Strains SF2 Research Articles

Methanotrophic Communities and Cultivation of Methanotrophs from Rice Paddy Fields Fertilized with Pig-livestock Biogas Digestive Effluent and Synthetic Fertilizer in the Vietnamese Mekong Delta.

Biogas digestive effluent (BDE) has been applied to rice fields in the Vietnamese Mekong Delta (VMD). However, limited information is available on the community composition and isolation of methanotrophs in these fields. Therefore, the present study aimed (i) to clarify the responses of the methanotrophic community in paddy fields fertilized with BDE or synthetic fertilizer (SF) and (ii) to isolate methanotrophs from these fields. Methanotrophic communities were detected in rhizospheric soil at the rice ripening stage throughout 2 cropping seasons, winter-spring (dry) and summer-autumn (wet). Methanotrophs were isolated from dry-season soil samples. Although the continued application of BDE markedly reduced net methane oxidation potential and the copy number of pmoA genes, a dissimilarity ordination ana-lysis revealed no significant difference in the methanotrophic community between BDE and SF fields (P=0.167). Eleven methanotrophic genera were identified in the methanotrophic community, and Methylosinus and Methylomicrobium were the most abundant, accounting for 32.3-36.7 and 45.7-47.3%, respectively. Type-I methanotrophs (69.4-73.7%) were more abundant than type-II methanotrophs (26.3-30.6%). Six methanotrophic strains belonging to 3 genera were successfully isolated, which included type I (Methylococcus sp. strain BE1 and Methylococcus sp. strain SF3) and type II (Methylocystis sp. strain BE2, Methylosinus sp. strain SF1, Methylosinus sp. strain SF2, and Methylosinus sp. strain SF4). This is the first study to examine the methanotrophic community structure in and isolate several methanotrophic strains from BDE-fertilized fields in VMD.

Eff ect of volatile organic compounds of entomopathogenic fungi of the genus Lecanicillium and their component, the acetic acid, on the female behaviour of the western flower thrips Frankliniella occidentalis (Pergande) (Thysanoptera, Thripidae)

The e ect of VOCs of six entomopathogenic fungal strains from genus Lecanicillium on the behavioral responses of the western ower thrips females Frankliniella occidentalis on vegetative bean plants and in a Y-shaped olfactometer was studied. The signi cant repellent reactions of thrips females and a decrease in a number of o spring by 33-34% were revealed after treatment the beans by conidial suspensions of L. lecanii strains F2 and Vl 29 in the concentration of 1 × 107 spores/ml. Strains Vl 21 ( L. muscarium ) and Vit 71 ( L. attenuatum ), which showed a repellent tendency towards thrips females, also caused a signi cant decrease of o spring number. The strains isolated from aphids (ARSEF 2332 of L. dimorphum and Vl 13 of L. longisporum ) showed a tendency towards attractiveness. In the Y-shaped olfactometer the repellency of the F2 strain and the attractiveness of the ARSEF 2332 strain were con rmed. The in uence of the other studied strains on the thrips behavioral reactions was unreliable. Acetic acid, tested in the olfactometer at doses corresponding to its content in the fungal VOCs over growing mycelium, caused di erent reactions of thrips females depending on the acid concentration: attractive reactions at the dose of 0.17 ppm; neutral - at the dose of 0.34 ppm; and weak repellent - at 0.85 ppm. The obtained data indicate that acetic acid, contained in the fungal VOCs, e ect on the behavioral responses of thrips females. The repellant e ect of the fungal spores of the genus Lecanicillium on thrips females and the negative impact on o spring number increase the e ectiveness of entomopathogenic fungi.

Selection of tomato progenies F2:3 for resistance to bacterial wilt via mixed modeling

Bacterial wilt is among the major diseases of tomatoes and is currently considered the most important in Brazil. The development of resistant cultivars is the strategy that presents greater efficiency and viability, being the mixed modeling approach allows the selection of genetic materials with major accuracy. This work defines strategies for the development and selection of segregating strains F2:3 from the estimation of genetic parameters and individual mean components using mixed models. A greenhouse experiment was conducted in a randomized complete block design with four replicates, in which the resistance of 43 progenies F2:3 segregants obtained from the F2 generation from the cross between the parents was evaluated, having as parents Yoshimatsu (Resistant) and IPA-7 (Susceptible). Components of variance and genetic parameters were estimated, followed by ranking by additive genetic values. The selection of F2:3 progenies resistant to bacterial wilt was more effective when the household information was used. Mixed model procedures for the selective accuracy of 77.98%, verified efficiency in the selection of F2:3 progenies resistant to bacterial wilt evaluated at 20 days after inoculation. A total of 240 resistant and moderately resistant individual progenies were selected with severity scores between 1 and 2.49. Families presented the new averages between 1.0 and 2.87, making 23 families F2:3 that will continue the tomato breeding program.

