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Simple Summary: Old World bollworm is a moth species that causes significant damage to a wide range of agricultural crops. This species was once confined to the eastern hemisphere but has recently spread throughout South America and the Caribbean and has the potential to establish in North America. Molecular detection methods for Old World bollworm have been developed to track its spread and rapidly differentiate it from the native sibling species, corn earworm. Currently, droplet digital PCR (ddPCR) is a preferred method for bulk screening as it is highly accurate and tolerant of PCR inhibitors; however, real-time PCR is less expensive and more widely available in molecular labs. Improvements to DNA extraction yield and purity are crucial for real-time PCR assay optimization, but these improvements must be time- and cost-efficient. In this study, we improve upon previously published bulk DNA extraction methods by reducing bench time and costly materials. Our results indicate that the addition of caffeine and RNase A improves DNA extraction, resulting in higher amounts of target DNA in real-time PCR. Such improvements will enable the use of high throughput screening methods across multiple platforms to improve the probability of detection of Old World bollworm. Helicoverpa armigera is among the most problematic agricultural pests worldwide due to its polyphagy and ability to evolve pesticide resistance. Molecular detection methods for H. armigera have been developed to track its spread, as such methods allow for rapid and accurate differentiation from the native sibling species H. zea. Droplet digital PCR (ddPCR) is a preferred method for bulk screening due to its accuracy and tolerance to PCR inhibitors; however, real-time PCR is less expensive and more widely available in molecular labs. Improvements to DNA extraction yield, purity, and throughput are crucial for real-time PCR assay optimization. Bulk DNA extractions have recently been improved to where real-time PCR sensitivity can equal that of ddPCR, but these new methods require significant time and specialized equipment. In this study, we improve upon previously published bulk DNA extraction methods by reducing bench time and materials. Our results indicate that the addition of caffeine and RNase A improves DNA extraction, resulting in lower Cq values during real-time PCR while reducing the processing time and cost per specimen. Such improvements will enable the use of high throughput screening methods across multiple platforms to improve the probability of detection of H. armigera. [ABSTRACT FROM AUTHOR]

The genus Acacia is a crucial source of industrial papermaking and timber, possessing significant economic value. Due to their nitrogen-fixing ability, rapid growth, and high tolerance to stress, Acacia species have become integral to short-term industrial timber forests, particularly in southern China and various other regions worldwide. Nevertheless, comparative genomic analyses of Acacia species remain limited, impeding the development of molecular markers for identifying and distinguishing between distinct Acacia species as well as distinguishing their wood counterparts from other tree species. To address this gap, we generated plastomic sequences from eight species commonly cultivated in Chinese plantation forests and compared them with existing data. Our findings revealed a generally conserved structure and gene content within the plastid genomes of Acacia. Notably, the dN/dS ratio of genes from distinct functional groups varied, particularly between ribosomal proteins and photosynthesis genes. Furthermore, phylogenetic analyses and sequence comparisons suggest that the introduction of Acacia to China may have been less diverse than previously thought or that interspecies introgression has occurred more frequently than previously documented. This study offers a valuable starting point for future research in this area and for improving timber quality through marker-assisted breeding. [ABSTRACT FROM AUTHOR]

Maintaining and planting sustainable forests is fundamental in perpetuating the essential functions of these ecosystems. A central aspect of managing forests for future resilience is the consideration of past migration and evolution of trees using genetic and genomic data to ensure that functionally appropriate diversity is conserved and utilized. In our study, we generated and compared genetic and genomic data from the plastome to better understand phylogeography and molecular evolution in the tree species Populus tremuloides (aspen). With these analyses, we found evidence of divergence and migration between northern and southern sites. Additionally, evidence of deep incomplete plastome sorting across the Salicaceae was found when examining insertion–deletion (indel) sites associated with DNA repair. By examining these indels in plastomic genes with introns across Salicaceae, we found a strong correlation between the abundance of DNA repair with genomic position and transcript abundance. From our findings, we conclude that previously ignored plastomic data are essential in understanding phylogeography and the evolution of key metabolic processes for improved aspen forest planning. Given the propensity of aspen forests to host high levels of biodiversity, rapidly sequester carbon, absorb excess nitrogen, and efficiently regulate snowmelt, improvements to planning and conservation will be highly impactful. [ABSTRACT FROM AUTHOR]

