Molecular tracing of <Emphasis Type="Italic">Bradyrhizobium </Emphasis>strains helps to correctly interpret <Emphasis Type="Italic">Acacia mangium </Emphasis>response to inoculation in a reforestation experiment in Madagascar

A field inoculation experiment using two Bradyrhizobium sp. strains was set up in Madagascar to test the growth response of Acacia mangium and to follow up the survival of inoculant strains using molecular tools. Three months after inoculation, one of the inoculant strains, AUST13c, exhibited a marked growth-promoting effect with a shoot height about 40% higher than that of the uninoculated control plants or TEL8-inoculated plants. The positive effect on tree growth initially observed with AUST13c was no more significant 6 months after transfer to the field and disappeared completely at 13 and 19 months. Analyses of nodule bacterial rRNA by PCR/RFLP displayed an early contamination of the different inoculation treatments by AUST13c 3 months after inoculation, spreading to almost all nodules of the trial 6 months later. This work clearly demonstrated that the progressive reduction of the positive effect of AUST13c inoculation on tree growth after field transplanting was not due to a progressive disappearance of this introduced strain but, on the contrary, was related to the widening spreading of AUST13c in all the plots. This was attributed to a higher competitiveness and effectiveness of AUST13c over the local strains and TEL8.     

Development and use of microsatellite markers for genetic diversity analysis of cañahua (<Emphasis Type="Italic">Chenopodium pallidicaule</Emphasis> Aellen)

Cañahua (Chenopodium pallidicaule Aellen) is a poorly studied, annual subsistence crop of the high Andes of South America. Its nutritional value (high in protein and mineral content) and ability to thrive in harsh climates make it an important regional food crop throughout the Andean region. The objectives of this study were to develop genetic markers and to quantify genetic diversity within cañahua. A set of 43 wild and cultivated cañahua genotypes and two related species (Chenopodium quinoa Willd. and Chenopodium petiolare Kunth) were evaluated for polymorphism using 192 microsatellite markers derived from random genomic cañahua sequences produced by 454 pyrosequencing of cañahua genomic DNA. Another 424 microsatellite markers from C. quinoa were also evaluated for cross-species amplification and polymorphism in cañahua. A total of 34 polymorphic microsatellite marker loci were identified which detected a total of 154 alleles with an average of 4.5 alleles per marker locus and an average heterozygosity value of 0.49. A cluster analysis, based on Nei genetic distance, clearly separated from wild cañahua genotypes from the cultivated genotypes. Within the cultivated genotypes, subclades were partitioned by AMOVA analysis into six model-based clusters, including a subclade consisting sole of erect morphotypes. The isolation by distance test displayed no significant correlation between geographic collection origin and genotypic data, suggesting that cañahua populations have moved extensively, presumably via ancient food exchange strategies among native peoples of the Andean region. The molecular markers reported here are a significant resource for ongoing efforts to characterize the extensive Bolivian and Peruvian cañahua germplasm banks, including the development of core germplasm collections needed to support emerging breeding programs.     

Field performance of three Bradyrhizobium japonicum strains with two soybean (Glycine max. L.) cultivars

Summary A field experiment was condutced in a clay loam soil to study the performance of three Bradyrhizobium japonicum strains; USDA 110, USDA 138 and TAL 379, in relation to their N₂-fixing potential and competitiveness on two soybean cultivars (Clark and Calland). Inoculation of soybean cultivars with these strains, either singly or in combination, induced significant increases in plant dry weight, N₂ fixation and seed yields. However, no significant differences were found between the rhizobial strains and/or their mixtures in N₂ fixation and increased seed yield for both cultivars. The two soybean cultivars gave similar responses to inoculation. No significant differences in seed yield were observed between Clark and Calland cultivars. The interaction between inoculant strain and soybean cultivar was not significant. The competition between strains for nodulation was assessed. Strain USDA 110 was the most competitive, followed by USDA 138. Strain TAL 379 was always less competitive on both cultivars. The incidence of double-strain occupancy of nodules varied from 8% to 40%.     