Point mutation of V252 in neomycin C epimerase enlarges substrate-binding pocket and improves neomycin B accumulation in Streptomyces fradiae

Neomycin, an aminoglycoside antibiotic with broad-spectrum antibacterial resistance, is widely used in pharmaceutical and agricultural fields. However, separation and purification of neomycin B as an active substance from Streptomyces fradiae are complicated. Although NeoN can catalyze conversion of neomycin C to neomycin B, the underlying catalytic mechanism is still unclear. In this study, the genomic information of high-yielding mutant S. fradiae SF-2 was elucidated using whole-genome sequencing. Subsequently, the mechanism of NeoN in catalyzing conversion of neomycin C to neomycin B was resolved based on NeoN–SAM–neomycin C ternary complex. Mutant NeoNV252A showed improved NeoN activity, and the recombinant strain SF-2-NeoNV252A accumulated 16,766.6 U/mL neomycin B, with a decrease in neomycin C ratio from 16.1% to 6.28%, when compared with the parental strain SF-2. In summary, this study analyzed the catalytic mechanism of NeoN, providing significant reference for rational design of NeoN to improve neomycin B production and weaken the proportion of neomycin C.Graphical

Effects of Cd-resistant fungi on uptake and translocation of Cd by soybean seedlings

In this study, three cadmium (Cd)-resistant fungal strains, temporarily named as F1, F2 and F3 were isolated from the roots of Cd-tolerant soybean cultivars, rhizosphere and bulk soils, respectively, in contaminated sites. Cd-resistant strains F1, F2 and F3 were characterized for their effect on biomass, Cd uptake and translocation of two soybean cultivars (Liaodou36 and Liaodou33) grown in Cd-contaminated soils. The results showed that Cd concentration decreased significantly in Cd-supplemented culture solutions inoculated with strains F1, F2 and F3 compared to non-inoculated controls, while cell counts significantly increased during the incubation. The increase in shoot biomass of two soybean cultivars inoculated with strains F1, F2 and F3 ranged from 13% to 29%, 16%–27% and 15%–32%, respectively, compared to controls. Strain F2 had a higher potential to reduce the water-soluble Cd content (23% and 40%) and EDTA-extractable Cd content in the rhizosphere soil of Liaodou36 and Liaodou33 seedlings compared to strains F1 and F3. A significant decrease of Cd contents was observed in the root and shoot of Liaodou33 inoculated with strain F2 compared to non-inoculated controls, and inoculation with strain F2 significantly reduced the TF and BCF of Liaodou33 in comparison with controls. Based on ITS rRNA gene sequence analyses, the strains F1, F2 and F3 were identified as Mucor circinelloides (similarity 99.81%), Curvularia lunata (similarity 99.31%) and Clonostachys rosea (similarity 99.17%). The results of our study demonstrated that the strain F2 had a higher Cd biosorption and immobilization potential than strains F1 and F3. The strain F2 promoted the growth and reduced Cd uptake and translocation of Liaodou33 in Cd-polluted soils. It is worth noting that our results might provide an effective technical support for Cd immobilization remediation and safe soybean production by inoculating moderate Cd-accumulating soybean cultivars with strain F2 in Cd-contaminated soils.

Applied microbiology and biotechnology uncovering the biosynthetic pathway of polysaccharide-based microbial flocculant in Agrobacterium tumefaciens F2.

