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Comparative cytogenetic mapping is a powerful approach to gain insights into genome organization of orphan crops, lacking a whole sequenced genome. To investigate the cytogenomic evolution of important Vigna and Phaseolus beans, we built a BAC-FISH (fluorescent in situ hybridization of bacterial artificial chromosome) map of Vigna aconitifolia (Vac, subgenus Ceratotropis), species with no sequenced genome, and compared with V. unguiculata (Vu, subgenus Vigna) and Phaseolus vulgaris (Pv) maps. Seventeen Pv BACs, eight Vu BACs, and 5S and 35S rDNA probes were hybridized in situ on the 11 Vac chromosome pairs. Five Vac chromosomes (Vac6, Vac7, Vac9, Vac10, and Vac11) showed conserved macrosynteny and collinearity between V. unguiculata and P. vulgaris. On the other hand, we observed collinearity breaks, identified by pericentric inversions involving Vac2 (Vu2), Vac4 (Vu4), and Vac3 (Pv3). We also detected macrosynteny breaks of translocation type involving chromosomes 1 and 8 of V. aconitifolia and P. vulgaris; 2 and 3 of V. aconitifolia and P. vulgaris; and 1 and 5 of V. aconitifolia and V. unguiculata. Considering our data and previous BAC-FISH studies, six chromosomes (1, 2, 3, 4, 5, and 8) are involved in major karyotype divergences between genera and five (1, 2, 3, 4, and 5) between Vigna subgenera, including mechanisms such as duplications, inversions, and translocations. Macrosynteny breaks between Vigna and Phaseolus suggest that the major chromosomal rearrangements have occurred within the Vigna clade. Our cytogenomic comparisons bring new light on the degree of shared macrosynteny and mechanisms of karyotype diversification during Vigna and Phaseolus evolution.

The genus Vigna Savi (Leguminosae Juss.) comprises approximately 150 species, classified into five subgenera, most of which exhibit a diploid chromosome number of 2n = 22. However, the wild species Vigna lasiocarpa (Benth) Verdc. (V. subg. Lasiospron) is notable for its dysploid chromosome number of 2n = 20. This study aimed to elucidate the chromosomal events involved in the karyotype evolution of V. lasiocarpa (Vla). We used oligopainting probes from chromosomes 1, 2, 3, and 5 of Phaseolus vulgaris L. and two barcode probes from the genome of V. unguiculata (L.) Walp. Additionally, bacterial artificial chromosomes (BACs) from V. unguiculata and P. vulgaris, along with a telomeric probe from Arabidopsis thaliana (L.) Heynh., were hybridized to V. lasiocarpa metaphase chromosomes to characterize Vla3, Vla7/5, and Vla9. Our findings revealed conserved oligo-FISH patterns on chromosomes 2, 6, 8, 10, and 11 between V. unguiculata and V. lasiocarpa. Paracentric and pericentric inversions were identified for Vla3 and Vla9, respectively. Our integrative approach revealed that the dysploid chromosome originated from an “end-to-end fusion” of homoeologous chromosomes 5 and 7. This is the first report on the chromosomal mechanisms underlying descending dysploidy in Vigna, providing new insights into the evolutionary dynamics of the genus.

The genus Vigna (Leguminosae) comprises about 150 species grouped into five subgenera. The present study aimed to improve the understanding of karyotype diversity and evolution in Vigna, using new and previously published data through different cytogenetic and DNA content approaches. In the Vigna subgenera, we observed a random distribution of rDNA patterns. The 35S rDNA varied in position, from terminal to proximal, and in number, ranging from one (V. aconitifolia, V. subg. Ceratotropis) to seven pairs (V. unguiculata subsp. unguiculata, V. subg. Vigna). On the other hand, the number of 5S rDNA was conserved (one or two pairs), except for V. radiata (V. subg. Ceratotropis), which had three pairs. Genome size was relatively conserved within the genus, ranging from 1C = 0.43 to 0.70 pg in V. oblongifolia and V. unguiculata subsp. unguiculata, respectively, both belonging to V. subg. Vigna. However, we observed a positive correlation between DNA content and the number of 35S rDNA sites. In addition, data from chromosome-specific BAC-FISH suggest that the ancestral 35S rDNA locus is conserved on chromosome 6 within Vigna. Considering the rapid diversification in the number and position of rDNA sites, such conservation is surprising and suggests that additional sites may have spread out from this ancestral locus.

