Skip to main content
NCBI home page
Ecology and Evolution logoLink to Ecology and Evolution
. 2019 May 26;9(12):6999–7004. doi: 10.1002/ece3.5266

Invasive Galega officinalis (Goat's rue) plants in Canada form a symbiotic association with strains of Neorhizobium galegae sv. officinalis originating from the Old World

Eden S P Bromfield 1,✉, Sylvie Cloutier 1, Catherine Robidas 1, Thu Van Tran Thi 1, Stephen J Darbyshire 1
PMCID: PMC6662265  PMID: 31380028

Abstract

The toxic legume plant, Galega officinalis, is native to the Eastern Mediterranean and Black Sea regions. This legume is considered to be a noxious weed, and its establishment in Canada may have resulted from ornamental planting and/or field trials. In its native range, a highly specific nitrogen‐fixing symbiosis with the bacterium, Neorhizobium galegae symbiovar (sv.) officinalis, is required for normal growth. In North America, nothing is known about the bacterial symbionts of G. officinalis. Our purpose was to determine the species and symbiovar identity of symbiotic bacteria associated with invasive plants of G. officinalis at five sites in the province of Ontario, Canada.  Sequence analysis of four housekeeping (16S rRNA, atpD, glnII, and recA) and two symbiosis (nodC and nifH) genes showed that all 50 bacterial isolates from root nodules of G. officinalis at the five Canadian sites were identical to strains of N. galegae sv. officinalis originating either from Europe or the Caucasus. Plant tests indicated that soils collected from four Canadian sites without a history of agriculture or presence of G. officinalis were deficient in symbiotic bacteria capable of eliciting nodules on this plant. Collectively our data support the hypothesis of anthropogenic co‐introduction of G. officinalis and its specific symbiotic bacterium into Canada from the Old World. Factors that may limit the spread of G. officinalis in new environments are discussed.

Keywords: Canada, Galega officinalis (Goat's rue), invasive, Neorhizobium galegae sv. officinalis

1. INTRODUCTION

Galega officinalis (Goat's rue) is a toxic leguminous plant that is considered to have originated in the Eastern Mediterranean and Black Sea regions but has been spread by anthropogenic means to Europe, Western Asia, and North Africa, and more recently to North and South America and New Zealand (CABI, 2019; Osterman et al., 2011; USDA‐ARS, 2019). While G. officinalis has a long history of ornamental and medicinal uses in Europe, it is usually considered a noxious weed because of its invasive characteristics and potential for poisoning livestock (CABI, 2019; CFIA, 2017; Chiej, 1984; EPPO, 2002; Tingey, 1971; USDA‐APHIS, 2010). Since its first introduction to North America in the late 19th Century as an ornamental or for evaluation in field trials (Bailey & Bailey, 1976; CABI, 2019; CFIA, 2017; Macoun, 1908; Tingey, 1971), G. officinalis has become established at several locations in Canada (Figure S1).

The rhizobia are root‐nodule bacteria that fix atmospheric nitrogen in symbiotic association with leguminous plants thereby supplying the host with nitrogen compounds necessary for growth. Galega officinalis and its non‐noxious relative, G. orientalis (fodder galega), form a highly specific symbiotic association with root‐nodule bacteria belonging to the species, Neorhizobium galegae (Andrews & Andrews, 2017; Lindstrom, 1989; Mousavi, Willems, Nesme, Lajudie, & Lindstrôm, 2015). No species other than N. galagae has been reported to induce root nodules on Galega plants (Österman et al., 2014).

Neorhizobium galagae is divided into two symbiovars (symbiotic varieties), officinalis and orientalis, that are capable of eliciting nodules on roots of G. officinalis and G. orientalis, but fix nitrogen only in association with the respective host plant species (Österman et al., 2014; Österman, Mousavi, Koskinen, Paulin, & Lindström, 2015; Radeva et al., 2001).

In preliminary surveys, we identified five sites in the province of Ontario, Canada, harboring established plants of G. officinalis that were nodulated by symbiotic bacteria.

