Enumeration of citrus endophytic bacterial communities based on illumine metagenomics technique

Citrus is a valuable crop in Pakistan. It is rich in vitamin C, other nutrients and antioxidants. Huanglongbing (HLB) caused by a bacterium "Candidatus liberibacter asiaticus" (CLas), africanus and americanus has an influence on citrus production around the world. Beside HLB there exist several other bacterial species in citrus groves in Pakistan. The structure and diversity of bacterial species in various ecosystems can be quickly examined using NGS. This approach is considerably quicker and more precise than outdated methods. Healthy or citrus greening infected leaf samples of Grapefruit (Citrus paradisi), C. aurantifolia, and C. reticulata Blanco were used for diversity analysis. In this study high throughput, NGS technique was used to access the population of both cultivable and non-cultivable bacterial endophytes from citrus leaves, by using PCR amplicons of 16S rDNA sequences (V5-V7 regions) with Illumina Hi seq. As a result, a total number of 68,722 sequences were produced from the test samples. According to the NGS-based diversity classification, the most common genera of exploited bacterial endophytes were Proteobacteria, Firmicutes, Bacteroides, Cyanobacteria, and Actinobacteria. C. aurantifolia and C. paradisi showed almost equal diversity, whereas C. reticulata Blanco had a higher proportion of Proteobacteria and Cyanobacteria in their leaves. To determine alpha diversity (AD), additional data was analyzed using statistical indices such as Shannon, Chao1, and Simpson. According to the inverse Simpson diversity index, the abundance of the microbial population in six different citrus samples was 0.48, 0.567, and 0.163, respectively. The metagenomics of microbiota in plant tissues was successfully recorded by NGS technology, which can help us learn more about the interactions between plants and microbes. This research is the first step toward a better understanding of 16SrRNA-based metagenomics from citrus in Pakistan using Illumina (Hi seq) Technology.