The low yield as bottleneck problem limits the application of microbial flocculant in water treatment. However, genetic information of microbial flocculant-producing strains can guide the regulation of microbial flocculant production, but it remains unknown. Agrobacterium tumefaciens F2 produced polysaccharide-based microbial flocculants in the fermentation medium but none in Luria Bertani medium; hence, the transcriptome was used to analyze the potentially associated genes with the production of microbial flocculants. Glucose, mannose, rhamnose, and galactose are the main sugar monomers, and genes (manA, glmM, manC, rfb genes, exo genes, etc.) with changed expression levels related to sugar monomers metabolism potentially participated in the biosynthesis of polysaccharide-based microbial flocculants. exoC, exoP, and manC were confirmed to participate in the biosynthesis via constructing the mutants F2-dexoC, F2-dexoP, and F2-dmanC. An exoF2 gene cluster was annotated due to the high percentage of matches between the genome sequences of strains F2 and C58, and exo genes in their genome sequences showed the similarity of 86~92%. The hypothetical pathway for the biosynthesis of polysaccharide-based microbial flocculants in strain F2 was proposed, laying the basis for the production yield regulation. KEY POINTS: • An exoF2 gene cluster in the polysaccharide biosynthesis was annotated. • exoC, exoP, and manC genes participated in the polysaccharide biosynthesis. • A hypothetical biosynthesis pathway of polysaccharide in flocculant was proposed. Graphical abstract.

First report about the screening, characterization, and fosmid library construction of Xanthomonas oryzae pv. oryzae strain with resistance to Fubianezuofeng

Bacterial blight (BB), which is caused by Xanthomonas oryzae pv. oryzae (Xoo), is a common bacterial disease that seriously harms rice production in major rice-growing areas worldwide. Fubianezuofeng (FBEZF), a sulfone bactericide that contains an oxadiazole moiety, exerts good control effect on BB. In this study, FBEZF-resistant strains of Xoo were screened for the first time in the laboratory to evaluate the risk of Xoo developing resistance to FBEZF. Three strains with moderate resistance to FBEZF, were obtained and named as F1, F2, and F3, which have resistance factors (RF) of 14.69, 15.72, and 11.12, respectively. FBEZF lacked positive cross-resistance to bismerthiazol, thiodiazole copper, zhongshengmycin and phenazino-1-carboxylic acid. The growth rates of the resistant strains F1 and F2 were similar to those of the wild-type strain in nutrient broth medium, but differed in nutrient agar medium. The extracellular polysaccharide production and pathogenicity of F1, F2, and F3 were reduced relative to those of the wild-type strain. A fosmid library containing 2304 transformants was constructed based on the genome of F2, and transformants 2193 and 2202 exhibited FBEZF resistance. The results are helpful for further study on the molecular mechanism of resistance to FBEZF in Xoo.

Uji Multilokasi Sepuluh Galur Padi Untuk Menghasilkan Varietas Unggul Baru

Reduced productive rice fields to non-paddy fields, most possible extension of the nationalrice cultivation area to the suboptimal lands reaching ± 91.9 million ha. Technologypackages that can be applied include the use of new improved rice varieties through theassembling of varieties that have high yield potential, resistant to suboptimal soil stress. Theobjective of the research is to produce new superior rice varieties that are available invarious environments. The study used a split-plot design, as the main plot is an environmentconsisting of rice paddies and gogo, as a subplot is the genotype of rice. The treatment ineach environment is prepared using the Randomized Complete Block Design (RCBD). Thetreatment consisted of 10 new rice strains and four varieties as a comparison. Variablesobserved (1) plant height; (2) the maximum number of shoots; (3) number of productiveshoots; (4) long panicle; (5) the number of grains per panicle; (6) the amount of graincontent of each panicle; (7) the number of empty grains per panicle; (8) weight of 1000grains of grain; (9) grain yield of each clump; (10) grain yield per hectare. Data wereanalyzed by variance if there was a difference between mean, median treatment, followed byLsd test at 5% level. The adaptability and yield stability of each strain was determined basedon the value of the coefficient of diversity (KK) (Francis and Kenneberg, 1978) in Syukur etal., (2012). The results showed that seven new rice strains had adaptations both planted asupland rice and lowland rice, namely strains B3, B4, F2, F3, H1, H4, and L2.