Our purpose was to determine the species and symbiovar identity of the root‐nodule bacteria associated with plants of G. officinalis established at these sites. Bacterial identification was achieved by phylogenetic analysis of four housekeeping and two symbiosis gene sequences.

To determine whether bacteria capable of symbiosis with G. officinalis occur naturally in Canadian soils, we carried out plant infection tests using soils collected from several sites that were without a history of agriculture or G. officinalis.

2. MATERIALS AND METHODS

Five plants of G. officinalis were dug up at random from each four sites (S1–S4) in a 50 km radius of Ottawa as well as from a single site (S5) about 800 km distant in Sault Ste. Marie, Ontario; site descriptions and coordinates are given in Table 1.

Table 1.

Site descriptions

Site Location Coordinates Soil texture Soil pH (1:1, soil:water)
(a) Sites with Galega officinalis plants
S1 Ottawa, Ontario 45°27′10″N, 75°35′19″W Silty clay 7.4
S2 Ottawa, Ontario 45°27′44″N, 75°37′32″W Silty clay loam 7.5
S3 Ottawa, Ontario 45°26′47″N, 75°39′9″W Sandy loam 7.7
S4 Ottawa, Ontario 45°30′10″N, 75°29′50″W Fine loamy sand 7.8
S5 Sault Ste. Marie, Ontario 46°30′59″N, 84°20′40″W Silty loam 7.8
(b) Sites without a history of agriculture or Galega officinalis
S6 Quyon, Québec 45°29′23″N, 76°23′18″W Silty loam 7.3
S7 Carp, Ontario 45°15′59″N, 76°08′52″W Medium to fine loamy sand 7.2
S8 Carp, Ontario 45°15′13″N, 76°08′24″W Fine sandy loam 7.1
S9 Kemptville, Ontario 45°03′49″N, 75°49′20″W Loam 7.4
Open in a new tab

Root nodules were collected from tap and lateral roots of the sampled plants and stored at 4°C in vials containing anhydrous silica gel (Date & Halliday, 1987). Bacteria were isolated from surface sterilized nodules, grown at 28°C on yeast extract mannitol (YEM) agar medium (Tang, Bromfield, Rodrigue, Cloutier, & Tambong, 2012) and purified by repeated streaking and single‐colony picking. Pure bacterial cultures were maintained at −80°C in 20% w/v glycerol.

The 50 bacterial isolates that were analyzed in this study are listed in Table S1; reference taxa are shown in Table S2.

Preparation of bacterial genomic DNA was as described by Tang et al., 2012. Amplification and sequencing of housekeeping (16S rRNA, atpD, glnII, and recA) and symbiosis (nifH [nitrogen fixation] and nodC [nodulation]) genes were carried out using primers and conditions described in Table S3. As suitable primers for amplification and sequencing of the nodC and nifH genes of N. galagae were not available in the literature, we designed primers based on the full genome sequence of N. galegae HAMBI 1141 sv. officinalis (GenBank accession no. HG938357) using Geneious Software (Biomatters Inc., USA) (Table S3). GenBank accession numbers of the nucleotide sequences used in this work are given in Tables S1 and S2.

Bayesian phylogenetic trees were inferred using MrBayes version 3.2.1 (Altekar, Dwarkadas, Huelsenbeck, & Ronquist, 2004) with default priors as previously described (Yu, Cloutier, Tambong, & Bromfield, 2014); Maximum Likelihood (ML) trees (Guindon et al., 2010) were inferred as detailed by Tang et al. (2012) using 1,000 nonparametric bootstrap replications to assess support. Best‐fit substitution models were selected using the Bayesian information criterion implemented in jModelTest version 2 (Darriba, Taboada, Doallo, & Posada, 2012). As tree topologies from Bayesian and ML analyses were similar, only the Bayesian trees are shown in this work.