Cytogenomic resources have accelerated synteny and chromosome evolution studies in plant species, including legumes. Here, we established the first cytogenetic map of V. angularis (Va, subgenus Ceratotropis) and compared this new map with those of V. unguiculata (Vu, subgenus Vigna) and P. vulgaris (Pv) by BAC-FISH and oligopainting approaches. We mapped 19 Vu BACs and 35S rDNA probes to the 11 chromosome pairs of Va, Vu, and Pv. Vigna angularis shared a high degree of macrosynteny with Vu and Pv, with five conserved syntenic chromosomes. Additionally, we developed two oligo probes (Pv2 and Pv3) used to paint Vigna orthologous chromosomes. We confirmed two reciprocal translocations (chromosomes 2 and 3 and 1 and 8) that have occurred after the Vigna and Phaseolus divergence (~9.7 Mya). Besides, two inversions (2 and 4) and one translocation (1 and 5) have occurred after Vigna and Ceratotropis subgenera separation (~3.6 Mya). We also observed distinct oligopainting patterns for chromosomes 2 and 3 of Vigna species. Both Vigna species shared similar major rearrangements compared to Pv: one translocation (2 and 3) and one inversion (chromosome 3). The sequence synteny identified additional inversions and/or intrachromosomal translocations involving pericentromeric regions of both orthologous chromosomes. We propose chromosomes 2 and 3 as hotspots for chromosomal rearrangements and de novo centromere formation within and between Vigna and Phaseolus. Our BAC- and oligo-FISH mapping contributed to physically trace the chromosome evolution of Vigna and Phaseolus and its application in further studies of both genera.

Gums and carrageenans are food additives widely used in food preparations to improve texture and as viscosifiers. Although they are typically added in small amounts, nowadays people tend to use more and more pre-prepared food. In this work, the content of a wide panel of trace elements in commercial products were analyzed. Carrageenans and gums (n = 13) were purchased in the Portuguese market and were from European suppliers. Samples were solubilized by closed-vessel microwave-assisted acid digestion and analyzed by ICP-MS. Globally, the content of essential trace elements decreased in the following order: Fe (on average, on the order of several tens of µg/g) > Mn > Zn > Cr > Cu > Co > Se > Mo (typically < 0.1 µg/g), while the content of non-essential/toxic trace elements decreased in the following order: Al > Sr > Rb > As > Li > Cd > Pb > Hg. The consumption of these food additives can significantly contribute to the daily requirements of some essential trace elements, namely Cr and Mo. The toxic trace elements Cd, As, Pb, and Hg were below the EU regulatory limits in all analyzed samples. Additional research is needed to define the potential risk of introducing toxic trace elements into food products through the use of these additives.

Mutations in granulin (GRN) have been associated with neurodegenerative diseases, such as frontotemporal lobar degeneration (FTLD) and neuronal ceroid lipofuscinosis (NCL). In Portugal, GRN mutations account for around half of all FTLD cases with known genetic origin. Here, we describe the generation and characterization of three human-induced pluripotent stem cell (hiPSC) lines from a Portuguese family harboring heterozygous and homozygous GRN mutation. hiPSCs were reprogrammed from human dermal fibroblasts by episomal nucleofection of the Yamanaka factors. The new generated lines were positive for pluripotency markers, could be further differentiated to cells expressing all trilineage markers, and presented a normal karyotype. They were also capable of differentiating into 3D brain organoids and presented a significant decrease in progranulin protein levels. Hence, these cell lines constitute suitable new tools to elucidate the pathophysiological mechanisms associated with the GRN mutations in the context of FTLD.