Complete Sequences of Multiple-Drug Resistant IncHI2 ST3 Plasmids in Escherichia coli of Porcine Origin in Australia

IncHI2 ST3 plasmids are known carriers of multiple antimicrobial resistance genes. Complete plasmid sequences from multiple drug resistant Escherichia coli circulating in Australian swine is however limited. Here we sequenced two related IncHI2 ST3 plasmids, pSDE-SvHI2 and pSDC-F2_12BHI2, from phylogenetically unrelated multiple-drug resistant Escherichia coli strains SvETEC (CC23:O157:H19) and F2_12B (ST93:O7:H4) from geographically disparate pig production operations in New South Wales, Australia. Unicycler was used to co-assemble short read (Illumina) and long read (PacBio SMRT) nucleotide sequence data. The plasmids encoded three drug-resistance loci, two of which carried class 1 integrons. One integron, hosting drfA12-orfF-aadA2, was within a hybrid Tn1721/21, with the second residing within a copper/silver resistance transposon, comprising part of an atypical sul3-associated structure. The third resistance locus was flanked by IS15DI and encoded neomycin resistance (neoR). An oqx-encoding transposon (quinolone resistance), similar in structure to Tn6010, was identified only in pSDC-F2_12BHI2. Both plasmids showed high sequence identity to plasmid pSTM6-275, recently described in Salmonella enterica serotype 1,4,[5],12:i:- that has risen to prominence and become endemic in Australia. IncHI2 ST3 plasmids circulating in commensal and pathogenic E. coli from Australian swine belong to a lineage of plasmids often in association with sul3 and host multiple complex antibiotic and metal resistance structures, formed in part by IS26.

Biodiesel production using lipase immobilised functionalized magnetic nanocatalyst from oleaginous fungal lipid

From the tannery effluent and oil contaminated soil different microbe were isolated and screened for high lipid content. Among all the isolates fungal strain SF2 showed high lipid content was selected and identified as Aspergillus niger. It was found that the fungi produced about 44.02% of lipid. The fatty acid composition was identified by gas chromatography. The physicochemical property of the extracted lipid was analysed. Magnetic nanoparticle was prepared and the lipase enzyme was immobilised on to the magnetic nanoparticle. Characterization of magnetic nanoparticle and enzyme immobilised magnetic nanoparticle was carried out. Esterase activity of both free enzyme and immobilised enzyme was analysed at different parameters in which immobilised enzyme showed better stability. The immobilised enzyme can be stored for 60 days without any loss in activity. Biodiesel was produced using the lipase immobilised on the magnetic nanoparticle. The optimum parameters for the fatty acid methyl ester production were investigated in terms of both physical and chemical parameters and the biodiesel yield under optimised reaction condition was 84%. The reusable stability of the catalyst was performed and it was found that the immobilised biocatalyst can be used for 5 cycles. The physio-chemical properties of the biodiesel was analysed and it was found to be within the limit specified by American society of testing materials standards. This work was firstly studied on the conversion of Aspergillus lipid into biodiesel by using lipase immobilised magnetic nanoparticle.

Investigating the Role of Endophytic Fungi in Gentiana scabra bge. by Cross-Growth Period Inoculation.

Gentiana scabra Bge. (gentian) is a Chinese medicinal plant. Endophytic fungi from the roots of gentian were isolated and cross-growth period inoculation was performed to study the roles of three Trichoderma spp. strains (F1, F2, and F9) in their original host plant. In treatments inoculated with F1, F2, and F9, gentiopicroside content increased 33.6, 23.7 and 13% than that in the control. Strains F1, F2, and F9 could also improve polysaccharide content by more than 6.6, 18.7 and 30% compared to the control. The incidence of spot blight in gentian inoculated with F1, F2, and F9 decreased by 31.2, 26.7 and 8.5%. Inconsistent changes in the activity of the three enzymes (superoxide dismutase, catalase and peroxidase) were observed when the plants were attacked by pathogens or inoculated with fungi. High enzymatic activity did not reflect mild disease. Cross-growth period inoculation, which takes into account the original living environment (gentian plant as "substrate" and different microorganisms as symbionts) of endophytic fungi, provides a new idea for studying effects of endophytes on their original hosts. This is the first research about the role of endophytic fungi in Gentiana scabra bge. in vivo.

The Migration of Multipartial Nuclear and Its Regulation by DCTN1 Gene in the Mycelium Cells of Volvariella volvacea

Eight Volvariella volvacea strains (F1, F2, F3, F4, F5, F6, F7, and F8) were grown in a favorable environment, and their vegetative hyphae were stained using eosin-crystal violet. Microscopic observations showed few or nonexistent nuclei in the cells at the tip of the hyphae, whereas more nuclei were observed in the sub-apical cells. Migrating nuclei were found in most strains, except F2 and F5. These nuclei had no definite shapes as they passed through intercellular channels. Migrating nuclei had the tendency to move towards the hyphal tips. Centrosomes and cytoskeleton were occasionally observed during this process. Likewise, DCTN1 gene transcription in the hyphae was analyzed by semi-quantitative RT-PCR. The transcriptional levels of strains F2 and F5 were significantly lower than those of the other strains, which implied that the nuclear migration was regulated by DCTN1 gene. This paper provides fundamental information on the mechanisms of nuclear migration and strain degeneration in V. volvacea.