Intrinsic antibiotic resistance and competition in fast- and slow-growing soybean rhizobia on a hybrid of Asian and US cultivars

Summary Six fast-growing soybean rhizobia (Rhizobium fredii) and thirteen slow-growing soybean rhizobia (Bradyrhizobium japonicum) were examined for resistance to 10 antibiotics. Axenic studies were carried out to determine the competitiveness of dual-strain inocula consisting of fast- and slow-growing rhizobia isolated from subtropical-tropical soils for nodule occupancy on a hybrid of Asian and US soybean cultivars. Nodule occupancy was determined by intrinsic resistance to erythromycin and neomycin. The results showed wide variability in resistance to 10 antibiotics for fast- and slow-growing rhizobia. The intrinsic antibiotic resistance of fast- and slow-growing rhizobia was extremely high against nalidixic acid (400 g ml^–1) and penicillin (200 g ml^–1). The competitive ability of inoculant strains for nodule occupancy varied for different combination sets and with the plant growing media. Our results show that fast-growing rhizobia nodulate a hybrid of Asian and US soybean cultivars. Fast-growing soybean rhizobia did not completely exclude nodulation by the slow-growing strains, which formed 0–79% nodules, depending on the strain used in the inoculum.     

Genetic diversity and potential for promotion of plant growth detected in nodule endophytic bacteria of soybean grown in Heilongjiang province of China

A total of 98 non-symbiotic endophytic bacterial strains isolated from soybean root nodules were classified into eight rDNA types in ARDRA analysis and 21 BOX types in BOX-PCR. The phylogenetic analysis of 16S rDNA identified these strains as Pantoea, Serratia, Acinetobacter, Bacillus, Agrobacterium, and Burkholderia. Limited genetic diversity was revealed among these bacteria since most of the strains (85.7%) were found in three very similar rDNA types corresponding to Pantoea agglomerans, and many strains shared the same BOX-PCR patterns. The inoculation of nodule endophytes had no significant effects on the growth and nodulation of soybean, but most of the strains produced indoleacetic acid (IAA), could solubilize mineral phosphate, and could fix nitrogen, implying that they are a valuable pool for discovering plant growth promoting bacteria. Our results demonstrated that the nodule endophytes were common in soybean and their diversity was affected by the plant's character and the soil conditions. The 99% similarities found in the nifH genes of Bradyrhizobium japonicum and of the endophytic Bacillus strains strongly indicated that horizontal transfer of symbiotic genes happened between the symbiotic bacteria and the endophytes.     

The role of inoculum identity in drought stress mitigation by arbuscular mycorrhizal fungi in soybean

It is well known that arbuscular mycorrhizal fungi (AMF) effects on plant growth largely depend on fungus identity. The objective of this study was to test whether three individual AMF isolates and their mixture mitigate drought stress (DS) differentially in soybean (Glycine max) genotype, predicting that under DS, the mixture of the AMF isolates would provide greater benefits to soybean plants than individual ones. In a greenhouse experiment, a drought-susceptible soybean genotype was inoculated with Septoglomus constrictum, Glomus sp., and Glomus aggregatum, known to be among the most abundant in agricultural and natural soils from central Argentina, and their mixture (Mx). Whereas under well-watered (WW) conditions, individual isolates and Mx treatment were similarly infective; under DS conditions, the Mx treatment showed lower rates of root colonization. Between WW and DS conditions, biomass was decreased in all treatments, although this effect was more marked in non-AM plants. Moreover, AMF strains improved water content and P and N concentrations. Under DS, the Mx treatment was unable to exceed the highest contents that were recorded by AMF isolates. However, under WW conditions, the Mx treatment showed a higher N content than individual isolates. Under both watering conditions, AM plants reduced oxidative damage evaluated as malondiadehyde and chlorophyll content and keep constant osmotic metabolites such as soluble sugars and proline content, without significant differences between AMF isolates and the Mx treatment. These results show that AMF play an important role in mitigating drought impacts on soybean, but that mixtures of AMF isolates did not perform as well as the best single strain inoculum, excluding complementarity effects and suggesting selection effect of AMF on DS alleviation in soybean.     

