Seed germination is a crucial developmental event in the plant life cycle. Proper germination significantly impacts the yield and quality. This study focuses on macadamia nuts, exploring the physiological, metabolic, and molecular biological characteristics during seed germination. The water content of macadamia seeds reached a peak in the seed imbibition stage, followed by a gradual decline. Key components such as fats, proteins, and soluble sugars decreased consistently. The enzyme activities of superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and glutathione peroxidase (GPX) increased throughout seed germination, reaching peak levels in leaf growth stages. A total of 1523 metabolites and 13,035 differentially expressed genes (DEGs) were detected by transcriptomic and metabolomic analyses, among which 1320 were transcription factors. The transcriptome and metabolome integration analysis showed significant overlaps between DEGs and differential metabolites in pathways such as phenylpropanoid biosynthesis. Quercetin, trifolin, rutin, myricetin, and quercetin 3-beta-D-sophoroside may play important roles in seed germination. FG2 and CYP75A/CYP75B1 were key genes in the flavonoid pathway. Hormones such as auxin (IAA), cytokinin (CTK), gibberellin (GA), and abscisic acid (ABA) exhibited stage-specific changes during germination. This study provided important theoretical basis and practical guidance for optimizing seed germination rates and enhancing stress resistance capabilities.

Macadamia integrifolia husk has great potential for use as a feedstuff in ruminants. However, little information regarding its effects on ruminal fermentation traits is available at present. Hence, this in vitro study aims to investigate the effects of Macadamia integrifolia husk as a substrate, supplemented with enzymes at different levels (0 g/kg cellulases + pectinases, CON; 0.5 cellulases + 0.5 g/kg pectinases, TRE1; 1.0 g/kg cellulases + 0.5 g/kg pectinases, TRE2; 1.5 cellulases + 0.5 g/kg pectinases, TRE3), on gas production, fermentation traits, and bacterial communities in goats. The results demonstrate that gas production was increased in TRE1 and TRE3 groups compared to the CON group at 6, 9, 12, 24, and 48 h (p < 0.05); DMD was the highest in the TRE3 group and the lowest in the CON group at 6, 12, 24, and 48 h (p < 0.05); and NDFD was the highest in the TRE1 group and the lowest in the CON group at 6 h, then the highest in the TRE3 group and the lowest in the CON group at 12, 24, and 48 h (p < 0.05). The concentrations of MCP, TVFAs, acetate, and propionate were the highest in the TRE3 group and the lowest in the CON group (p< 0.05). The abundance of Bacillota was the highest in the TRE1 group and the lowest in the TRE2 group (p < 0.05), whereas that of Synergistota and Actinomycetota was the lowest in the CON group and the highest in the TRE2 group (p < 0.05). Acidobacteriota was the most abundant in the TRE3 group and the least abundant in the TRE1 group (p < 0.05). The most abundant genus was norank_p_Bacteroidota, comprising approximately 29.3%, 30.6%, 30.4%, and 31.7% of the total bacteria in the CON, TRE1, TRE2, and TRE3 groups, respectively; this was followed by Succiniclasticum, comprising approximately 6.3%, 5.8%, 6.1%, and 7.0% of the total bacteria in the CON, TRE1, TRE2, and TRE3 groups, respectively. This study provides new insights regarding the use of Macadamia integrifolia husk as a feedstuff in goats; in particular, supplementation with cellulases and pectinases could effectively improve its utilization. The results suggest that the optimal supplementary levels of cellulases and pectinases are 1.5 g/kg and 0.5 g/kg, respectively.

The pericarp of Macadamia integrifolia represents a promising but underexplored source of functional flavonoids. To systematically elucidate their biosynthesis and enhance the industrial potential of this by-product, we conducted integrated transcriptomic and metabolomic profiling of pericarps across five developmental stages (50, 80, 110, 140, and 170 days after flowering). Our analysis reveals, for the first time, a distinct temporal shift in both gene expression and metabolite accumulation. Early stages were characterized by high expression of PAL, 4CL, CHS, and FLS, coupled with abundant flavonols and anthocyanins. In contrast, late stages exhibited upregulation of CHI and F3’5’H, redirecting the metabolic flux toward flavanones and isoflavones. This dynamic profile was closely associated with jasmonate and gibberellin signaling pathways and was likely regulated by key transcription factors (MYB, NAC, bHLH). These findings provide a multi-omics framework that elucidates the temporal flavonoid biosynthesis in macadamia pericarp, thereby laying the groundwork for its future industrial valorization.