Simple Summary: Most insect species, like most other animal species, host a number of different viruses, and because the number of insect species are so great, the diversity of viruses in insects is expected to be larger than in any other animal lineage. Despite this expectation, few studies are conducted to characterize such viruses and fewer still are carried out to describe the evolutionary dynamics between host and virus. Here we studied a virus that infects an agriculturally important moth species corn earworm (Helicoverpa zea) by comparing host and viral genomes. From this comparison it was found that the virus has incorporated several host genes among which the gene cytosolic serine hydroxymethyltransferase is not known from any other virus and thus may provide a unique study system for better understanding the function of this gene. Additionally, undescribed genes were found in both the virus and host that possess sequences originating from bacteria. Lastly the prevalence of the virus was tested across a broad geographic range and found to be present in nearly all host populations tested. Given these findings, more studies should be initiated to characterize insect viruses for novel molecular interactions with possible uses in gene editing and insect biological control. Insect viruses have been described from numerous lineages, yet patterns of genetic exchange and viral prevalence, which are essential to understanding host–virus coevolution, are rarely studied. In Helicoverpa zea, the virus HzNV-2 can cause deformity of male and female genitalia, resulting in sterility. Using ddPCR, we found that male H. zea with malformed genitalia (agonadal) contained high levels of HzNV-2 DNA, confirming previous work. HzNV-2 was found to be prevalent throughout the United States, at more than twice the rate of the baculovirus HaSNPV, and that it contained several host-acquired DNA sequences. HzNV-2 possesses four recently endogenized lepidopteran genes and several more distantly related genes, including one gene with a bacteria-like sequence found in both host and virus. Among the recently acquired genes is cytosolic serine hydroxymethyltransferase (cSHMT). In nearly all tested H. zea, cSHMT contained a 200 bp transposable element (TE) that was not found in cSHMT of the sister species H. armigera. No other virus has been found with host cSHMT, and the study of this shared copy, including possible interactions, may yield new insights into the function of this gene with possible applications to insect biological control, and gene editing. [ABSTRACT FROM AUTHOR]

Curcuma alismatifolia (Zingiberaceae) is an ornamental species with high economic value due to its recent rise in popularity among floriculturists. Cultivars within this species have mixed genetic backgrounds from multiple hybridization events and can be difficult to distinguish via morphological and histological methods alone. Given the need to improve identification resources, we carried out the first systematic study using plastomic data wherein genomic evolution and phylogenetic relationships from 56 accessions of C. alismatifolia were analyzed. The newly assembled plastomes were highly conserved and ranged from 162,139 bp to 164,111 bp, including 79 genes that code for proteins, 30 tRNA genes, and 4 rRNA genes. The A/T motif was the most common of SSRs in the assembled genomes. The Ka/Ks values of most genes were less than 1, and only two genes had Ka/Ks values above 1, which were rps15 (1.15), and ndhl (1.13) with petA equal to 1. The sequence divergence between different varieties of C. alismatifolia was large, and the percentage of variation in coding regions was lower than that in the non-coding regions. Such data will improve cultivar identification, marker assisted breeding, and preservation of germplasm resources. [ABSTRACT FROM AUTHOR]

Identifying alleles associated with adaptation to new environments will advance our understanding of evolution from the molecular level. Previous studies have found that the Populus davidiana southwest population in East Asia has differentiated from other populations in the range. We aimed to evaluate the contributions of the ancestral-state bases (ASBs) relative to derived bases (DBs) in the local adaptation of P. davidiana in the Yunnan–Guizhou Plateau from a quantitative perspective using whole-genome re-sequencing data from 90 P. davidiana samples from three regions across the species range. Our results showed that the uplift of the Qinghai–Tibet Plateau during the Neogene and associated climate fluctuations during the Middle Pleistocene were likely an important factor in the early divergence of P. davidiana. Highly differentiated genomic regions between populations were inferred to have undergone strong linked natural selection, and ASBs are the chief means by which populations of P. davidiana adapt to novel environmental conditions; however, when adapting to regions with high environmental differences relative to the ancestral range, the proportion of DBs was significantly higher than that of background regions, as ASBs are insufficient to cope with these environments. Finally, a number of genes were identified in the outlier region. [ABSTRACT FROM AUTHOR]

