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Simple Summary: The Asian citrus psyllid is an insect pest of citrus trees that transmits the causal agent of huanglongbing disease. Double-stranded RNA molecules trigger RNA interference (RNAi), a naturally occurring process in eukaryotes that modulates the expression of genes involved in host innate immunity. Use of dsRNA biopesticides offers an alternative to traditional insecticides. In this study, the concentration of dsRNA required to achieve optimal knockdown and mortality for screening exogenous dsRNA by oral delivery was 200 ng/μL. Implementing this pre-established dsRNA regime for laboratory screening of ACP RNAi will enable high-throughput discovery of dsRNA targets and aid in evaluating RNAi biopesticide formulations. Establishing a working threshold can also prevent the use of excessive amounts of dsRNA, which is expensive, can negatively influence RNAi efficiency, and potentially trigger off-target effects. The Asian citrus psyllid (ACP) is a citrus pest and insect vector of "Candidatus Liberibacter asiaticus", the causal agent of citrus greening disease. Double-stranded RNA (dsRNA) biopesticides that trigger RNA interference (RNAi) offer an alternative to traditional insecticides. Standardized laboratory screening of dsRNA requires establishing the minimal effective concentration(s) that result in effective RNAi "penetrance" and trigger RNAi, resulting in one or more measurable phenotypes, herein, significant gene knockdown and the potential for mortality. In this study, knockdown was evaluated for a range of dsRNA concentrations of three ACP candidate genes, clathrin heavy chain (CHC), vacuolar ATPase subunit A (vATPase-A), and sucrose non-fermenting protein 7 (Snf7). Gene knockdown was quantified for ACP teneral adults and 3rd instar nymphs allowed a 48 h ingestion-access period (IAP) on 10, 50,100, 200, and 500 ng/µL dsRNA dissolved in 20% sucrose followed by a 5-day post-IAP on orange jasmine shoots. Significant gene knockdown (p < 0.05) in ACP third instar nymphs and adults ranged from 12–34% and 18–39%, 5 days post-IAP on dsRNA at 10–500 and 100–500 ng/µL, respectively. The threshold concentration beyond which no significant gene knockdown and adult mortality was observed post-48 h IAP and 10-day IAP, respectively, was determined as 200 ng/µL, a concentration indicative of optimal RNAi penetrance. [ABSTRACT FROM AUTHOR]

Simple Summary: Whiteflies (Bemisia tabaci) 'B mitotype' represent an insect pest of fruit, vegetable, ornamental and weed plants. B. tabaci is a cryptic species comprising at least eight endemic haplotypes, of which haplotypes 6 and/or 8 are recognized invasives. The objective of this study was to identify the prevalence and relationships among putative native and exotic begomoviruses and North Africa–Middle East haplotypes in Oman. Several begomoviral species were identified from B. tabaci adults collected from infested plant species, with 67% and 33% representing native and exotic species, respectively. Logistic regression and correspondence analyses predicted 'strong' and 'close' virus–vector associations involving haplotypes 5 and 2 and the exotic chili leaf curl virus and endemic tomato yellow leaf curl virus-OM, respectively. Irrigated agriculture and global trade expansion have facilitated diversification and spread of begomoviruses (Geminiviridae), transmitted by the Bemisia tabaci (Gennadius) cryptic species. Oman is situated on major crossroads between Africa and South Asia, where endemic/native and introduced/exotic begomoviruses occur in agroecosystems. The B. tabaci 'B mitotype' belongs to the North Africa–Middle East (NAFME) cryptic species, comprising at least eight endemic haplotypes, of which haplotypes 6 and/or 8 are recognized invasives. Prevalence and associations among native and exotic begomoviruses and NAFME haplotypes in Oman were investigated. Nine begomoviral species were identified from B. tabaci infesting crop or wild plant species, with 67% and 33% representing native and exotic species, respectively. Haplotypes 2, 3, and 5 represented 31%, 3%, and 66% of the B. tabaci population, respectively. Logistic regression and correspondence analyses predicted 'strong'- and 'close' virus–vector associations involving haplotypes 5 and 2 and the exotic chili leaf curl virus (ChiLCV) and endemic tomato yellow leaf curl virus-OM, respectively. Patterns favor a hypothesis of relaxed virus–vector specificity between an endemic haplotype and the introduced ChiLCV, whereas the endemic co-evolved TYLCV-OM and haplotype 2 virus–vector relationship was reinforced. Thus, in Oman, at least one native haplotype can facilitate the spread of endemic and introduced begomoviruses. [ABSTRACT FROM AUTHOR]

