National Clonal Germplasm Repository for Citrus

Huanglongbing (HLB) is a devastating citrus disease that threatens the citrus industry worldwide. HLB is associated with the bacteria Candidatus Liberibacter asiaticus (CLas) and as of today, there are no tools for economically viable disease management. Several wild Australian limes have been identified to be HLB resistant and their resistance is hypothesized to be conferred by resistance genes (R-genes), which mediate pathogen-specific defense responses. The aim of this study was to gain insight into the genomic features of R-genes in Australian limes, in comparison to susceptible citrus cultivars. In this study, we used five citrus genomes, including three Australian limes (Citrus australasica, C. glauca and C. inodora) and two cultivated citrus species (C. clementina and C. sinensis). Our results indicate up to 70% of the R-genes were identified in the unannotated regions in the original genome annotation of each species, owing to the use of a R-gene specific pipeline. Surprisingly, the two cultivated species harbored 15.8 to 104% more R-genes than the Australian limes. In all species, over 75% of the R-genes occurred in clusters and nearly 80% were concentrated in three chromosomes (Chr3, 5 and 7). The syntenic R-gene based phylogenic classification grouped the five species according to their HLB-resistance levels, reflecting the association between these R-genes and their distinct Australian origins. Domain structure analysis revealed substantial similarities in the R-genes between wild Australian limes and cultivated citrus. Investigation of chromosomal sites underlying Australian specific R genes revealed diversifying selection signatures on several chromosomal regions. The findings in this study will aid in the development of tools for genome-assisted breeding for HLB-resistant varieties.

Simple SummaryThe cotton seed bug, Oxycarenus hyalinipennis, is an invasive pest native to southern Europe and North Africa. It feeds on plants in the mallow family, including the agricultural commodities cotton, okra, hibiscus, cocoa, and kenaf. The O. hyalinipennis is a major economic threat to the U.S. cotton industry. Based on host and climate availabilities and the lack of natural enemies, O. hyalinipennis has a high likelihood for establishment throughout the southern regions in the U.S. The fundamental life history and reproductive behavior of O. hyalinipennis have been inadequately explored until now; therefore, it is difficult to develop corresponding precautionary measures. In this research, we have conducted a comprehensive study on the life cycle of O. hyalinipennis, including egg development, metamorphosis, adult mating behavior, adult lifespan, and survivorship. Our results provide valuable information for scientists and growers to develop efficient management strategies in IPM for timely infestation intervention to protect the cotton crop and other agricultural commodities from O. hyalinipennis damage.Understanding the fundamental life cycle and reproductive behavior of a pest insect is essential for developing efficient control strategies; however, much of this knowledge remains elusive for a multitude of insects, including the cotton seed bug, Oxycarenus hyalinipennis. Here, we report the results of our comprehensive study on the cotton seed bug’s life cycle, including mating behavior, adult lifespan, and egg-to-adulthood development. Our findings showed that adult males and females began mating as early as three days after emerging (75%), and the frequency of mating increased to 100% by the fifth day. Mated females commenced oviposition on cotton seeds as early as two days after mating, with a cumulative mean number of 151 fertile eggs oviposited during the first oviposition cycle. Furthermore, around 10% of eggs from both mated and unmated females remained unfertilized. The first instar nymphs began emerging approximately seven days following oviposition. To track their development, we monitored the newly hatched nymphs daily until they reached adulthood. There were five nymphal stages, which cumulatively took roughly 28 to 30 days. Notably, mating positively influenced the survivorship and lifespan of adult O. hyalinipennis. Mated males and females exhibited median lifespans of 28 and 25 days, respectively. In contrast, unmated males and females only lived for a median lifespan of 9.5 days, about one-third that of the mated O. hyalinipennis. Our study provides key insights into the O. hyalinipennis life history for new IPM strategies.

The date palm (Phoenix dactylifera L.) is one of the oldest cultivated fruit crops. It may have originated as a species from a primitive Phoenix ancestor and shows genes shared with or derived from other Phoenix species. Most probably, the date palm originated in the arid subtropical deserts of the Middle East near current Iraq and Iran. It was probably first domesticated and cultivated in that area approximately 5000 years bp, its cultivation later expanding into Egypt around 3500 years bp (before present era), and into India, western North Africa, and Southern Europe by 2000 years bp. The date palm may be considered “intermediately” domesticated as it maintains some characteristics of undomesticated crops but also has some characteristics of intermediate and fully domesticated crops. In addition to archeological evidence of its cultivation, various artistic representations of date cultivation show both cultivation and simple forms of most cultural practices used in modern date production, particularly artificial pollination. The latter led to an association of the date palm with ancient fertility goddesses, and it was later integrated into the three great monotheistic religions. Date cultivation later expanded outside the traditional area of date cultivation and is today cultivated in over 40 countries around the world, although the ancient center of date culture remains the most import region producing dates.

Citrus is among the most economically important fruit crops. Its vast species diversity and global production was observed by N.I. Vavilov during his international plant explorations from the early to mid-1900s. Currently, ex situ citrus collections located around the world conserve and protect citrus genetic resources, as revealed in a survey conducted in 2021. Responses were received from 43 collections in 27 countries, of which 35 provided data regarding collection composition, management practices, and security, as well as other information. The six largest citrus collections have between 1000 and 1735 accessions. The largest accession holdings are mandarins and sweet oranges, although all citrus fruit types are maintained: mandarin, sweet orange, lemon, pummelo, grapefruit, hybrids, lime, sour orange, citron, kumquat, papeda, finger lime, and crop wild relatives. Diseases pose significant threats to collections, though some collections are maintained in a clean-plant state as a result of intensive sanitation efforts. National and regional quarantine regulations often limit the export and import of citrus plants or propagative materials, thus limiting the availability of materials at an international level. Resources, both financial and human, are necessary to ensure the long-term safety and security of citrus collections on a global scale. Future efforts to develop citrus genebanking communities will provide opportunities for improved conservation, as well as collaborations and training.