Production of a novel bioflocculant and its flocculation performance in aluminum removal

ABSTRACTA novel bioflocculant CBF with high flocculating activity, produced by mixed culture of Rhizobium radiobacter F2 and Bacillus sphaericus F6 from soil, was investigated with regard to its production and flocculation performance in Al(III) removal. The most preferred carbon source, nitrogen source and C/N ratio (w/w) for strains F2 and F6 to produce CBF were glucose, urea and 20, respectively. The optimal inoculum size for CBF production was 10 % (v/v). The optimal initial pH, culture temperature and shaking speed were 7–8, 30°C and 140 r/min for 24 h, respectively, under which the flocculating activity of the bioflocculant reached 98.52 %. According to literature review, flocculant dosage, coagulant aid dosage, pH, hydraulic condition of coagulation and sedimentation time are considered as influencing parameters for CBF flocculation performance in Al(III) removal by L16(45) orthogonal design. The optimal conditions for Al(III) removal obtained through analysis and verification experiments were as follows: CBF, 28 mg/L; coagulant aid, 1.5 mL/L; initial pH, 8.0; and hydraulic conditions of coagulation: stir speed, 160 r/min; stir time, 40 s; and sedimentation time, 30 min. Under the optimal conditions, the removal efficiency of Al(III) was 92.95 %. Overall, these findings indicate that bioflocculant CBF offers an effective alternative method of decreasing Al(III) during drinking water treatment.

Association between a naturally arising polymorphism within a functional region of HIV-1 Nef and disease progression in chronic HIV-1 infection.

HIV-1 Nef mediates downregulation of HLA class I (HLA-I) through a number of highly conserved sequence motifs. We investigated the in vivo implication(s) of naturally arising polymorphisms in functional motifs in HIV-1 Nef that are associated with HLA-I downregulation, including the acidic cluster, polyproline, di-arginine and Met-20 regions. Plasma samples from treatment-naive, chronically HIV-1 infected subjects were collected after obtaining informed consent, and viral RNA was extracted and amplified by nested RT-PCR. The resultant nef amplicons were sequenced directly, and subtype-B sequences with an intact open reading frame (n = 406) were included in our analyses. There was over-representation of isoleucine at position 20 (Ile-20) in our dataset when compared to sequences in the Los Alamos sequence database (17.7 vs. 6.9 %, p = 0.0309). The presence of having Ile-20 in Nef was found to be associated with higher median plasma viral load (p = 0.013), independent of associated codons or viral lineage effects, whereas no clinical association was found with polymorphisms in the other functional motifs. Moreover, introduction of a Met-20-to-Ile mutation in a laboratory strain SF2 Nef resulted in a modest, albeit not statistically significant, increase in HLA class I downregulation activity (p = 0.06). Taken together, we have identified a naturally arising polymorphism, Ile-20, within HIV-1 subtype B Nef that is associated with poorer disease outcome.

Draft Genome Sequence of Asaia sp. Strain SF2.1, an Important Member of the Microbiome of Anopheles Mosquitoes

Asaia spp. are abundant members of the microbiota of Anopheles mosquitoes, the principle vectors of malaria. Here, we report the draft genome sequence of Asaia sp. strain SF2.1. This strain is under development as a platform to deliver antimalarial peptides and proteins to adult female Anopheles mosquitoes.

Dynamic range of Nef functions in chronic HIV-1 infection

HIV-1 Nef is required for efficient viral replication and pathogenesis. However, the extent to which Nef's functions are maintained in natural sequences during chronic infection, and their clinical relevance, remains incompletely characterized. Relative to a control Nef from HIV-1 strain SF2, HLA class I and CD4 down-regulation activities of 46 plasma RNA Nef sequences derived from unique chronic infected individuals were generally high and displayed narrow dynamic ranges, whereas Nef-mediated virion infectivity, PBMC replication and CD74 up-regulation exhibited broader dynamic ranges. 80% of patient-derived Nefs were active for at least three functions examined. Functional co-dependencies were identified, including positive correlations between CD4 down-regulation and virion infectivity, replication, and CD74 up-regulation, and between CD74 up-regulation and PBMC replication. Nef-mediated virion infectivity inversely correlated with patient CD4± T-cell count. Strong functional co-dependencies and the polyfunctional nature of patient-derived Nef sequences suggest a phenotypic requirement to maintain multiple Nef functions during chronic infection.