To determine whether symbiotic bacteria capable of eliciting nodules on G. officinalis occur naturally in Canada, soil samples were collected from four sites (S6–S9, within a 150 km radius of Ottawa, Ontario) that were without a history of agriculture or G. officinalis (Table 1). Thirty random soil samples from each site were collected to a depth of 15 cm with aseptic precautions; soil samples were pooled to form a composite sample for each site. Seeds collected from plants of G. officinalis growing in Ottawa, Ontario, were surface sterilized, grown in axenic Leonard jar assemblies supplied with nitrogen‐free nutrient solution (Tang et al., 2012) and inoculated with soil suspensions (1:1, soil:water). Plants were maintained in a temperature‐controlled growth chamber for 35 days using conditions described by Bromfield et al. (2010). Controls consisted of uninoculated plants and plants inoculated with HAMBI 1141, a nitrogen‐fixing reference strain of N. galegae sv. officinalis and with a bacterial isolate (G122) from Galega plants growing at site S2 (Ottawa).

3. RESULTS AND DISCUSSION

Galega officinalis plants sampled from the five Canadian sites (S1 to S5) were vigorous, showed no signs of nitrogen deficiency, and were extensively nodulated by symbiotic bacteria (Figure S2). Effective nitrogen fixation by bacterial symbionts was indicated by the presence of leghemoglobin (a red colored hemoprotein required for nitrogen fixation), in the interior of the nodules. All five of these sites had moist soils with pH values above 7.0 (Table 1) and showed evidence of significant anthropogenic disturbance and import of soil material. This suggests that the G. officinalis plants at these sites were probably introduced along with imported soil material.

Isolates of symbiotic bacteria from nodules of G. officinalis at Canadian sites produced colonies on YEM agar medium after 7 days at 28°C that were beige, round, convex, ca. 1–1.5 mm diameter and similar to those of N. galagae reference strains, HAMBI 540T (sv. orientalis) and HAMBI 1141 (sv. officinalis).

As was expected based on the high specificity of the Galega–bacterial symbiosis (Österman et al., 2014, 2015; Radeva et al., 2001), analysis of almost full‐length 16S rRNA gene sequences (1,400 bp) indicated that all 50 bacterial isolates from sites S1 to S5 belonged to the genus Neorhizobium as they were placed in a phylogenetic cluster with HAMBI 540T, the type strain of the species, N. galegae (Figure 1).

Figure 1.

Figure 1

Open in a new tab

Bayesian phylogenetic tree of 16S rRNA gene sequences of reference taxa and 50 bacterial isolates from Galega officinalis plants at five Canadian sites (HKY + I substitution model). Only posterior probabilities >90% are shown. Scale bar represents expected number of substitutions per site

Further analyses based on BLASTn database searches revealed that each of the 50 isolates had atpD, glnII, and recA gene sequences that were identical to one of four N. galegae sv. officinalis strains: HAMBI 1183 (=G6 = IAM14208) and HAMBI 1186 (=G10) probably originating from Kew Gardens, United Kingdom (K. Lindström and P. Oivanen, University of Helsinki, Finland, personal communication; Lindstrom, 1989); HAMBI 2425 (=IT1) originating from Italy (Terefework, Kaijalainen, & Lindström, 2001); and HAMBI 2544 (=G032) originating from the Russian Caucasus (Andronov et al., 2003).

A phylogenetic tree (Figure 2) of concatenated atpD–glnII–recA housekeeping gene sequences (1,383 bp) confirmed that the 50 bacterial isolates were divided into four lineages (multiple locus genotypes) each represented by a strain of N. galegae sv. officinalis. The identification of symbiovars in root‐nodule bacteria is based on differences in symbiosis gene sequences (Rogel, Ormeño‐Orrillo, & Romero, 2011). In this connection, BLASTn database searches confirmed that all 50 isolates of N. galegae belong to symbiovar officinalis as they possess nodC (795 bp) and nifH (650 bp) symbiosis gene sequences that are at least 99.9% similar to N. galegae sv. officinalis HAMBI 1141.

Figure 2.

