As it is seen from Figure 1(a) and Table 3 the nod- ules of Ammodendron conollyi, formed after inoculation, were pink and large (up to 7 mm in diameter). In vari- ants of Ammodendron conollyi plant inoculation the av- erage nodule number per plant was 1-5 and their bio- mass comprised 4.5-27.5 mg. At the same time the ni- trogen fixation intensity did not influence essentially on the height of shoot part and length of roots. High nitro- gen-fixing activity was recorded in direct inoculation with own nodule bacteria, isolated from Ammodendron conollyi (AC1-1, AC2, AC12-1, AC18-1, AC21). The cross-inoculation of Ammodendron conollyi plants by nodule bacteria isolated from Astragalus villossimus, has led to visibly low nitrogen-fixing activity in comparison with results of direct inoculation of the same plants with own nodule bacteria. It should be noted that cross- inoculation of Astragalus villossimus plants by Am- modendron conollyi nodule bacteria (AC8-1, AC11, AC15) gave comparatively higher values of nitrogen fixation than their direct inoculation with own Astraga- lus villossimus nodule bacteria, during this there was an increase of nodules amount in Astragalus villossimus plants. As it is seen from Table 3, AС1-1 strain was more specific towards maternal Ammodendron conollyi plants during direct inoculation than Astragalus villos- simus plants during their cross-inoculation. It is interest- ing to note that, on the whole, nitrogen-fixing activities of Ammodendron conollyi plants inoculated with direct inoculation and cross-inoculation were in several times higher than nitrogen-fixing activities of inoculated As- tragalus villossimus plants and Astragalus unifoliolatus plants. Highly-efficient symbiosis (under term “symbio- sis efficiency” is meant the increase of biomass of in- oculated plants in comparison with biomass of non-in- oculated plants) in Ammodendron conollyi plants was observed during inoculation with such nodule bacteria as AC1-1, AC8-1, AC11, AC12-1, AC18-1, AV1, AV8-1, AV9-1, AV30, when efficiency (biomass) of shoot part increased from 143.2 to 168.6 % in comparison with control. At observation of Astragalus villossimus plants (Table 3) during direct inoculation with AV1, AV2, AV3, AV9, AV26-1 and cross-inoculation by AC11, AC15, AC21 and AU30-1 numerous nodules were formed (av- erage nodule number 10-29 and 1-2.5 mm size, their
15 Read more
Lupinus is known to form endophytic associations with both nodulating and non-nodulating bacteria. In this study, multilocus sequence analysis (MLSA) was used to analyze phylogenetic relationships among root nodule bacteria associated with Lupinus and soybean. Out of 17bacterial strains analyzed, 13 strains isolated from root nodules of Lupinus spp. were obtained from the Na- tional Rhizobium Germplasm Resource Collection, USDA. Additionally, two strains of root-nodule bacteria isolated each from native Lupinus and domes- tic soybean were examined. Sequences of the 16S rRNA gene and three housekeeping genes ( atpD , dnaK and glnII ) were used. All the reference genes were retrieved from the existing complete genome sequences only. The clus- tering of 12 of the strains was consistent among single and concatenated gene trees, but not USDA strains 3044, 3048, 3504, 3715, and 3060. According to the concatenated phylogeny, we suggest that USDA 3040, 3042, 3044, 3048, 3051, 3060, 3504, 3709 and 3715 are Bradyrhizobium , USDA 3063 and 3717 are Mesorhizobium , USDA 3043 is Burkholderia and USDA 3057a is Micro- virga . The two strains isolated from native lupines in this study are Burkhol- deria and Rhizobium , whereas the two from domestic soybean are Bradyrhi- zobium . This study emphasizes the robustness of MLSA, the diversity of bac- terial species that are capable of nodulating lupine and the substantial capabil- ity of Burkholderia spp. to colonize lupine root nodules.
23 Read more
Root nodule bacteria (RNB) or “rhizobia” are a type of plant growth promoting bacteria, typified by their ability to fix nitrogen for their plant host, fixing nearly 65% of the nitrogen currently utilized in sustainable agricultural production of legume crops and pastures. In this study, we sequenced the genomes of 110 RNB from diverse hosts and biogeographical regions, and undertook a global exploration of all available RNB genera with the aim of identifying novel genetic determinants of symbiotic association and plant growth promotion. Specifically, we performed a subtractive comparative analysis with non-RNB genomes, employed relevant transcriptomic data, and leveraged phylogenetic distribution patterns and sequence signatures based on known precepts of symbiotic- and host-microbe interactions. A total of 184 protein families were delineated, including known factors for nodulation and nitrogen fixation, and candidates with previously unexplored functions, for which a role in host-interaction, -regulation, biocontrol, and more, could be posited. These analyses expand our knowledge of the RNB purview and provide novel targets for strain improvement in the ultimate quest to enhance plant productivity and agricultural sustainability.