Differentiation among Maize (<Emphasis Type="Italic">Zea mays</Emphasis> L.) Landraces from the Tarasca Mountain Chain,Michoacan, Mexico and the <Emphasis Type="Italic">Chalqueño</Emphasis> Complex

The classification of Mexican maize (Zea mays L.) begun since the early 20th century, it was consolidated during the middle of this century, but recent additions and rearrangements have been performed by several authors employing new methods of analysis and collections from diverse origin; nevertheless, maize from the State of Michoacan, Mexico has received little attention in regard to its systematic classification. Maize populations from the Tarasca Mountain Chain in Michoacan are commonly considered in literature, as belonging to the Chalqueño race; however, closer observations indicate that significant differences do exist, suggesting the necessity of performing an in-depth study on this respect. Thirty nine native maize populations from the Tarasca Mountain Chain region were evaluated along with 19 typical populations of the Chalqueño, Celaya and Conico races coming from the States of Mexico, Puebla, Hidalgo, Querétaro and Oaxaca. Populations were evaluated in Aranza, Michoacan and Montecillo, Mexico State. Seventeen morphological characters were scored and analyzed by one-way analyses of variance and multivariate techniques. Populations were also genetically analyzed through 17 isozyme loci. Native populations had some alleles not found either in the Chalqueño, Celaya or Conico races, and possess larger genetic diversity. Local populations were congregated into a discrete group apart from the typical Chalqueño populations, suggesting that landraces from the Tarasca Mountain Chain region might not be considered as belonging to the Chalqueño race, but they integrate by themselves a different race.     

Hydrogen metabolism and nitrogen fixation of soybean (Glycine max. L.) inoculated with different strains ofRhizobium spp.

Summary Hydrogenase activities and N₂-fixing capacities of soybean nodules (Glycine max. cv. Hodgson), inoculated with strains ofBradyrhizobium japonicum andRhizobium fredii from different geographical regions, were measured after 35 days of culture under controlled conditions. Of the strains tested, 47% induced nodules with bacteroids which recycled H₂. The data obtained suggest that H₂-recycling ability is not a major factor influencing early N₂-fixation which depends essentially on the precocity and intensity of the initial nodulation.     

Genetic variability of soybean bradyrhizobia populations under different soil managements

Due to their ecologic and economic importance, bradyrhizobia have been extensively studied in recent years. Since 1992, Bradyrhizobium elkanii SEMIA 587 and SEMIA 5019 and Bradyrhizobium japonicum SEMIA 5079 and SEMIA 5080 have been widely used in most Brazilian soybean fields. The objective of this work was to estimate the genetic variability of bradyrhizobial isolates recovered from soils under rhizobial inoculation and different soil managements. Only 25% of the isolates demonstrated high similarities to the original strains, and a strong correlation was obtained between the bradyrhizobial genetic variability and soil management. A high level of genetic diversity was observed both within isolates (H = 5.46) as well as among the different soil practices. Soil under no-tillage presented a higher bradyrhizobia diversity compared with bradyrhizobia isolated from soil under conventional tillage. Serological characterization also indicated that B. elkanii strains SEMIA 587 and SEMIA 5019 were more competitive and presented a higher nodular occupancy capacity than strains belonging to B. japonicum species in Southern Brazilian soils.     