Figure 2

Open in a new tab

Bayesian phylogenetic tree (GTR + G + I substitution model) of atpD–glnII–recA concatenated housekeeping gene sequences (1,383 bp) for reference taxa and 50 bacterial isolates from Galega officinalis plants at five Canadian sites. Roman numerals in bold type refer to lineage numbers. Posterior probabilities >90% are shown. Bar, expected substitutions per site

Data for the frequency of bacterial strains at sites S1–S5 (Table 2) show that at least three of the four strains were detected at each of the Ottawa sites (S1–S4, separated by distances of between 5 and 50 km) whereas at the geographically distant Sault Ste. Marie site (S5, about 800 km from Ottawa) only one strain was encountered. These differences in the relative occurrence of N. galegae sv. officinalis strains between geographically disparate locations may reflect the import of G. officinalis plant material and associated bacterial symbiont into Canada from different sources, or other factors such as sampling error and genetic drift.

Table 2.

Frequency of strains of Neorhizobium galegae sv. officinalis at five Canadian sites based on sequence analysis of atpD, glnII, and recA genes

Lineagea Strainb Bacterial isolates from Total isolates
Ottawa Sault Ste. Marie
Site S1 Site S2 Site S3 Site S4 Site S5
I HAMBI 1183 – 2 – – – 2
II HAMBI 1186 5 4 3 1 – 13
III HAMBI 2544 4 3 4 8 – 19
IV HAMBI 2425 1 1 3 1 10 16
Open in a new tab

Data are based on 10 bacterial isolates from each site.

a

As defined in Figure 2.

b

Strains of Neorhizobium galegae sv. officinalis originating either from Europe or the Caucasus.

In plant tests, negative control plants and plants inoculated with suspensions of soil collected from four sites (S6–S9) without a history of G. officinalis lacked root nodules. These plants had small yellow shoots due to the absence of symbiotic nitrogen fixation. In contrast, positive control plants inoculated with N. galegae sv. officinalis HAMBI 1141 and isolate G122 from site S2 (Ottawa) possessed large dark green shoots and numerous nitrogen‐fixing root nodules. These results indicate that bacterial symbionts of G. officinalis are absent from Canadian soils where the plant host does not occur.

Collectively our data support the hypothesis of anthropogenic co‐introduction of G. officinalis and its specific bacterial symbiont into Canada probably from parts of Europe or the Caucasus. The transport of bacteria either on seed or in soil containing plant material is a possible mechanism that could account for the co‐introduction of G. officinalis and its specific symbiont to Canada.

The fact that symbiotic G. officinalis plants were only found at Canadian sites with soils above pH 7.0 (range 7.4–7.8; Table 1) is consistent with reports from the United States (Oldham & Ransom, 2009) and Spain (González‐Andrés, Redondo, Pescador, & Urbano, 2004) of plants (presumably symbiotic) growing in soils with pH ranges of 7.3–7.5 and 7.7–8.2, respectively. This suggests that the apparent adaptation of G. officinalis to soils above pH 7.0 together with its high level of symbiotic specificity may serve as important factors limiting the spread of the plant in new environments where the specific nitrogen‐fixing bacterial symbiont (N. galegae sv. officinalis) is absent. This is consistent with the observation that G. officinalis is established at only a few localized sites in Canada (Figure S1) despite having been cultivated as early as 1897 (Bailey & Bailey, 1976; Macoun, 1908) and herbarium records showing that the plant was grown in gardens throughout the first half of the 20th Century.

CONFLICT OF INTEREST

The authors declare that there are no conflicts of interest.

AUTHOR CONTRIBUTIONS

ESPB, SC, and SJD conceived the experiments. CR, SC, ESPB, SJD, and TVTT performed the experiments; SC and ESPB analyzed the data. ESPB drafted the manuscript. All authors read, contributed to, and approved the final manuscript.

