were clearly distinguished from reference type strains based on phylogenetic analysis, DNA-DNA hybridization, fatty acid composition data analysis, and comparison of a range of physiological and biochemical characteristics. It is evident from the genotypic and phenotypic data that strains NBD5(T) and NBD8 represent a novel species of the genus Sphingomonas, for which the name Sphingomonas morindae sp. nov. is proposed. The type strain is NBD5(T) ( = DSM 29151(T) = KCTC 42183(T) = CICC 10879(T)).

Detection and Identification of Phytoplasmas in Pomegranate Trees with Yellows Symptoms

AbstractSymptoms resembling those associated with phytoplasma presence were observed in pomegranate (Punica granatum L.) trees in June 2012 in the Aegean Region of Turkey (Aydın province). The trees exhibiting yellowing, reduced vigour, deformations and reddening of the leaves and die‐back symptoms were analysed to verify phytoplasma presence. Total nucleic acids were extracted from fresh leaf midribs and phloem tissue from young branches of ten symptomatic and five asymptomatic plants. Nested polymerase chain reaction assays using universal phytoplasma‐specific 16S rRNA and tuf gene primers were performed. Amplicons were digested with Tru1I, Tsp509I and HhaI restriction enzymes, according to the primer pair employed. The phytoplasma profiles were identical to each other and to aster yellows (16SrI‐B) strain when digestion was carried out on 16Sr(I)F1/R1 amplicons. However, one of the samples showed mixed profiles indicating that 16SrI‐B and 16SrXII‐A phytoplasmas were present when M1/M2 amplicons were digested, the reamplification of this sample with tuf cocktail primers allowed to verify the presence of a 16SrXII‐A profile. One pomegranate aster yellows strain AY‐PG from 16S rRNA gene and the 16SrXII‐A amplicon from tuf gene designed strain STOL‐PG were directly sequenced and deposited in GenBank under the Accession Numbers KJ818293 and KP161063, respectively. To our knowledge, this is the first report of 16SrI‐B and 16SrXII‐A phytoplasmas in pomegranate trees.

Elizabethkingia endophytica sp. nov., isolated from Zea mays and emended description of Elizabethkingia anophelisKämpfer et al. 2011.

A slightly yellow bacterial strain (JM-87(T)), isolated from the stem of healthy 10 day-old sweet corn (Zea mays), was studied for its taxonomic allocation. The isolate revealed Gram-stain-negative, rod-shaped cells. A comparison of the 16S rRNA gene sequence of the isolate showed 99.1, 97.8, and 97.4% similarity to the 16S rRNA gene sequences of the type strains of Elizabethkingia anophelis, Elizabethkingia meningoseptica and Elizabethkingia miricola, respectively. The fatty acid profile of strain JM-87(T) consisted mainly of the major fatty acids C15:0 iso, C17:0 iso 3-OH, and C15:0 iso 2-OH/C16:1ω7c/t. The quinone system of strain JM-87(T) contained, exclusively, menaquinone MK-6. The major polyamine was sym-homospermidine. The polar lipid profile consisted of the major lipid phosphatidylethanolamine plus several unidentified aminolipids and other unidentified lipids. DNA-DNA hybridization experiments with E. meningoseptica CCUG 214(T) ( = ATCC 13253(T)), E. miricola KCTC 12492(T) ( = GTC 862(T)) and E. anophelis R26(T) resulted in relatedness values of 17% (reciprocal 16%), 30% (reciprocal 19%), and 51% (reciprocal 54%), respectively. These DNA-DNA hybridization results, in addition to some differentiating biochemical properties, clearly indicate that strain JM-87(T) is a representative of a novel species, for which the name Elizabethkingia endophytica sp. nov. is proposed. The type strain is JM-87(T) ( = CIP 110885(T) = LMG 28604(T) = CCM 8570(T)).