Root nodule bacteria are free-living soil bacteria, belonging to diverse genera within the Alphaproteobacteria and Betaproteobacteria, that have the capacity to form nitrogen-fixing symbioses with legumes. The symbiosis is specific and is governed by signaling molecules produced from both host and bacteria. Sequencing of several model RNB genomes has provided valuable insights into the genetic basis of symbiosis. However, the small number of se- quenced RNB genomes available does not currently reflect the phylogenetic diversity of RNB, or the variety of mechanisms that lead to symbiosis in different legume hosts. This prevents a broad understanding of symbiotic interactions and the factors that govern the biogeography of host-microbe symbioses.
In the first experiment, 23 strains of nodule bacteria listed in Table 1 were assessed for their capacity to nodulate three provenances of L. ambigua (CRSLAM-37, 39, 41, see Table S2). Three seedlings, representing each provenance, were placed in a 1 kg pot then inoculated with a single test strain as described by Yates et al. (2007). All treatments, plus uninoculated and nitrogen-supplied controls, were replicated four times in this split-plot design, and pots arranged in completely randomised blocks. The split-plot design allows the inclusion of more than one plant genotype in each pot which contains a single inoculant (Howieson et al., 1995; Yates et al., 2007). Sterile DI water was provided every second day and 20 ml of nutrient solution (Howieson et al., 1995) weekly through a sterilised watering tube inserted below the soil surface. After 56 days the plants were carefully removed, the roots washed free of soil and the presence of nodules recorded. Shoots were dried at 70 °C then weighed to
34 Read more
The influence of seed inoculation with a bio-preparation of nodule bacteria Rhizobium leguminosarum, fertilization of 0, 40, 80, 120 kg N/ha and molybdenum topdressing on the yield of green mass, dry ma�er, seed, average number of pods per plant, average number of seeds per pod, average nodule number per plant, nodule dry weight as well as plant and seed protein concentration were studied during the two years of investigations on two soils – Mollic Gley- sols and Eutric Cambisols. The highest values of all investigated parameters were obtained in the inoculated seed variants with molybdenum application, except the average nodule number per plant where the highest values were achieved in variants without molybdenum. The effect of nitrogen fertilization depended on the soil type, i.e. its che- mical properties. The largest number of the investigated parameters obtained the highest values as a result of fertili- zation with 40 kg N/ha on Mollic Gleysols (3.96% humus). Thus, seed yield was 4.02 t/ha, nodule dry ma�er 0.482 g per plant whereas seed protein concentration was 26.91%. The largest number of the investigated parameters on Eut- ric Cambisols (1.07% humus) obtained the highest values with fertilization of 80 kg N/ha where seed yield amounted to 3.65 t/ha, nodule dry ma�er 0.456 g per plant while seed protein concentration was 26.48%.
acid-tolerant rod that was trapped in 2001 from acidic soil collected from Karijini National Park (Australia) using Gastrolobium capitatum as a host. WSM2232 was effective in nitrogen fixation with G. capitatum but subsequently lost symbiotic competence during long-term storage. Here we describe the features of genome sequence information and its annotation. The 7,208,311 bp standard-draft genome is arranged into 72 scaffolds of 72 contigs containing 6,322 protein-coding genes and 61 RNA- only encoding genes. The loss of symbiotic capability can now be attributed to the loss of nodulation and nitrogen fixation genes from the genome. This rhizobial genome is one of 100 sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacte- ria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.
13 Read more
The indigenous populations of NAB associated with common bean ranged from 17 bacteria cells per gram of dry soil to 120000 bacteria cells per gram of dry soil. These observations provide evidence that indigenous common bean associated bacteria are widespread in Western Kenya. The high population levels of common bean associated indigenous bacteria in the field sites could be attributed to the legumes’ widespread integration in the cropping system in Kenya . Variation among the field sites in population sizes of root nodule bacteria (i.e. 17 to >120000 cells per gram of dry soil) observed in this study is a common phenomenon. This can be attributed to differences in levels of soil pH, plant nutrients, soil type, soil moisture, temperature and crop/soil management, among other factors . The results are in agreement with the findings of several researchers, Ref.  reported inhibition of NAB by high aluminium concentration and low pH. The low population of NAB and legume nodulation noted with the Kakamega soil could be attributed to its low pH (4.56), high aluminium concentration (9.425 Cmol/kg), and low organic carbon content (0.3125 Cmol/kg) that have been shown to adversely affect both survival of NAB and nodulation process in legumes .