Symbiotic effectiveness of Bradyrhizobium japonicum in acid soils can be predicted from their sensitivity to acid soil stress factors in acidic agar media

In acid soil, low pH, reduced availability of nutrients, and toxicity of Al and Mn limit plant growth and the survival and effectiveness of rhizobia. The symbiosis between legumes and rhizobia is particularly sensitive to acid soil stress. A pot experiment evaluated whether Bradyrhizobium japonicum strain growth on acidic agar media would predict ability to colonize the rhizosphere and form effective nodules in acidic soils. Three Indonesian strains of B. japonicum with similar effectiveness at neutral pH in sand culture but with different tolerance of acid soil stress factors in agar media, and an acid-tolerant commercial strain (CB1809) of comparable effectiveness, were tested in three acid soils using the Al tolerant soybean (Glycine max cv PI 416937). At 7 days after inoculation all strains had achieved large rhizosphere populations, but by day 14 the rhizosphere population of the acid-sensitive strain had decreased, while the more acid-tolerant strains increased. The acid-tolerant strains had significantly greater nodulation and symbiotic effectiveness than plants inoculated with the acid-sensitive strain. Laboratory prescreening of B. japonicum for acid, Al and Mn tolerance in acid media successfully identified strains which were symbiotically competent in low pH soils.     

Improvement of simazine degradation by inoculation of corn and soybean plants with rhizobacteria

We isolated the strains of aerobic bacteria, that were able to use the simazine (2-chloro4,6-bis(ethylamino)-s-triazine) herbicide as a sole source of nitrogen, from the roots of corn plants and the nodules of soybean plants. Partial base sequences of 16S rRNA genes and physiological characteristics of the representative isolates revealed that they belonged to Agrobacterium radiobacter and Bradyrhizobium japonicum, respectively. Inoculation of corn plants with the isolates of A. radiobacter resulted in a faster decrease in the amount of simazine in vermiculite soil. Neither bacteria nor corn plants alone were able to promote the decrease in the amount of simazine. Soybean plants modulated by the isolates of B. japonicum were more sensitive to simazine in a hydroponic medium compared to non-nodulated soybeans, though a significant decrease in the amount of simazine in the media was observed. The results obtained in the present study indicate that the rhizosphere of corn and soybean plants containing root nodules is an appropriate habitat for simazine-degrading bacteria to degrade the herbicide.     

Competitiveness of native Bradyrhizobium japonicum strains in two different soil types

Interstrain competitiveness is a key factor affecting the performance of rhizobium inoculant. In the present study five native strains of Bradyrhizobium japonicum, namely SSF 4, SSF 5, SSF 6, SSF 7 and SSF 8, were assessed for their competitiveness in nodulating soybean using serological methods. The strains were inoculated individually or with the type strain USDA 110 at a 1:1 ratio. Nodule occupancy determined by immunofluorescence and dot immunoblot assay revealed that under in vitro conditions SSF 8 is more competitive than USDA 110 whereas the others were less competitive. The competitive ability of these strains was also estimated in pot culture in the field. In red soil both SSF 8 and USDA 110 were equally competitive whereas in black soil SSF 8 competed better than USDA 110 and produced more nodules. In a black soil field trial using a randomized block design, USDA 110 or SSF 8, when inoculated alone, occupied the majority of the nodules and enhanced nodule dry weight and shoot biomass. SSF 8 was more competitive when the strains were co-inoculated. Received: 1 November 1996     

Diversity,phylogeny and host specificity of soybean and peanut bradyrhizobia

A study on the diversity, phylogeny, and host specificity of soybean (Glycine max L.) and peanut (Arachis hypogaea L.) bradyrhizobia was conducted based on the 16S ribosomal RNA (rRNA) restriction fragment length polymorphisms (RFLPs), 16S rRNA sequencing, and 16S–23S rRNA intergenetic spacer (IGS) RFLP assays. Based on 16S rRNA RFLP assay, tested bradyrhizobia were divided into five genotypes, which could be further clustered into five groups by IGS RFLP assays. According to the 16S rRNA sequencing, strains of IGS-II, IV, and V were phylogenetically related to Bradyrhizobium liaoningense, Bradyrhizobium japonicum, and Bradyrhizobium elkanii, while strains of IGS-Ic and IGS-III related to Bradyrhizobium yuanmingense and Bradyrhizobium canariense, respectively. All isolates could crossly nodulate Phaseolus vulgaris, forming small white nodules. Strains of IGS-II originally isolated from peanut could efficiently nodulate Glycine soja, and two strains isolated from soybean could also nodulate peanut.     

