Genetic Diversity and Population Structure of Sweet Orange (Citrus sinensis) Germplasm in Inhambane Province, Mozambique

Background/Objectives: Sweet orange (Citrus sinensis (L.) Osbeck) is an economically important fruit crop that contributes substantially to food security and smallholder income in Mozambique. Despite this, the genetic diversity of the locally grown germplasm has not been characterized at the molecular level, limiting its improvement and conservation programs. This study assessed the genetic diversity and population structure of germplasm from 94 sweet orange trees sampled across four districts of Inhambane Province using DArTSeq single-nucleotide polymorphism (SNP) markers. Methods: After filtering 8111 SNPs for call rate (≥0.80) and minor allele frequency (≥0.01), 1263 markers were retained, of which 1144 were anchored to the nine chromosomes of the reference genome. Results: Sparse non-negative matrix factorization identified K = 1, indicating a single undifferentiated gene pool, supported by a smooth PCA scree with one weak axis. DAPC assigned individuals to their district only 38.3% of the time (random expectation = 25%; maximum a-score = 0.10), and the first two PCoA axes explained 10.33% of variation with complete district overlap, indicating no detectable geographic structure. Diversity was low, with observed heterozygosity (Ho = 0.247) exceeding expected heterozygosity (He = 0.138) and a negative inbreeding coefficient (Fis = −0.222). The pattern indicated a heterozygote excess consistent with the fixation of the heterozygous interspecific-hybrid genome under clonal propagation. A hierarchical analysis of molecular variance showed that differentiation among districts was negligible (0.04%), whereas 4.16% of variation was partitioned among orchards (farms) within districts, indicating that the little of the existing structure resides at the orchard level, confounded with propagation method and cultivar, rather than among districts. Most variation was partitioned within individuals (76.0%), and pairwise FST values (0.0005–0.0035) were uniformly low. Conclusions: These results indicate that the sweet orange orchards stem from a single, highly heterozygous gene pool redistributed through the exchange of seed and vegetative planting material. This underscores the need to introduce diverse external germplasm to broaden the genetic base for sustainable improvement in Mozambique.