In Saudi Arabia (SA), the citrus greening disease is caused by 'Candidatus Liberibacter asiaticus' (CLas) transmitted by the Asian citrus psyllid (ACP) Diaphorina citri. The origin and route(s) of the ACP-CLas pathosystem invasion in SA have not been studied. Adult ACP were collected from citrus trees in SA and differentiated by analysis of the mitochondrial cytochrome oxidase I (mtCOI) and nuclear copper transporting protein (atox1) genes. A phylogenetic analysis of the Wolbachia spp. surface protein (wsp) gene was used to identify the ACP-associated Wolbachia spp. A phylogenetic analysis of the atox1 and mtCOI gene sequences revealed one predominant ACP haplotype most closely related to the Indian subcontinent founder populations. The detection and identification of CLas in citrus trees were carried out by polymerase chain reaction (PCR) amplification and sequencing of the 16S rDNA gene. The CLas-integrated prophage genomes were sequenced, annotated, and used to differentiate CLas populations. The ML and ASTRAL trees reconstructed with prophages type 1 and 2 genome sequences, separately and concatenated, resolved two major lineages, CLas-1 and -2. The CLas-1 clade, reported here for the first time, consisted of isolates from SA isolates and Pakistan. The CLas-2 sequences formed two groups, CLas-2-1 and -2-2, previously the 'Asiatic' and 'Floridian' strains, respectively. Members of CLas-2-1 originated from Southeast Asia, the USA, and other worldwide locations, while CLas-2-2 was identified only in Florida. This study provides the first snapshot into the status of the ACP-CLas pathosystem in SA. In addition, the results provide new insights into the pathosystem coevolution and global invasion histories of two ACP-CLas lineages with a predicted center of origin in South and Southeast Asia, respectively. [ABSTRACT FROM AUTHOR]

Simple Summary: The whitefly Bemisia tabaci taxon consists of an undefined number of morphologically identical genetic variants of which only a few, including the B, harbor invasive haplotypes. These haplotypes have potential to upsurge and become important pests and plant virus vectors in irrigated agroecosystems worldwide. In the 1980s, unprecedented outbreaks associated with the B variant were reported worldwide, however, the precise origin(s) of the invasive haplotypes has not been determined. In this study, available B. tabaci mitochondrial gene sequences were examined for patterns of conserved single nucleotide changes (SNPs). The whitefly sequence records represented North Africa-Middle Eastern habitats, the proposed B variant center of origin, and distant locales recently invaded by haplotype(s) of the B variant. Unexpectedly, the analysis revealed eight SNPs groups (haplotypes) demonstrating that the genetic architecture of the B mitoype is more complex than previously recognized. Also, the distribution patterns of the eight B haplotypes were tightly linked to well-defined eco-geographic regions, suggesting the different groups have diversified by geographic isolation. Contrary to claims that collectively, the B variant is invasive, only two of the eight haplotypic groups have established in geographical locations outside of their zone of endemism. The Bemisia tabaci cryptic species contains 39 known mitotypes of which the B and Q are best recognized for having established outside their extant endemic range. In the 1980s, previously uncharacterized haplotype(s) of the B mitotype rapidly established in tropical and subtropical locales distant from their presumed center of origin, leading to displacement of several native mitotypes and extreme damage to crops and other vegetation particularly in irrigated agroecosystems. To trace the natural and evolutionary history of the invasive B haplotypes, a phylo-biogeographic study was undertaken. Patterns of single nucleotide polymorphisms (SNPs) and signatures potentially indicative of geographic isolation were investigated using a globally representative mitochondrial cytochrome oxidase I gene (mtCOI) sequence database. Eight haplotype groups within the North Africa-Middle East (NAFME) region were differentiated, NAFME 1–8. The NAFME 1–3 haplotypes were members of the same population that is associated with warm desert climate niches of the Arabian Peninsula and east coastal Africa-Ethiopia. The NAFME 4 and 5 haplotypes are endemic to warm and cold semi-arid niches delimited by the Irano-Turanian floristic region, itself harboring extensive biodiversity. Haplotypes 6 and 7 co-occurred in the Middle East along eastern Mediterranean Sea landmasses, while NAFME 8 was found to be endemic to Cyprus, Turkey, and desert micro-niches throughout Egypt and Israel. Contrary to claims that collectively, the B mitotype is invasive, NAFME 6 and 8 are the only haplotypes to have established in geographical locations outside of their zone of endemism. [ABSTRACT FROM AUTHOR]

The Bemisia tabaci complex of whiteflies contains globally important pests thought to contain cryptic species corresponding to geographically structured phylogenetic clades. Although mostly morphologically indistinguishable, differences have been shown to exist among populations in behavior, plant virus vector capacity, ability to hybridize, and DNA sequence divergence. These differences allow for certain populations to become invasive and cause great economic damage in a monoculture setting. Although high mitochondrial DNA divergences have been reported between putative conspecifics of the B. tabaci species complex, there is limited data that exists across the whole genome for this group. Using data from 2184 orthologs obtained from whole genome sequencing (Illumina), a phylogenetic analysis using maximum likelihood and coalescent methodologies was completed on ten individuals of the B. tabaci complex. In addition, automatic barcode gap discovery methods were employed, and results suggest the existence of five species. Although the divergences of the mitochondrial cytochrome oxidase I gene are high among members of this complex, nuclear divergences are much lower in comparison. Single-copy orthologs from whole genome sequencing demonstrate divergent population structures among members of the B. tabaci complex and the sequences provide an important resource to aid in future genomic studies of the group. [ABSTRACT FROM AUTHOR]