Effects of ectomycorrhizal fungus <Emphasis Type="Italic">Boletus edulis</Emphasis> and mycorrhiza helper <Emphasis Type="Italic">Bacillus cereus</Emphasis> on the growth and nutrient uptake by <Emphasis Type="Italic">Pinus thunbergii</Emphasis>

The present greenhouse study was undertaken to evaluate the effects of co-inoculating the ectomycorrhizal (ECM) fungus Boletus edulis with the mycorrhiza helper bacterium Bacillus cereus HB12 or HB59 on the growth and nutrient uptake of Pinus thunbergii. The inoculation with mycorrhiza helper bacterium significantly (P?≤?0.05) increased the ectomycorrhizal colonization. Treatments with dual inoculum (the mycorrhiza helper bacterium plus mycorrhiza) significantly (P?≤?0.05) increased the P. thunbergii growth. Bacteria–mycorrhizae interactions resulted in a great utilization of phosphate and potassium. The single inoculation resulted in a higher root activity than the control while the co-inoculation led to the highest root activity. The 6-CFDA staining assay showed that B. cereus enhanced fungal activity in ectomycorrhizal symbiosis. The results conclusively suggest that B. cereus isolated from the rhizosphere of P. thunbergii can potentially be used as individual inoculant or co-inoculated with ECM fungi to increase the production in sustainable ecological systems. These results support the potential use of B. cereus (HB12 or HB59) and B. edulis as mixed inoculants stimulating growth of P. thunbergii.     

Polymorphism of <Emphasis Type="Italic">Got2</Emphasis> DNA sequences sheds light on <Emphasis Type="Italic">Aegilops tauschii</Emphasis> Coss. intraspecies divergence and origin of <Emphasis Type="Italic">Triticum aestivum</Emphasis> L.

DNA sequences of nuclear gene Got2 was studied in 60 accessions of Aegilops tauschii, 29 of subsp. tauschii and 31 of subsp. strangulata. It was found that Got2 allozyme polymorphism in Ae. tauschii is due to a single, unique, mutation which led to replacement of glutamic acid by isoleucine in residue 256 of the enzyme molecule, encoded by Got2. As revealed by Got2 DNA sequences variation, initially in its history Ae. tauschii was presented by subsp. strangulata, and among phylogenetic lineages of subsp. strangulata, the lineage “t-9¹s” (TauL3) is the most ancient, a relict one. Subspecies tauschii is relatively “young”. Initially it was presented by the lineage marked by combination of allozyme alleles Got2 ¹⁰⁵ and Acph1 ¹⁰⁰. In the past it inhabited the Continental area from Caucasia to Pakistan, but later on it was forced out by newly originated, now—a major lineage of subsp. tauschii, marked by Got2 ¹⁰⁰. This lineage extended the Continental area of the species up to Kirgizstan, but actually failed to penetrate into pre-Caspian area, occupied by subsp. strangulata. These results essentially differ from those obtained previously, using chloroplast DNA (cpDNA) sequences polymorphism. As revealed by cpDNA, the major, “usual”, subsp. strangulata (TauL2) is “younger” than subsp. tauschii, which resided on phylogenetic tree between relict lineage “t-9¹s”of subsp. strangulata—and major subsp. strangulata. But both cpDNA and Got2 DNA sequences indicate that the level of genetic variation in subsp. tauschii is much lower than in subsp. strangulata. According to Got2 DNA sequences variation, it was Ae. tauschii subsp. strangulata lineage “k-109″ which donated genome D to Triticum aestivum L. This lineage includes accessions: k-109 from South-Eastern Precaspian Azerbaijan; KU-2105, KU-2159 from Western Precaspian Iran; KU-2080 from Eastern Precaspian Iran.     