Supporting information

 

Click here for additional data file. (524.3KB, pdf)

ACKNOWLEDGMENTS

Funding by Agriculture and Agri‐Food Canada (Grant nos. 1318 and 1666) is gratefully acknowledged. We thank Pekka Oivanen and Kristina Lindström, University of Helsinki, Finland, for supplying strains of N. galegae and for providing information on the origins of bacterial strains. We are grateful to Xiaoyuan Geng and Ben Stewart, Agriculture and Agri‐Food Canada, Ottawa, for providing information on soil characteristics.

Bromfield ESP, Cloutier S, Robidas C, Tran Thi TV, Darbyshire SJ. Invasive Galega officinalis (Goat's rue) plants in Canada form a symbiotic association with strains of Neorhizobium galegae sv. officinalis originating from the Old World. Ecol Evol. 2019;9:6999–7004. 10.1002/ece3.5266

Data Availability Statement: Nucleotide sequences of bacterial isolates from G. officinalis plants in Canada were deposited in DDBJ/GenBank, under Accession Numbers: KT869496‐KT869545 (16S rRNA), KT869546‐KT869595 (atpD), KT869596‐KT869645 (glnII), KT869646‐KT869695 (recA), KT869696‐KT869745 (nifH), and KT869746‐KT869795 (nodC).Voucher specimens of Galega officinalis plants collected at Canadian sites S1–S5 are available at the Vascular Plant Herbarium (DAO), Agriculture and Agri‐Food Canada, 960 Carling Ave., Ottawa, Ontario, K1A 0C6, Canada. http://www.agr.gc.ca/eng/science-and-innovation/agriculture-and-agri-food-research-centres-and-collections/national-collection-of-vascular-plants-dao.All other data are presented in the Supporting Information.

DATA ACCESSIBILITY

Nucleotide sequences of bacterial isolates from G. officinalis plants in Canada were deposited in DDBJ/GenBank, under Accession Numbers: KT869496‐KT869545 (16S rRNA), KT869546‐KT869595 (atpD), KT869596‐KT869645 (glnII), KT869646‐KT869695 (recA), KT869696‐KT869745 (nifH), and KT869746‐KT869795 (nodC).

Voucher specimens of Galega officinalis plants collected at Canadian sites S1–S5 are available at the Vascular Plant Herbarium (DAO), Agriculture and Agri‐Food Canada, 960 Carling Ave., Ottawa, Ontario, K1A 0C6, Canada. http://www.agr.gc.ca/eng/science-and-innovation/agriculture-and-agri-food-research-centres-and-collections/national-collection-of-vascular-plants-dao.

All other data are presented in the Supporting Information.

