Transcriptomic Profiling of Various Developmental Stages of Aphis Aurantii to Provide a Genetic Resource for Gene Expression and SSR Analysis.

Abstract

The tea aphid, Aphis aurantii Boyer de Fonscolombe (Hemiptera: Aphididae), is found in regions where the tea plant grows, and has become one of the most important pests in tea gardens in the tropics and subtropics (Han et al., 2012; Deng et al., 2019). This species is also known as the black citrus aphid and destroys citrus orchards (Wang and Tsai, 2001). Ap. aurantii is a polyphagous aphid with over 190 genera hosts, including many other economically important plants in addition to tea and citrus, such as coffee, cacao, loquat, litchi, mango, and camellia (Carver, 1978; Deng et al., 2019). This aphid directly damages trees by sucking the phloem sap out of the shoot tip or from new fresh leaves and injecting its saliva, which causes phytotoxicity and stunting in the plant (Guidolin and Consoli, 2018). Moreover, this aphid secretes honeydew when sap-feeding, and sooty molds frequently grow on the honeydew, which hinders photosynthetic activity (Sevim et al., 2012). Aphids are wide-spread pests that feed on a wide range of fruits and vegetables, most of them are vectors of plant viruses (Huang and Qiao, 2014; Hulle et al., 2020). Except the tea aphid, there have been previous functional studies on genes associated with the development (Ding et al., 2017; Ye et al., 2019), reproduction (Shang et al., 2018; Ullah et al., 2019b), wing development (Shang et al., 2020b), response to the stress (Gao et al., 2018; Jing et al., 2018), and pest control (Mohammed et al., 2018; Ullah et al., 2019a) of other aphids pests (e.g., Ap. citricidus, Ap. gossypii, Acyrthosiphon pisum). Moreover, functional studies of the new insecticide targets in aphids were also conducted in these aphids (Ye et al., 2019; Shang et al., 2020a). However, most previously published papers have focused on the ecology (Alizadeh Kafeshani et al., 2018; Guidolin and Consoli, 2018; Li et al., 2019), control (Aslam et al., 2015; Gholamzadeh-Chitgar and Pourmoradi, 2017), and mitochondrial genome (Wang et al., 2019) in tea aphid Ap. aurantii. There have been few functional studies that focused on the development and reproduction of Ap. aurantii, because there are limited reference sequences. Quantitative and qualitative transcriptome analyses can reveal the integrated biochemical and physiological processes at a molecular level that are associated with specific aspects of the organism, such as identification of the critical genes during the different developmental stages in insects (Morandin et al., 2018; Liu et al., 2020). For instance, RNA-Seq was used to elucidate the underlying molecular mechanisms of metamorphic development of Henosepilachna vigintioctopunctata (Zhang et al., 2018). Using available insect genomes, comparative transcriptome analysis was conducted to analyze gene expression during all developmental stages of Zeugodacus cucurbitae (Wei et al., 2020), Bactrocera dorsalis (Liu et al., 2020). In addition, gene expression has been studied by RNA-Seq in multiple tissues to identify tissue-specific genes involved in female fertility in B. dorsalis [e.g., vitellogenin and vitelline membrane protein in female (Wei et al., 2018, 2019)]. In aphids, RNA-Seq was used to analyze the gene expression between dispersing and non-dispersing morphs (Shang et al., 2016, 2020b). Thus, RNA-Seq technology allows us to determine the gene expression to underlie certain biological functions of critical genes and identify potential targets of new environmentally friendly insecticides for pest control. Simple sequence repeats (SSRs), also known as microsatellites, are short, tandemly arranged, repeating motifs (1–6 bp), which are widely distributed throughout the genomes of eukaryotic organisms (Temnykh et al., 2001). SSRs are co-dominant, hypervariable, neutral, and reproducible molecular markers; therefore, they have become the most widely used molecular markers in population genetic and conservation studies to evaluate the level of genetic variation in a species (King, 2012). Transcriptomic sequencing is also a highly efficient approach to identify SSRs in insects with no accessible genome. This method was extensively used to identify and analyze the SSRs in Liposcelis entomophila (Wei et al., 2013) and H. vigintioctopunctata (Zhang et al., 2018). SSRs were also identified in specific tissues in B. dorsalis (Wei et al., 2015). In this study, RNA-Seq was conducted on samples from six stages of Ap. aurantii with four biological replicates. A comprehensive transcriptome was sequenced, the transcripts were de novo assembled, and the gene's functional annotation was performed. Gene expression during development and SSRs were analyzed. These results will be valuable for the future functional studies of genes involved in Ap. aurantii development, reproduction, and wing differentiation.