Azospirillum brasilense Az39 and Bradyrhizobium japonicum E109, inoculated singly or in combination,promote seed germination and early seedling growth in corn (Zea mays L.) and soybean (Glycine max L.)

Inoculants are biological formulations that combine a stable microorganism population and various types of compounds produced and released during fermentation, such as phytohormones and plant growth regulators. Azospirillum brasilense strain Az39 and Brayrhizobium japonicum strain E109 were previously shown to produce indole 3-acetic acid (IAA), gibberellic acid (GA₃) and zeatin (Z). We tested the hypothesis that such compounds are responsible for early growth promotion in inoculated corn (Zea mays L.) and soybean (Glycine max L.) seedlings. Seeds were inoculated with Az39, E109, or both, and kept in a chamber at 20–30 °C under a controlled photoperiod to evaluate seed germination. To evaluate root and shoot length and dry weight, and number of nodules and percentage of nodulated seedlings, in soybean, seedlings were kept in a growth chamber for 14 days under similar photoperiod and temperature conditions. Az39 and E109, singly or in combination, showed the capacity to promote seed germination, nodule formation, and early development of corn and soybean seedlings. Both strains were able to excrete IAA, GA₃ and Z into the culture medium, at a concentration sufficient to produce morphological and physiological changes in young seed tissues.     

Involvement of Bradyrhizobium japonicum denitrification in symbiotic nitrogen fixation by soybean plants subjected to flooding

Denitrification by Bradyrhizobium japonicum bacteroids contributes to nitric oxide (NO) production within soybean nodules in response to flooding conditions. However, the physiological relevance of NO production by denitrification in B. japonicum-Glycine max symbiosis is still unclear. In this work, soybean plants were inoculated with B. japonicum strains lacking the nirK or norC genes which encode the copper-containing nitrite reductase and the c-type nitric oxide reductase enzymes, respectively. 14 days flooding increased nodule number of plants inoculated with the WT and norC strains, but not of plants inoculated with the nirK mutant. However, nodule dry weight was not affected by 14 days flooding regardless of the strain used for inoculation. Supporting this observation, individual nodule growth was significantly higher in plants inoculated with nirK than those inoculated with WT or norC after 14 days flooding. Nodule functioning was strongly inhibited by flooding since leghemoglobin content of the nodules induced by any of the strains was significantly decreased after 7 or 14 days flooding compared to control plants. However, this effect was more relevant in nodules of plants inoculated with the WT or norC mutant than in those inoculated with the nirK mutant. Nitrogen fixation was also estimated by analyzing nitrogen content derived from biological nitrogen fixation in shoots, using the ¹⁵N isotope dilution technique. By using this approach, we observed that the negative effect of 14 days flooding on nitrogen fixation was more pronounced in plants inoculated with the norC mutant. However, nitrogen fixation of plants inoculated with nirK showed the highest tolerance to 14 days flooding. These findings allowed us to demonstrate the previously proposed hypothesis which suggests that NO formed by copper-containing nitrite reductase in soybean nodules, in response to flooding, has a negative effect on nitrogenase activity. We propose that inoculation of soybeans with a B. japonicum nirK mutant, which does not produce NO from nitrate, increases the tolerance of symbiotic nitrogen fixation to flooding.     

Symbiotic efficiency and compatibility of native rhizobia in northern Thailand with different soybean cultivars