DNA originating from organellar genomes are regularly discovered in nuclear sequences during genome assembly. Nevertheless, such insertions are sometimes omitted during the process of nuclear genome assembly because the inserted DNA is assigned to organellar genomes, leading to a systematic underestimation of their frequency. With the rapid development of high-throughput sequencing technology, more inserted fragments from organelle genomes can now be detected. Therefore, it is necessary to be aware of the insertion events from organellar genomes during nuclear genome assembly to properly attribute the impact and rate of such insertions in the evolution of nuclear genomes. Here, we investigated the impact of intracellular gene transfer (IGT) from the plastome to the nuclear genome using genome assemblies that were refined through time with technological improvements from two model species, Arabidopsis thaliana and Oryza sativa. We found that IGT from the plastome to the nuclear genome is a dynamic and ongoing process in both A. thaliana and O. sativa, and mostly occurred recently, as the majority of transferred sequences showed over 95% sequence similarity with plastome sequences of origin. Differences in the plastome-to-nuclear genome IGT between A. thaliana and O. sativa varied among the different assembly versions and were associated with the quality of the nuclear genome assembly. IGTs from the plastome to nuclear genome occurred more frequently in intergenic regions, which were often associated with transposable elements (TEs). This study provides new insights into intracellular genome evolution and nuclear genome assembly by characterizing and comparing IGT from the plastome into the nuclear genome for two model plant species. [ABSTRACT FROM AUTHOR]

The genus Dalbergia contains more than 200 species, several of which are trees that produce traditional medicines and extremely high-value timber commonly referred to as rosewood. Due to the rarity of these species in the wild, the high value of the timber, and a growing international illicit trade, CITES (Convention on International Trade in Endangered Species of Wild Fauna and Flora) has listed the entire genus in appendix II and the species Dalbergia nigra in appendix I because species in this genus are considered at risk of extinction. Given this, and the fact that species or even genus level determination is nearly impossible from cut timber morphology, alternative molecular methods are needed to identify and track intercepted rosewood. To better identify rosewood using molecular methods, we sequenced and assembled eight chloroplast genomes including D. nigra as well as conducted comparative analyses with all other available chloroplast genomes in Dalbergia and closely related lineages. From these analyses, numerous repeats including simple sequence repeats (SSR) and conserved nucleotide polymorphisms unique to subclades within the genus were detected. From phylogenetic analysis based on the CDS from 77 chloroplast genes, the groups Siam rosewood and scented rosewood resolved as monophyletic, supporting the morphological traits used to delimit these species. In addition, several instances of paraphyly and polyphyly resulting from mismatches between taxonomic determinations and phylogenetic tree topology were identified. Ultimately, the highly variable regions in the chloroplast genomes will provide useful plastid markers for further studies regarding the identification, phylogeny, and population genetics of Dalbergia species, including those frequently intercepted in illegal trade. [ABSTRACT FROM AUTHOR]

Qat (Catha edulis, Celastraceae) is a woody evergreen species with great economic and cultural importance. It is cultivated for its stimulant alkaloids cathine and cathinone in East Africa and southwest Arabia. However, genome information, especially DNA sequence resources, for C. edulis are limited, hindering studies regarding interspecific and intraspecific relationships. Herein, the complete chloroplast (cp) genome of Catha edulis is reported. This genome is 157,960 bp in length with 37% GC content and is structurally arranged into two 26,577 bp inverted repeats and two single-copy areas. The size of the small single-copy and the large single-copy regions were 18,491 bp and 86,315 bp, respectively. The C. edulis cp genome consists of 129 coding genes including 37 transfer RNA (tRNA) genes, 8 ribosomal RNA (rRNA) genes, and 84 protein coding genes. For those genes, 112 are single copy genes and 17 genes are duplicated in two inverted regions with seven tRNAs, four rRNAs, and six protein coding genes. The phylogenetic relationships resolved from the cp genome of qat and 32 other species confirms the monophyly of Celastraceae. The cp genomes of C. edulis, Euonymus japonicus and seven Celastraceae species lack the rps16 intron, which indicates an intron loss took place among an ancestor of this family. The cp genome of C. edulis provides a highly valuable genetic resource for further phylogenomic research, barcoding and cp transformation in Celastraceae. [ABSTRACT FROM AUTHOR]