Rhizobium mesoamericanum STM6155 (INSCD = ATYY01000000) is an aerobic, motile, Gram-negative, non-spore- forming rod that can exist as a soil saprophyte or as an effective nitrogen fixing microsymbiont of the legume Mimosa pudica L.. STM6155 was isolated in 2009 from a nodule of the trap host M. pudica grown in nickel-rich soil collected near Mont Dore, New Caledonia. R. mesoamericanum STM6155 was selected as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) genome sequencing project. Here we describe the symbiotic properties of R. mesoamericanum STM6155, together with its genome sequence information and annotation. The 6,927,906 bp high-quality draft genome is arranged into 147 scaffolds of 152 contigs containing 6855 protein-coding genes and 71 RNA-only encoding genes. Strain STM6155 forms an ANI clique (ID 2435) with the sequenced R. mesoamericanum strain STM3625, and the nodulation genes are highly conserved in these strains and the type strain of Rhizobium grahamii CCGE501 T . Within the STM6155 genome, we have identified a chr chromate efflux gene cluster of six genes arranged into two putative operons and we postulate that this cluster is important for the survival of STM6155 in ultramafic soils containing high concentrations of chromate.
11 Read more
Bradyrhizobium sp. Ai1a-2 is is an aerobic, motile, Gram-negative, non-spore-forming rod that was isolated from an effective nitrogen fixing root nodule of Andira inermis collected from Tres Piedras in Costa Rica. In this report we describe, for the first time, the genome sequence information and annotation of this legume microsymbiont. The 9,029,266 bp genome has a GC content of 62.56% with 247 contigs arranged into 246 scaffolds. The assembled genome contains 8,482 protein-coding genes and 102 RNA-only encoding genes. This rhizobial genome was sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project proposal.
Legumes engage in nitrogen-fixation symbioses with bacterial partners from at least 13 genera of Proteobac- teria [1-4]. Despite the high extent of phylogenetic diver- sity of root nodule bacteria, the very broad distribution of one particular genus (Bradyrhizobium) across host legume clades suggests that bacteria in this genus may have been the first legume symbionts . Bradyrhizo- bium interacts with the widest diversity of legume clades (at least 24 of ca. 33 nodule-forming legume tribes; ) and is associated with nodulating groups that represent early branching lineages  in all three legume subfam- ilies [8,9]. Analysis of basal Bradyrhizobium lineages that are associated with early-diverging legume groups may thus shed light on the origins of this symbiosis.
Nodule bacteria are known for their nitrogen-fixing abilities in legumes like soybean. However, in the presence of a high abundance of non-nitrogen-fixing bacteria, there could be some functional and metabolic differences. To determine the functional potential of nodule communities and pos- sible contributions to the plant or plant-bacterial interac- tions, PICRUSt was used to describe the functional profile of the nodule bacteriome, which was followed by further statistical analysis and visualization in STAMP. Based on KEGG classification, after excluding human disease and other eukaryotic-related system pathways, 28 level 2 func- tional pathways and 118 level 3 pathways were found to be statistically significant due to cultivar (Additional file 1: Table S1). Nitrogen related pathways, in particular, like amino acid metabolism, metabolism of other amino acids, and nucleotide metabolism were found to be significantly different in nodules due to cultivar. In level 3 pathways, “Phenylalanine, tyrosine and tryptophan biosynthesis”; “Val- ine, leucine and isoleucine metabolism ” ; and “ D-Arginine and D-Ornithine metabolism” in addition to “Cysteine and Methionine metabolism” were found to be significantly dif- ferent between cultivars. In addition, “ sucrose and starch metabolism” was more enriched in Fendou-78 compared to the nodules of other cultivars. No pathways were found to be significantly different between the irrigated and non-irrigated samples on any KEGG level. This may seem contradictory to the differences in diazotroph populations associated with water status; however, this may be due to a lack of sensitivity of PICRUSt to differences in function at the species level. Overall, these results indicate that based on the bacteriome, there are changes in amino acid and carbon cycling potential that would be expected to differ due to cultivars with different nodule bacteriomes.