Studies on phenotypic traits of different shell color strains F<SUP>2</SUP> of Manila clam(<I>Ruditapes philippinarum</I>)

The study on phenotypic traits of different shell color strains F1 and F2(C:the control strains;Tr;two-band red strain;Tw:two-band white strain;Ab:agate-black strain;Or:ocean-red strain;Pw:pearl-white strain;Z:zebra strain;W:wave strain.) of Manila clam was conducted in September 2007.The results showed that the shell length of Pw F1 was the greatet,and significantly greater than others except Tr(P0.05),the shell length of Z was the smallest,and significantly smaller than others(P0.05),there were no obvious differences in shell length among C,Or,W,between Tr and Tw,among Tw,Ab,Or,among Ab,Or,W(P0.05).Both fresh weight and fecundity of Pw F1 were the greatet,that of Tr were the second greatet and significantly greater than others(P0.05),those of Z were the smallest,and significantly smaller than others(P0.05),there were no remarkable differences between C and W,among Tw,Ab,Or in fresh weight and between C and Z,among Tr,Ab,Or,W in fecundity(P0.05).No significant differences in egg-diameter,the fertilized rates and size of D larvae were observed among all strains(P0.05),the hatching rate of C was significantly lower than those of other strains except Tw(P0.05).By end of pelagic period(9 d),the shell lengths of W and Z were significantly greater than those of other strains(P0.05),there were no remarkable differences in shell length among Tw,Ab,Or,Pw,Z,W(P0.05) ;the sequences of survival rates were following:Ab,ZOr,WTw,PwTrC,there were significant differences among them(P0.05).No remarkable difference in both metamorphic time and metamorphic size among all strains was observed(P0.05).There were such a sequence in metamorphic rates as follows:AbZTwOrPwWTrC,there were significant differences among them(P0.05).By the end of indoor rearing period(240 d),the shell length of Ab were significantly greater than those of other strains except Or,Tr(P0.05),the shell length of Z were significantly smaller than those of other strains except C,W(P0.05),there were no remarkable differences in shell length between C and Pw,W,Tr,Tw,Or,and between Tw and Pw;the survival rate of Z was remarkably higher than those of other strains except Ab(P0.05),the survival rate of C was remarkably lower than those of other strains except Tr,Tw(P0.05).On day 360,both shell length and fresh weight were the greatet and remarkably greater than those of other strains(P0.05),the shell length of Ab and Z were significantly smaller than those of other strains(P0.05),the survival rate of Ab and Z were significantly higher than those of other strains except Or,Pw(P0.05),the survival rate of Or was significantly higher than that of C(P0.05),there was no remarkable difference in survival rate among other strains(P0.05).The order of relative yields was as follows:Z(678.63%±56.80%)Ab(554.88%±69.42%)Or(527.23%±76.21%)W(475.97%±90.25%)Pw(405.90%±55.19%)Tw(224.89%±47.85%)Tr(178.50%±34.50%)C(100.00%±22.10%).

Endophytic Bacteria Improve Seedling Growth of Sunflower Under Water Stress, Produce Salicylic Acid, and Inhibit Growth of Pathogenic Fungi

Endophytic bacterial strains SF2 (99.9% homology with Achromobacter xylosoxidans), and SF3 and SF4 (99.9% homology with Bacillus pumilus) isolated from sunflower grown under irrigation or drought were selected on the basis of plant growth-promoting bacteria (PGPB) characteristics. Aims of the study were to examine effects of inoculation with SF2, SF3, and SF4 on sunflower cultivated under water stress, to evaluate salicylic acid (SA) production by these strains in control medium or at Ψa = -2.03 MPa, and to analyze effects of exogenously applied SA, jasmonic acid (JA), bacterial pellets, and bacterial supernatants on growth of pathogenic fungi Alternaria sp., Sclerotinia sp., and Verticillum sp. Growth response to bacterial inoculation was studied in two inbred lines (water stress-sensitive B59 and water stress-tolerant B71) and commercial hybrid Paraiso 24. Under both water stress and normal conditions, plant growth following inoculation was more strongly enhanced for Paraiso 24 and B71 than for B59. All three strains produced SA in control medium; levels for SF3 and SF4 were higher than for SF2. SA production was dramatically higher at Ψa = -2.03 MPa. Exogenously applied SA or JA caused a significant reduction of growth for Sclerotinia and a lesser reduction for Alternaria and Verticillum. Fungal growth was more strongly inhibited by bacterial pellets than by bacterial supernatants. Our findings indicate that these endophytic bacteria enhance growth of sunflower seedlings under water stress, produce SA, and inhibit growth of pathogenic fungi. These characteristics are useful for formulation of inoculants to improve growth and yield of sunflower crops.