Color serves as a crucial visual signal for attracting pollinating insects and directly affects the fruit set rate in woody crops. This study investigated the molecular mechanisms underlying flower color formation in macadamia. The results demonstrated that darker flower colors were associated with higher fruit set rates: the rates for purple, pink, pinkish-white, and white flowers were 2.78, 1.99, 1.35, and 1.31, respectively. High-throughput sequencing identified 1359 differentially accumulated metabolites, including benzoic acid, 4-hydroxybenzaldehyde, and isorhamnetin. Transcriptional regulators such as ERF, MYB, and WRKY were significantly up-regulated in darker flowers. KEGG analysis revealed two key metabolic pathways, in which genes including HCT (shikimate hydroxycinnamoyl transferase) and F3GalTase (flavonol 3-O-galactosyltransferase), as well as related metabolites such as p-coumaric acid, chlorogenic acid, and myricetin, showed higher expression levels in darker flowers. Anthocyanin content was highest in pink and pinkish-purple varieties (462.79 and 446.35 μg/g, respectively), and lower in white and light pink varieties (140.52 and 167.97 μg/g). In conclusion, flower color intensity is positively correlated with both fruit set rate and anthocyanin content. Genes involved in the flavonoid and phenylpropanoid pathways, along with transcription factors such as WRKY and MYB, collectively regulate flower color formation. This study provides a theoretical basis for macadamia flower color breeding.

Macadamia integrifolia is a valuable tropical and subtropical crop, yet its large-scale propagation remains constrained by difficulties in adventitious rooting. ‘Guire 1’, the dominant cultivar in Guangxi, China, accounts for over 65% of the local planting area, but its rooting ability and the underlying regulatory mechanisms remain poorly understood. Using one-year-old lignified branches of ‘Guire 1’, we examined structural, physiological, and molecular characteristics during the rooting process. Measurements included nutrient content, oxidase activity, and endogenous hormone levels across developmental stages, while integrated transcriptomic and metabolomic analyses were conducted to identify key pathways and genes. The findings demonstrated that macadamia cuttings rooted through the callus formation pathway. During rooting, malondialdehyde (MDA), soluble sugar, and soluble protein levels gradually declined, with no significant changes in later stages. Activities of superoxide dismutase (SOD) and peroxidase (POD) gradually decreased, with peroxidase maintaining relatively high activity in later stages. Free proline, abscisic acid (ABA), and jasmonic acid (JA) contents peaked at early stages and then decreased, whereas indole-3-acetic acid (IAA) peaked at the root primordium initiation stage. Zeatin riboside (ZR) and salicylic acid (SA)were highest at the initial stage. During the rooting process, a total of 1722 differential metabolites (DAMs), 14,482 differentially expressed genes (DEGs), and 699 transcription factors were identified. Integrated analyses identified plant hormone signal transduction, phenylpropanoid biosynthesis, flavonoid biosynthesis, and starch and sucrose metabolism as the crucial pathways involved in macadamia cutting rooting. These pathways regulated hormone balance, lignin synthesis, vascular differentiation, reactive oxygen species scavenging, and energy supply during rooting. This study elucidates the rooting mechanism in macadamia cuttings, supporting germplasm preservation and large-scale propagation techniques.

Macadamia is an emerging woody nut with high economic value. Although genome assemblies are now available for all four species, a comprehensive evolutionary analysis integrating divergence time estimation, lineage-specific gene family dynamics, and whole-genome duplication (WGD) history has not yet been undertaken. Here, we performed a comparative genomic analysis of four macadamia species (M. integrifolia, M. tetraphylla, M. jansenii, and M. ternifolia) alongside 11 other plant species. A total of 72,992 gene families were identified, with species-specific gene families numbering 277 in M. integrifolia, 234 in M. tetraphylla, 236 in M. ternifolia, and 183 in M. jansenii. KEGG enrichment analysis revealed that M. integrifolia-unique genes were enriched in terpenoid biosynthesis and ABC transporter pathways, whereas the unique genes of the other three species were predominantly associated with phenylpropanoid biosynthesis and amino acid/carbohydrate metabolism. Phylogenetic analysis revealed a well-supported monophyletic macadamia clade, with M. integrifolia and M. tetraphylla forming one subclade, and M. ternifolia and M. jansenii forming another. Divergence time estimation indicated that the genus diverged approximately 10.3 million years ago (Mya), with species-level divergences occurring at 7.8–8.4 Mya. Genomic synteny analysis revealed a conserved genomic architecture, with 57.36%–59.68% of genes being shared in syntenic blocks among species. Gene family dynamics revealed lineage-enriched patterns in the terpenoid biosynthesis pathways: expansion in M. integrifolia but contraction in the other three species. WGD analysis detected two WGD events. On a relative Ks scale, the more recent WGD event overlapped with the species-level divergence, whereas the older WGD event was shared by the common ancestor of the genus. These findings provide a comprehensive genomic foundation for understanding macadamia evolution and may facilitate breeding and conservation efforts.