18 Read more
subgroups were later confirmed by 16S rRNA sequencing (see Table 2). Additionally, the symbiotic phenotypes of these strains were assayed on either the original host C. cajan or the more promiscuous legume V. unguiculata (Lewin et al. 1987). A similar procedure was carried out for a few isolates, such as strain CI-34D1, which were not recognized by MALDI-TOF MS as a potential RNB. In fact, strain CI-34D1 was identified by MALDI-TOF MS as a Brevibacillus sp. and 16S rDNA sequencing showed it was related to Brevibacillus reuszeri. As shown in Table 2, only the C. cajan isolates identified by MALDI-TOF MS as members of known rhizobial genera were found to nodulate and fix nitrogen with both pigeon pea and cowpea. As expected, the non-RNB strain CI-34D1 failed to nodulate either of the tested host plants. In further tests, endosymbiotic bacteria from nodules collected from roots of clover (Trifolium spp.) or French seradella (Ornithopus sativus) growing in Australian fields were analyzed directly by MALDI-TOF MS using the protocol established by Ziegler et al. (2012), which does not require isolation or cultivation of the RNB. Nodules were found to harbor bacteria identified by MALDI-TOF MS as either R. leguminosarum (clover symbionts) or Bradyrhizobium canariense (French seradella symbionts). To verify the accuracy of MALDI-TOF MS-based results, nodule bacteria were isolated and the 16S rDNA sequences obtained for five (A- BRK13, A-G150, A-G169, A-Q1.7, and A-WB) out of the 41 isolates identified as R.
39 Read more
Bradyrhizobium elkanii USDA 76 T (INSCD = ARAG00000000), the type strain for Bradyrhizobium elkanii, is an aerobic, motile, Gram-negative, non-spore-forming rod that was isolated from an effective nitrogen-fixing root nodule of Glycine max (L. Merr) grown in the USA. Because of its significance as a microsymbiont of this economically important legume, B. elkanii USDA 76 T was selected as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria sequencing project. Here the symbiotic abilities of B. elkanii USDA 76 T are described, together with its genome sequence information and annotation. The 9,484,767 bp high-quality draft genome is arranged in 2 scaffolds of 25 contigs, containing 9060 protein-coding genes and 91 RNA-only encoding genes. The B. elkanii USDA 76 T genome contains a low GC content region with symbiotic nod and fix genes, indicating the presence of a symbiotic island integration. A comparison of five B. elkanii genomes that formed a clique revealed that 356 of the 9060 protein coding genes of USDA 76 T were unique, including 22 genes of an intact resident prophage. A conserved set of 7556 genes were also identified for this species, including genes encoding a general secretion pathway as well as type II, III, IV and VI secretion system proteins. The type III secretion system has previously been characterized as a host determinant for Rj and/or rj soybean cultivars. Here we show that the USDA 76 T genome contains genes encoding all the type III secretion system components, including a translocon complex protein NopX required for the introduction of effector proteins into host cells. While many bradyrhizobial strains are unable to nodulate the soybean cultivar Clark (rj1), USDA 76 T was able to elicit nodules on Clark (rj1), although in reduced numbers, when plants were grown in Leonard jars containing sand or vermiculite. In these conditions, we postulate that the presence of NopX allows USDA 76 T to introduce various effector molecules into this host to enable nodulation.
11 Read more
Bradyrhizobium sp. Th.b2 is an aerobic, motile, Gram-negative, non-spore-forming rod that was isolated from an effective nitrogen-fixing root nodule of Amphicarpaea bracteata collected in Johnson City, New York. Here we describe the features of Bradyrhizobium sp. Th.b2, together with high-quality permanent draft genome sequence information and annotation. The 10,118,060 high-quality draft genome is arranged in 266 scaffolds of 274 contigs, contains 9,809 protein-coding genes and 108 RNA-only encoding genes. This rhizobial genome was sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.
A total of 110 strains of nodule bacteria was isolated from plants Onobrychis transcaucasica and Onobrychis choras- sanica. Nodulation study of bacteria in both Onobrychis plant species in microvegetation experiment gave a very low nodulation on plant roots. The intensive nodulation of Onobrychis plants was recorded in vegetation experiment and for Onobrychis transcaucasica the efficiently-nodulating strains were found OT102, OT103, OT117, OT121, OT130, OT136, ОT139, ОT140, while for Onobrychis chorassanica plants – ОC106, ОC107, ОC109, ОC112, ОT103, ОT117 and ОT123 strains. Nucleotide sequencing of the 16S rRNA gene and BLAST analysis showed that nodule bacteria of Onobrychis plants were related to Rhizobium, Burkholderia, Enterobacter and Pantoea genera. It has been shown a possibility of growing up of Onobrychis plants at minimal additional moisture of sabulous soils in the Kyzyl Kum De- sert, creating artificial pastures and thereby immobilizing the desert blown sands.