Symbiotic efficiency and compatibility of 81 isolates of native bradyrhizobia from irrigated areas in northern Thailand with four soybean cultivars and one cowpea cultivar were evaluated under laboratory conditions. Effectiveness and / or compatibility of the tested isolates were compared with those of a standard strain (Bradyrhizobium japonicum CB 1809) by using plants grown on plastic seed bags. Effectiveness of the isolates was also estimated using uninoculated control plants grown in a nitrogen-free solution. Nodulation of a wide range of host plants by the majority of the tested isolates was observed, which agreed well with the results of our previous field experiment (Shutsrirung et al. 2002: Soil Sci. Plant Nutr., 48, 491–499). Up to 75% of the tested isolates induced a higher growth efficiency than that of the uninoculated control in association with one of the tested cultivars, Black soybean. Comparision with uninoculated control plants, enable to estimate the proportion of the tested isolates leading to effective growth promotion (E + e) of each cultivar, namely, Black soybean (local Thai cultivar), 75%; Cowpea, 82%; SJ5 (commercial Thai cultivar), 33%; Bragg (US cultivar), 33%; and Improved Pelican (US cultivar), 9%. These results indicated that although isolates with a high infectiveness with both “Asian-type” and “US-type” soybeans could be found, a high frequency of isolates leading to inefficient nodules was observed in the US cultivar, suggesting the presence of genetic differences in the soybean cultivars that express high-preference (efficient nodules) or low-preference (inefficient nodules) for a certain group of tested isolates. Based on the results of this laboratory experiment together with our previous field experiment, native rhizobial populations in the irrigated area of northern Thailand could be separated into three groups; Group 1: rhizobium strains showing a high effectiveness with only Asian cultivars, Group 2: strains showing a high effectiveness with only US origin cultivars, and Group 3: strains showing a high effectiveness with both Asian and US origin cultivars. The majority of the native rhizobial populations belonged to Group 1. The isolates in Group 3 may display a high potential for manipulating useful rhizobial inoculant.     

Nuclear and cytoplasmic contributions from <Emphasis Type="Italic">Erianthus arundinaceus</Emphasis> (Retz.) Jeswiet in a sugarcane hybrid clone confirmed through genomic in situ hybridization and cytoplasmic DNA polymorphism

A hybrid between Erianthus arundinaceus (Retz.) Jeswiet and Saccharum spontaneum L. which are wild related species of sugarcane (Saccharum L., Family Poaceae), was repeatedly crossed as female parent with sugarcane commercial varieties to develop near commercial sugarcane clones. The cytoplasm type of the hybrid derivatives were confirmed to be of E. arundinaceus through the mitochondrial and chloroplast DNA polymorphism of nad 4/3-4 intron segment and psbC–trnS segment, respectively. The E. arundinaceus × S. spontaneum hybrid with somatic chromosome number 2n = 62 was confirmed to have 30 chromosomes from E. arundinaceus through genomic in situ hybridization (GISH). The (E. arundinaceus × S. spontaneum) × sugarcane hybrid (2n = 118) had 24 chromosomes from E. arundinaceus whereas its next generation hybrid with sugarcane (2n = 108) had only 12 Erianthus chromosomes. The commercial sugarcane hybrid Co 15015, which is the third generation hybrid with 2n = 106 was confirmed to have two E. arundinaceus chromosomes through GISH. It is the first report of sugarcane with both alien cytoplasm and chromosome contributions from E. arundinaceus.     

Occurrence and nature of mixed infections in nodules of field-grown soybeans (Glycine max)

Summary Mixed infections of Bradyrhizobium japonicum strains in early and late nodules of four soybean cultivars were studied in a field soil. Nodule occupants were identified by immunofluorescence using serogroup specific antibodies prepared against B. japonicum strains USDA 110, USDA 123, and USDA 138. Double infection was determined directly by combined examination of the same microscopic field by fluorescence and phase contrast microscopy. Double strain occupancy was observed consistently, and its occurrence did not differ substantially in pouch, soil pot, and field experiments, ranging in incidence from 12% to 32%. No significant differences in the incidence or nature of double infection could be attributed to cultivar, seed inoculation, or plant maturity. Strains reactive to strain USDA 123-fluorescent antibody were dominant in both singly and doubly infected nodules irrespective of cultivar, plant age, or seed inoculation with strain USDA 110.Paper no. 15092 in the Scientific Journal Series of the Minnesota Agricultural Experiment Station, St. Paul