A cytogenetic map was established for the trifoliate orange, Poncirus trifoliata. Chromosome size, centromere position and CMA + heterochromatin content were determined for each chromosome pair, together with the position of BAC clones previously described as chromosome-specific markers. Although P. trifoliata is a true biological species, heteromorphisms for the size of the heterochromatic bands in two chromosome pairs were observed. A minimum set of four BACs was proposed for chromosome identification in P. trifoliata, as well as a chromosome nomenclature that can be applied to all citrus species. The Ctv locus was assigned to chromosome 9 and the remaining linkage groups can now be integrated to this physical map. This standard nomenclature for trifoliate orange will be valuable for genome-sequencing projects in all citrus species.

Background: West Nile virus (WNV) was first sequenced in Brazil in 2019, when it was isolated from a horse in the Espírito Santo state. Despite multiple studies reporting serological evidence suggestive of past circulation since 2004, WNV remains a low priority for surveillance and public health, such that much is still unknown about its genomic diversity, evolution, and transmission in the country. Methods: A combination of diagnostic assays, nanopore sequencing, phylogenetic inference, and epidemiological modeling are here used to provide a holistic overview of what is known about WNV in Brazil. Results: We report new genetic evidence of WNV circulation in southern (Minas Gerais, São Paulo) and northeastern (Piauí) states isolated from equine red blood cells. A novel, climate-informed theoretical perspective of the potential transmission of WNV across the country highlights the state of Piauí as particularly relevant for WNV epidemiology in Brazil, although it does not reject possible circulation in other states. Conclusion: Our output demonstrates the scarceness of existing data, and that although there is sufficient evidence for the circulation and persistence of the virus, much is still unknown on its local evolution, epidemiology, and activity. We advocate for a shift to active surveillance, to ensure adequate preparedness for future epidemics with spill-over potential to humans.

Mandarins are among the main citrus species and constitute a morphologically diverse and taxonomically controversial group. Cytogenetic studies were limited to the determination of its karyotype formula and the localization of ribosomal DNA (rDNA) sites. The cytogenetic map of Poncirus trifoliata, established by fluorescent in situ hybridization (FISH) using single-copy bacterial artificial chromosomes (BACs) together with the localization of rDNA sequences and heterochromatin (CMA/DAPI banding pattern), represented an important milestone for the cytogenetic studies of the citrus species. In this work, the cytogenetic map of Citrus reticulata cv. Cravo, one of the pure species of the genus, was built using BACs from P. trifoliata, including the citrus tristeza virus resistance locus (Ctv), to detect homeologies and structural karyotypic changes between these species. Except for chromosomes 2 and 3, heterochromatin distribution and position of BACs were maintained in comparison to P. trifoliata, indicating a high synteny between both karyotypes and supporting a close phylogenetic relationship between these species. A break of colinearity was observed between BAC clones and the CMA + bands (including rDNA loci) from chromosomes 2 and 3, which could be mainly explained by amplification and reduction of the satellite DNA sequences that constitute the heterochromatic bands of these chromosome pairs.

Pummelo (Citrus maxima) is considered as one of the true citrus species. Together with mandarin (C. reticulata), it gave rise to the hybrid sweet orange (C. sinensis) and other important citrus crops. Although these species have 2n = 18, each has a unique heterochromatin distribution. The aims of this study were to identify chromosome homoeologies between pummelo and other true citrus species, to investigate the karyotypic changes involved in the chromosomal evolution between true citrus and to shed light into the origin of sweet orange hybrid karyotype. Mitotic metaphase chromosomes of pummelo and sweet orange were double stained with the fluorochromes CMA/DAPI (Chromomycin A3/4’-6-diamidino-2-phenylindole), and identified by FISH (Fluorescence in Situ Hybridization) with chromosome-specific BAC (Bacterial Artificial Chromosome) markers. The results were compared to previously established cytogenetic maps of mandarin, C. medica and Poncirus trifoliata. Only chromosomes 1, 4 and 8 were maintained unaltered among species, with chromosomes 2 and 3 being among the least conserved in heterochromatin distribution. BACs were conserved in position among homoeologs and the markers mapped to chromosomes 2 and 3 indicated that sweet orange karyotype largely conserved one chromosome from pummelo and one from mandarin. Despite conserved synteny, expansion and contraction of heterochromatic blocks accounted for the differences between karyotypes, even between the hybrid sweet orange and pummelo.

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