12 Read more
coronaria or Sulla). WSM1592 was isolated from a nodule recovered from H. coronarium roots located in Ottava, bordering Sassari, Sardinia in 1995. WSM1592 is highly effective at fixing nitrogen with H. coronarium, and is currently the commercial Sulla inoculant strain in Australia. Here we describe the features of R. sullae strain WSM1592, together with genome sequence information and its annotation. The 7,530,820 bp high-quality permanent draft genome is arranged into 118 scaffolds of 118 contigs containing 7.453 protein-coding genes and 73 RNA-only encoding genes. This rhizobial genome is sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.
There are several types of cancer from which lung cancer is becoming the prime factor in cancer deaths due to increasing rate of smoking and air pollution in different countries. According to survey report 31% of cancer deaths for males and 26% for females are caused by the lung cancer  there has been a great effort to improve the diagnosis and treatment of the lung cancer. Medical imaging is the main tool for cancer diagnosis and lot of researchers are focused on this area. Computed Tomography (CT) is considered to be the best modality for cancer diagnosis. Time and cost are two crucial factors in diagnosis of a lung cancer. The survival from lung cancer is directly related to lung nodule growth at its detection time. The earlier detection leads to the higher the chances of successful treatment. To overcome this scenario the automated detection of cancerous lung nodule system is required. It can eliminate the health risks and complications involved in invasive operations significantly and improve the chances for successful treatment of the lung cancer patient. It will help radiologist to improve the diagnosis efficiency by calculating the quantity of nodule growth in each stage accurately.
16SrRNA gene of the isolates was highly conserved but with variable regions which make it a good marker in studying evolutionary diversity. This is in tandem with other studies which have shown that the 16SrRNA gene is efficient in defining the genera because it is conserved but have variable regions, just enough to determine genetic diversity in organisms . However, it has limitations in identifying species, due to the possible occurrence of genetic recombination and horizontal gene transfer resulting in sequence mosaicism [44,45], and perhaps this might be the reason why members of different genera clustered together on the phylogenetic tree. Another limitation of identifying bacteria based on the analysis of 16SrRNA genes is that species that are closely related may not always be differentiated because of the sequence conservation of 16SrRNA gene . To overcome these difficulties, the use of other genes including protein- coding genes with greater sequence divergence than 16SrRNA genes, are recommended as alternative genetic markers for identification of the nodule associated bacteria .
To investigate the protective effects and the possible mechanisms of garlic oil (GO) against N-nitrosodiethylamine (NDEA)-induced hepatocarcinoma in rats, Wistar rats were gavaged with GO (20 or 40 mg/kg) for 1 week, and then were gavaged with GO and NDEA (10 mg/kg) for the next 20 weeks. The changes of morphology, histology, the biochemical indices of serum, and DNA oxidative damage of liver were examined to assess the protective effects. Lipid peroxidation (LPO), antioxidant defense system, and apoptosis-related proteins were measured to investigate potential mechanisms. At the end of the study (21 weeks), GO administration significantly inhibited the increase of the nodule incidence and average nodule number per nodule-bearing liver induced by NDEA, improved hepatocellular architecture, and dramatically inhibited NDEA-induced ele- vation of serum biochemical indices (alanine aminotransferase , aspartate aminotransferase, alka- line phosphatase and gamma-glutamyl transpeptidase) and hepatic 8-hydroxy-2’-deoxyguanosine (8-OHdG) levels in a dose-dependent manner. The mechanistic studies demonstrated that GO counteracted NDEA-induced oxidative stress in rats illustrated by the restoration of glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx), glutathione-S-transferase (GST) levels, and the reduction of the malondialde- hyde (MDA) levels in liver. Furthermore, the mRNA and protein levels of Bcl-2, Bcl-xl, andβ -arrestin-2 were significantly decreased whereas those of Bax and caspase-3 were significantly increased. These data suggest that GO exhibited significant protection against NDEA-induced hepatocarcinogenesis, which might be related with the enhancement of the antioxidant activity and the induction of apoptosis.
12 Read more