REFERENCES

  1. Altekar, G. , Dwarkadas, S. , Huelsenbeck, J. P. , & Ronquist, F. (2004). Parallel metropolis coupled Markov chain Monte Carlo for Bayesian phylogenetic inference. Bioinformatics, 20, 407–415. 10.1093/bioinformatics/btg427 [DOI] [PubMed] [Google Scholar]
  2. Andrews, M. , & Andrews, M. E. (2017). Specificity in legume‐rhizobia symbioses. International Journal of Molecular Sciences, 18, 705 10.3390/ijms18040705 [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Andronov, E. E., Terefework, Z., Roumiantseva, M. L., … K. (2003). Symbiotic and genetic diversity of Rhizobium galegae isolates collected from the Galega orientalis gene center in the Caucasus. Applied and Environmental Microbiology, 69, 1067–1074. 10.1128/AEM.69.2.1067-1074.2003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bailey, L. G. , & Bailey, E. Z. (1976). Hortus Third. A concise dictionary of plants cultivated in the United States and Canada (p. 1290). New York, NY: Macmillan Publishing Co. [Google Scholar]
  5. Bromfield, E. S. P. , Tambong, J. T. , Cloutier, S. , Prévost, D. , Laguerre, G. , & van Berkum, P. (2010). Ensifer, Phyllobacterium and Rhizobium species occupy nodules of Medicago sativa (alfalfa) and Melilotus alba (sweet clover) grown at a Canadian site without a history of cultivation. Microbiology, 156, 505–520. [DOI] [PubMed] [Google Scholar]
  6. CABI (2019). Galega officinalis (goatsrue) In Invasive species compendium. Wallingford, UK: CAB International; Retrieved from www.cabi.org/isc/datasheet/24756 [Google Scholar]
  7. CFIA (2017). Weed seed: Galega officinalis (goat's rue). Ottawa, Ontario, Canada: Canadian Food Inspection Agency; Retrieved from http://inspection.gc.ca/plants/seeds/testing-grading/seeds-identification/galega-officinalis/eng/1405080573911/1405080574833 [Google Scholar]
  8. Chiej, R. (1984). The Macdonald encyclopedia of medicinal plants (p. 447). London, UK: MacDonald. [Google Scholar]
  9. Darriba, D. , Taboada, G. L. , Doallo, R. , & Posada, D. (2012). jModelTest 2: More models, new heuristics and parallel computing. Nature Methods, 9, 772 10.1038/nmeth.2109 [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Date, R. A. , & Halliday, J. (1987). Collection, isolation, cultivation and maintenance of rhizobia In Elkan G. H. (Ed.), Symbiotic nitrogen fixation technology (pp. 6999–27). New York, NY: Marcel Dekker. [Google Scholar]
  11. EPPO (2002). Global database. Paris, France: European and Mediterranean Plant Protection Organization; Retrieved from https://gd.eppo.int/taxon/GAGOF [Google Scholar]
  12. González‐Andrés, F. , Redondo, P. A. , Pescador, R. , & Urbano, B. (2004). Management of Galega officinalis L. and preliminary results on its potential for milk production improvement in sheep. New Zealand Journal of Agricultural Research, 47, 233–245. [Google Scholar]
  13. Guindon, S. , Dufayard, J. F. , Lefort, V. , Anisimova, M. , Hordijk, W. , & Gascuel, O. (2010). New algorithms and methods to estimate maximum‐likelihood phylogenies: Assessing the performance of PhyML 3.0. Systematic Biology, 59(3), 307–321. 10.1093/sysbio/syq010 [DOI] [PubMed] [Google Scholar]
  14. Lindstrom, K. (1989). Rhizobium galegae, a new species of legume root nodule bacteria. International Journal of Systematic and Evolutionary Microbiology, 39, 365–367. 10.1099/00207713-39-3-365 [DOI] [Google Scholar]
  15. Macoun, W. T. (1908). List of herbaceous perennials tested in the arboretum and botanic garden: Central Experimental Farm, Ottawa, Canada, with descriptions of flowers, and other notes. Canadian Department of Agriculture Bulletin 5, 2nd Series. [Google Scholar]
  16. Mousavi, S. A. , Willems, A. , Nesme, X. , de Lajudie, P. , & Lindstrôm, K. (2015). Revised phylogeny of Rhizobiaceae: Proposal of the delineation of Pararhizobium gen. nov., and 13 new species combinations. Systematic and Applied Microbiology, 38, 84–90. 10.1016/j.syapm.2014.12.003 [DOI] [PubMed] [Google Scholar]
  17. Oldham, M. , & Ransom, C. V. (2009). Goatsrue (Galega officinalis) seed biology. Weed Science, 57, 149–154. [Google Scholar]