Simple Summary: Invasive species are a constant threat to agriculture throughout the world against which early detection is one of the primary defenses. The Old World bollworm is one of the most important invasive agricultural pests in the world. While historically absent from the Americas, this species was first found in South America in 2013 and poses an ongoing threat of spreading into North America. Surveys are conducted each year, which result in hundreds or thousands of traps that must be screened for this species. Unfortunately, the most common non-target is the native corn earworm, which is nearly identical morphologically to the Old World bollworm and cannot be easily separated. Molecular methods have been developed to screen these trap samples, but the required equipment is expensive and not commonly available. This study details improvements to current molecular methods that will allow for screening of bulk trap samples using standard laboratory instruments and protocols. The ability to perform these methods in nearly any molecular biology lab will greatly enhance our ability to detect and exclude this important pest. Helicoverpa armigera (Hübner) is one of the most important agricultural pests in the world. This historically Old World species was first reported in Brazil in 2013 and has since spread throughout much of South America and into the Caribbean. Throughout North America, H. armigera surveys are ongoing to detect any incursions. Each trap is capable of capturing hundreds of native Helicoverpa zea (Boddie). The two species cannot be separated without genitalic dissection or molecular methods. A ddPCR assay is currently used to screen large trap samples, but this equipment is relatively uncommon and expensive. Here, we optimized a newly designed assay for accurate and repeatable detection of H. armigera in bulk samples across both ddPCR and less costly, and more common, real-time PCR methods. Improvements over previously designed assays were sought through multiple means. Our results suggest bulk real-time PCR assays can be improved through changes in DNA extraction and purification, so that real-time PCR can be substituted for ddPCR in screening projects. While ddPCR remains a more sensitive method for detection of H. armigera in bulk samples, the improvements in assay design, DNA extraction, and purification presented here also enhance assay performance over previous protocols. [ABSTRACT FROM AUTHOR]

Pterocarpus is a genus of trees mainly distributed in tropical Asia, Africa, and South America. Some species of Pterocarpus are rosewood tree species, having important economic value for timber, and for some species, medicinal value as well. Up to now, information about this genus with regard to the genomic characteristics of the chloroplasts has been limited. Based on a combination of next-generation sequencing (Illumina Hiseq) and long-read sequencing (PacBio), the whole chloroplast genomes (cp genomes) of five species (rosewoods) in Pterocarpus (Pterocarpus macrocarpus, P. santalinus, P. indicus, P. pedatus, P. marsupium) have been assembled. The cp genomes of five species in Pterocarpus have similar structural characteristics, gene content, and sequence to other flowering plants. The cp genomes have a typical four-part structure, containing 110 unique genes (77 protein coding genes, 4 rRNAs, 29 tRNAs). Through comparative genomic analysis, abundant simple sequence repeat (SSR)loci (333–349) were detected in Pterocarpus, among which A /T single nucleotide repeats accounted for the highest proportion (72.8–76.4%). In the five cp genomes of Pterocarpus, eight hypervariable regions, including trnH-GUG_psbA, trnS-UGA_psbC, accD-psaI, ndhI-exon2_ndhI-exon1, ndhG_ndhi-exon2, rpoC2-exon2, ccsA, and trnfM-CAU, are proposed for use as DNA barcode regions. In the comparison of gene selection pressures (P. santalinus as the reference genome), purifying selection was inferred as the primary mode of selection in maintaining important biological functions. Phylogenetic analysis shows that Pterocarpus is a monophyletic group. The species P. tinctorius is resolved as early diverging in the genus. Pterocarpus was resolved as sister to the genus Tipuana. [ABSTRACT FROM AUTHOR]

Heimia myrtifolia is an important medicinal plant with several pharmacologically active alkaloids and is also used as an ornamental landscape plant. The purpose of this study is to complete and characterize the chloroplast (cp) genome of H. myrtifolia and compare genomic features to other Myrtales species’ cp genomes. The analysis showed that H. myrtifolia has a total length of 159,219 bp with a typical quadripartite structure containing two identical inverted repeats (IRs) of 25,643 bp isolated by one large single copy (LSC) of 88,571 bp and one small single copy (SSC) of 18,822 bp. The H. myrtifolia cp genome contains 129 genes with eight ribosomal RNAs, 30 transfer RNAs, and 78 protein coding genes, in which 17 genes are duplicated in two IR regions. The genome organization including gene type and number and guanine-cytosine (GC) content is analyzed among the 12 cp genomes in this study. Approximately 255 simple sequence repeats (SSRs) and 16 forward, two reverses, and two palindromic repeats were identified in the H. myrtifolia cp genome. By comparing the whole H. myrtifolia cp genome with 11 other Myrtales species, the results showed that the sequence similarity was high between coding regions while sequence divergence was high between intergenic regions. By employing the full cp genomes for phylogenetic analysis, structural and sequence differences were characterized between H. myrtifolia and 11 Myrtales species illustrating what patterns are common in the evolution of cp genomes within the Myrtales. The first entire cp genome in the genus Heimia provides a valuable resource for further studies in these medicinally and ornamentally important taxa. [ABSTRACT FROM AUTHOR]