The objective of this study was to evaluate the effect of pollen source on the nutritional composition of seeds of the ‘Medjool’ date (Phoenix dactylifera L.) under conditions of northwest Mexico. The content of nutrients (protein, lipids, fiber, and ash), sugars (total, reducing, and non-reducing), and most abundant minerals, were evaluated, as well as antioxidant activities. The pollen of four cultivars (‘Deglet Noor’, ‘Khadrawy’, ‘Medjool’, and ‘Zahidi’) was selected. Four groups of three bunches were pollinated in each of the female palms. For the four treatments, the following ranges were obtained: Seeds had a weight of 1.32–1.41 g and low concentrations of protein (4.44–5.49 g/100 g) and lipids (9.62–10.19 g/100 g). The content of dietary fiber and ash was 59.65–72.59 g/100 g and 1.09–1.42 g/100 g, respectively. Total sugars content was 4.50–6.96 g/100 g, while the insoluble solids were 92.21–96.03 g/100 g. The predominant mineral was potassium with 399.97–459.79 mg/100 g dry weight, followed by sulfur with 135.51–162.67 mg/100 g dry weight. Finally, for antioxidant activities, DPPH (2,2′-diphenyl-1-picrylhydrazyl) and ABTS (2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)) assay results were 0.0014–0.0094 IC50 g/L and 0.176–0.299 IC50 g/L, respectively. The pollen sources had an effect on the physicochemical properties of the seeds of the ‘Medjool’ cultivar date. However, some of the parameters (diameter, total acidity, pH) did not show statistically significant differences. The pollen sources from the ‘Khadrawy’ and ‘Medjool’ cultivars were the ones that induced differences in physical, proximate, and nutritional parameters, while pollen from the ‘Zahidi’ source induced the largest differences for minerals. The results suggest that selection of the source of pollen has a significant effect on the nutritional properties of the date seed from the cultivar ‘Medjool’, and that its use could be feasible as a functional food or by-product of the date production.

Huanglongbing (HLB) is a highly destructive citrus disease and is associated with a nonculturable bacterium, 'Candidatus Liberibacter asiaticus'. 'Ca. L. asiaticus' in the United States was first found in Florida in 2005 and is now endemic there. In California, 'Ca. L. asiaticus' was first detected in Hacienda Heights in Los Angeles County in 2012 and has now been detected in multiple urban locations in southern California. Knowledge of 'Ca. L. asiaticus' strain diversity in California is important for HLB management. In this study, genomic diversity among 10 'Ca. L. asiaticus' strains from six California locations were analyzed using a next-generation sequencing (NGS) (Illumina MiSeq and HiSeq) approach. Draft genome sequences of 'Ca. L. asiaticus' strains were assembled. Sequences of the 16S ribosomal RNA gene and nrdB confirmed 'Ca. L. asiaticus' identity. Prophages were detected in all 'Ca. L. asiaticus' strains. The California 'Ca. L. asiaticus' strains formed four prophage typing groups (PTGs): PTG1, with type 1 prophage only (strains from Anaheim, San Gabriel, and Riverside); PTG2, with type 2 prophage only (strains from Hacienda Heights); PTG1-3, with both type 1 and 3 prophages (a strain from Cerritos); and PTG1-2, with both type 1 and type 2 prophages (a strain from La Habra). Analyses of the terL sequence showed that all California 'Ca. L. asiaticus' strains were not introduced from Florida but likely from locations in Asia. Miniature inverted-repeat transposable elements were found in all 'Ca. L. asiaticus' strains, yet, a jumping-out event was detected in the 'Ca. L. asiaticus' strain from Cerritos. Altogether, this study demonstrated that the NGS approach focusing on prophage variation was sensitive and effective in revealing diversity of 'Ca. L. asiaticus' strains in California.

In the original version of this Article, the affiliation of the first author, Maria F. Torres, ‘Department of Biological Sciences, University of Cincinnati, Cincinnati, 45221, OH, USA’ was incorrectly assigned as a present address and should have been listed as a full affiliation. This error has been corrected in both the PDF and HTML versions of the Article.

Citrus paradisi Macfad. ‘Duncan’ was transformed with constructs coding for the wild-type and mutant RNA-dependent RNA polymerase (RdRp) of Citrus tristeza virus (CTV) for exploring replicase-mediated pathogen-derived resistance (RM-PDR). The RdRp gene was amplified from a CTV genome and used to generate the wild-type and 2 mutant RdRp constructs for plant transformation. One mutant had the key amino acids GDD changed to AAA (RdRp-mGDD), and the second mutant had a deletion encompassing the GDD domain (RdRp-DGDD). Etiolated epicotyl segments of Duncan grapefruit (Citrus paradisi Macfad. 'Duncan') were transformed with each of these constructs using the Agrobacterium-mediated transformation method. From 4540 transformed epicotyl segments, 1402 kanamycin-resistant shoots were regenerated. After testing for expression of green fluorescent protein (GFP) and \Beta-glucuronidase (GUS) reporter genes by fluorescence microscopy and histochemical staining, respectively, 146 GUS-positive plants were rooted and 97 surviving plants were established in soil in pots. A total of 70 plants were tested for the presence of the GUS gene and CTV RdRp transgenes by polymerase chain reaction (PCR). A total of 51 GUS and CTV transgene-positive transgenic plants (15 with RdRp, 21 with RdRp-mGDD, and 15 with RdRp-DGDD) were identified.

Author(s): Ramos, C.; Castillo, J.; Fernandez, O.; Rangel, B.; Manjunath, K. L.; Lee, R. F.