Biological nitrogen fixation by two Acacia species and associated root-nodule bacteria in a suburban Australian forest subjected to prescribed burning

Purpose: Prescribed burning is a forest management practice which can lead to nitrogen (N)-limited conditions. This study aimed to explore whether biological N fixation (BNF) remained the main source of N acquisition for two understorey Acacia species in a Eucalyptus-dominated suburban forest of subtropical Australia, 3 to 6 years after prescribed burning. Root-nodule bacteria associated with these acacias were also characterised to unravel the differences in rhizobial communities between sites and species. Material and methods: Two sites, burned 3 and 6 years before sample collection, were selected within a dry subtropical forest of south-east Queensland, Australia. Leaves were collected from individuals of Acacia disparrima and A. leiocalyx at each site to determine leaf total carbon (C) and N content, C and N isotope composition (δ C and δ N) and the percentage of N derived from atmospheric N . Nodules were harvested from both acacia species at each site to isolate root nodule bacteria. Bacterial isolates were processed for 16S rDNA gene sequencing. Results and discussion: Generally, no differences were found in plant physiological variables between the two acacia species. Six years after the fire, both species still depended upon BNF for their N supply, with a higher dependence in winter than in summer. Fire, although of low intensity, was likely to have created a N-limited environment which induced the reliance of legumes on BNF. Root nodule bacteria were dominated by non-rhizobial endophytes, mainly from the Firmicutes phylum. No difference in nodule bacterial diversity was found between sites. The relative abundance of rhizobial genera varied amongst plant species and sites, with a shift in dominance from Bradyrhizobium to Rhizobium species between sites 1 and 2. Conclusions: Our results show that even 6 years after burning, ecosystem remained under N stress and BNF was still the main mechanism for N acquisition by the understorey legumes. 2 2 13 1

Additional file 5: of Identification of candidate genes related to calanolide biosynthesis by transcriptome sequencing of Calophyllum brasiliense (Calophyllaceae)

Coding sequences alignment between PAL-like C. brasiliense unigenes and Arabidopsis PAL1 protein. (TXT 8 kb

Additional file 3: Figures S1-S9. of Identification of candidate genes related to calanolide biosynthesis by transcriptome sequencing of Calophyllum brasiliense (Calophyllaceae)

Figure S1: Gene ontology classification of C. brasiliense unigenes. Figure S2: Biological processes assigned to differentially expressed genes. Figure S3: Re-constructed C. brasiliense metabolic network from glucose to trans-cinnamate. Figure S4: Phylogenetic relationships, primary protein structures and identities percent of trans-cinnamate 4-hydroxylases proteins. Figure S5: Structural and phylogenetic analysis of 4-coumarate: CoA ligases (4CL) from C. brasiliense. Figure S6: Alignment of the 4-coumaroyl 2′-hydroxylases from C. brasiliense. Figure S7: Schematic representation of the Wagner-Meerwein rearrangement. Figure S8: C. brasiliense prenyltransferases. Figure S9: Phylogenetic tree of selected C. brasiliense cytochrome P450. (PPTX 1219 kb

Additional file 11: of Identification of candidate genes related to calanolide biosynthesis by transcriptome sequencing of Calophyllum brasiliense (Calophyllaceae)

Coding sequences alignment between CYP-like unigenes from C. brasiliense and CYP proteins from different plant species involved in furanocoumarins biosynthesis. (TXT 74 kb

Additional file 6: of Identification of candidate genes related to calanolide biosynthesis by transcriptome sequencing of Calophyllum brasiliense (Calophyllaceae)

Coding sequences alignment between C. brasiliense and Arabidopsis PAL proteins. (TXT 16 kb

Additional file 8: of Identification of candidate genes related to calanolide biosynthesis by transcriptome sequencing of Calophyllum brasiliense (Calophyllaceae)

Coding sequences alignment between C. brasiliense and Arabidopsis 4CL proteins. (TXT 18 kb