  18. Osterman, J. , Chizhevskaja, E. P. , Andronov, E. E. , Fewer, D. P. , Terefework, Z. , Roumiantseva, M. L. , … Lindström, K. (2011). Galega orientalis is more diverse than Galega officinalis in Caucasus–whole‐genome AFLP analysis and phylogenetics of symbiosis‐related genes. Molecular Ecology, 20, 4808–4821. [DOI] [PubMed] [Google Scholar]
  19. Österman, J. , Marsh, J. , Laine, P. K. , Zeng, Z. , Alatalo, E. , Sullivan, J. T. , … Lindström, K. (2014). Genome sequencing of two Neorhizobium galegae strains reveals a noeT gene responsible for the unusual acetylation of the nodulation factors. BMC Genomics, 15, 500 10.1186/1471-2164-15-500 [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Österman, J. , Mousavi, S. A. , Koskinen, P. , Paulin, L. , & Lindström, K. (2015). Genomic features separating ten strains of Neorhizobium galegae with different symbiotic phenotypes. BMC Genomics, 16, 348 10.1186/s12864-015-1576-3 [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Radeva, G. , Jurgens, G. , Niemi, M. , Nick, G. , Suominen, L. , & Lindstrom, K. (2001). Description of two biovars in the Rhizobium galegae species: Biovar orientalis and biovar officinalis. Systematic and Applied Microbiology, 24, 192–205. 10.1078/0723-2020-00029 [DOI] [PubMed] [Google Scholar]
  22. Rogel, M. A. , Ormeño‐Orrillo, E. , & Romero, E. M. (2011). Symbiovars in rhizobia reflect bacterial adaptation to legumes. Systematic and Applied Microbiology, 34(2), 96–104. 10.1016/j.syapm.2010.11.015 [DOI] [PubMed] [Google Scholar]
  23. Tang, J. , Bromfield, E. S. P. , Rodrigue, N. , Cloutier, S. , & Tambong, J. T. (2012). Microevolution of symbiotic Bradyrhizobium populations associated with soybeans in east North America. Ecology and Evolution, 2, 2943–2961. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Terefework, Z. , Kaijalainen, S. , & Lindström, K. (2001). AFLP fingerprinting as a tool to study the genetic diversity of Rhizobium galegae isolated from Galega orientalis and Galega officinalis . Journal of Biotechnology, 91, 169–180. 10.1016/S0168-1656(01)00338-8 [DOI] [PubMed] [Google Scholar]
  25. Tingey, D. C. (1971). Goatsrue, a potential forage crop, turned out to be a weed. Utah Science, 32, 25–28. [Google Scholar]
  26. USDA‐APHIS (2010). Federal noxious weed list. Retrieved from https://www.aphis.usda.gov/plant_health/plant_pest_info/weeds/downloads/weedlist.pdf [Google Scholar]
  27. USDA‐ARS (2019). USDA, agricultural research service, national plant germplasm system. Germplasm Resources Information Network (GRIN‐Taxonomy). Beltsville, MD: National Germplasm Resources Laboratory; Retrieved from https://npgsweb.ars-grin.gov/gringlobal/taxonomydetail.aspx?xml:id=70971. Accessed 4 March 2019. [Google Scholar]
  28. Yu, X. , Cloutier, S. , Tambong, J. T. , & Bromfield, E. S. P. (2014). Bradyrhizobium ottawaense sp. nov., a symbiotic nitrogen fixing bacterium from root nodules of soybeans in Canada. International Journal of Systematic and Evolutionary Microbiology, 64, 3202–3207. [DOI] [PMC free article] [PubMed] [Google Scholar]

Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

 

Click here for additional data file. (524.3KB, pdf)

Data Availability Statement

Nucleotide sequences of bacterial isolates from G. officinalis plants in Canada were deposited in DDBJ/GenBank, under Accession Numbers: KT869496‐KT869545 (16S rRNA), KT869546‐KT869595 (atpD), KT869596‐KT869645 (glnII), KT869646‐KT869695 (recA), KT869696‐KT869745 (nifH), and KT869746‐KT869795 (nodC).

Voucher specimens of Galega officinalis plants collected at Canadian sites S1–S5 are available at the Vascular Plant Herbarium (DAO), Agriculture and Agri‐Food Canada, 960 Carling Ave., Ottawa, Ontario, K1A 0C6, Canada. http://www.agr.gc.ca/eng/science-and-innovation/agriculture-and-agri-food-research-centres-and-collections/national-collection-of-vascular-plants-dao.

All other data are presented in the Supporting Information.

Articles from Ecology and Evolution are provided here courtesy of Wiley

RESOURCES