Endophytic bacteria in sunflower (Helianthus annuus L.): isolation, characterization, and production of jasmonates and abscisic acid in culture medium

This study was designed to isolate and characterize endophytic bacteria from sunflower (Helianthus annuus) grown under irrigation and water stress (drought) conditions, to analyze growth of isolated bacteria under drought condition, and to evaluate the ability of bacteria isolated from plants cultivated under drought to produce jasmonates (JAs) and abscisic acid (ABA). Bacteria were isolated from soil samples collected when sunflower plants were at the end of the vegetative stage. A total of 29 endophytic strains were isolated from plants grown under irrigation or drought condition. Eight strains (termed SF1 through SF8) were selected based on nitrogen-fixing ability. All eight strains showed positive catalase and oxidase activities; five strains (SF2, SF3, SF4, SF5, SF7) solubilized phosphates; none of the strains produced siderophores. Strains SF2, SF3, SF4, and SF5, the ones with the highest phosphate solubilization ability, strongly inhibited growth of the pathogenic fungi Verticillum orense and Sclerotinia sclerotiorum but had less inhibitory effect on Alternaria sp. Among the eight strains, SF2 showed 99.9% sequence homology with Achromobacter xiloxidans or Alcaligenes sp., while the other seven showed 99.9% homology with Bacillus pumilus. Strains SF2, SF3, and SF4 grown in control medium produced jasmonic acid (JA), 12-oxo-phytodienoic acid (OPDA), and ABA. These three strains did not differ in amount of JA or OPDA produced. ABA content was higher than that of JA, and production of both ABA and JA increased under drought condition. The characteristics of these isolated bacterial strains have technological implications for inoculant formulation and improved growth of sunflower crops.

Antigen Expression Kinetics and Immune Responses of Mice Immunized with Noninfectious Simian-Human Immunodeficiency Virus DNA

In a previous report we demonstrated that three injections of an rt-deleted noninfectious genome of the simian-human immunodeficiency virus SHIV(KU2) induced protection against AIDS in macaques (D. K. Singh, Z. Liu, D. Sheffer, G. A. Mackay, M. Smith, S. Dhillon, R. Hegde, F. Jia, I. Adany, and O. Narayan, J. Virol 79:3419-3428, 2005). To make this DNA safer, we deleted two more genes, the integrase gene and vif, along with the 3' long terminal repeat. We also replaced the gag, pro, and nef genes (SIVmac239 origin) with those of human immunodeficiency virus (HIV) type 1 strain SF2. The resultant construct, designated delta4SHIV(KU2) DNA, was used in this study to evaluate gene expression and immunogenicity in BALB/c mice. DNA-transfected human embryonic kidney epithelial cells (HEK 293) produced all of the major viral proteins and released p24 in the supernatant for 12 days. Inoculation of the vaccine DNA into the gastrocnemius muscles resulted in intense mononuclear cell infiltration at the inoculated sites and the production of viral p24 in myocytes, in infiltrating mononuclear cells, and in cells in the spleen and draining lymph nodes between 3 and 10 days postinoculation. Expression of p24 in the muscle cells peaked at day 7 and became undetectable after day 12. The same 12-day period of expression of p24 was observed in mice that were given a second injection 4 weeks after the first. Evaluation of immune responses in BALB/c mice revealed that the DNA induced enzyme-linked immunospot and antigen-specific proliferative cell-mediated immunity responses. The responses were stronger in mice that were coinjected with a second plasmid expressing granulocyte-macrophage colony-stimulating factor. Since new waves of viral antigen production could be induced with each boosting injection of the vaccine DNA, this DNA could be a safe and efficient agent to induce long-term protection against HIV.