Linnaeus, 1753 (Euphorbiaceae) is one of the most diverse and complex genera among the angiosperms, showing a huge diversity in morphologic traits and ecologic patterns. In order to improve the knowledge of the karyotype organization of (2n = 18) and (2n = 12), cytogenetic studies were performed by means of conventional staining with Giemsa, genome size estimations with flow cytometry, heterochromatin differentiation with chromomycin A (CMA) and 4',6-diamidino-2-phenylindole (DAPI) and Giemsa C-banding, fluorescent hybridization (FISH) with 45S and 5S rDNA probes, and impregnation with silver nitrate (AgNO ). Our results revealed small metacentric chromosomes, CMA /DAPI heterochromatin in the pericentromeric regions of all chromosomes and CMA /DAPI in the distal part of chromosome arms carriers of nucleolar organizing regions (NORs). The DNA content measurements revealed small genomes for both species: with 2C = 0.77 pg and with 2C = 1.41 pg. After FISH procedures, , and presented three and four pairs of terminal 45S rDNA sites, respectively, colocalizing with CMA heterochromatic blocks, besides only one interstitial pair of 5S rDNA signals. Additionally, the maximum number of active NORs agreed with the total number of observed 45S rDNA sites. This work represents the first analysis using FISH in the subfamily Euphorbioideae, revealing a significant number of chromosomal markers, which may be very helpful to understand evolutionary patterns among species.

Objectives To compare the regenerative properties of human stem cells of the apical papilla (SCAPs) embedded in a platelet-rich plasma (PRP) scaffold, when implanted in vivo using an organotypic model composed of human root segments, with or without the presence of the bioactive cements – ProRoot MTA or Biodentine. Material and methods SCAPs were isolated from third molars with incomplete rhizogenesis and expanded and characterized in vitro using stem cell and surface markers. The pluripotency of these cells was also assessed using adipogenic, chondrogenic, and osteogenic differentiation protocols. SCAPs together with a scaffold of PRP were added to the root segment lumen and the organotypic model implanted on the dorsal region of immunodeficient rats for a period of 4 months. Results Presence of SCAPs induced de novo formation of dentin-like and pulp-like tissue. A barrier of either ProRoot MTA or Biodentine did not significantly affect the fraction of sections from roots segments observed to contain deposition of hard material ( P > 0.05). However, the area of newly deposited dentin was significantly greater in segments containing a barrier of Biodentine compared with ProRoot MTA ( P < 0.001). Conclusions and clinical relevance SCAPs offer a viable alternative to other dental stem cells (DSCs) in their regenerative properties when enclosed in the microenvironment of human tooth roots. The present study also suggests that the presence of bioactive materials does not hinder or impede the formation of new hard tissues, but the presence of Biodentine may promote greater mineralized tissue deposition.

Key message Inversions and translocations are the major chromosomal rearrangements involved in Vigna subgenera evolution, being Vigna vexillata the most divergent species. Centromeric repositioning seems to be frequent within the genus. Oligonucleotide-based fluorescence in situ hybridization (Oligo-FISH) provides a powerful chromosome identification system for inferring plant chromosomal evolution. Aiming to understand macrosynteny, chromosomal diversity, and the evolution of bean species from five Vigna subgenera, we constructed cytogenetic maps for eight taxa using oligo-FISH-based chromosome identification. We used oligopainting probes from chromosomes 2 and 3 of Phaseolus vulgaris L. and two barcode probes designed from V. unguiculata (L.) Walp. genome. Additionally, we analyzed genomic blocks among the Ancestral Phaseoleae Karyotype (APK), two V. unguiculata subspecies ( V. subg. Vigna ), and V. angularis (Willd.) Ohwi & Ohashi ( V. subg. Ceratotropis ). We observed macrosynteny for chromosomes 2, 3, 4, 6, 7, 8, 9, and 10 in all investigated taxa except for V. vexillata (L.) A. Rich ( V. subg. Plectrotropis ), in which only chromosomes 4, 7, and 9 were unambiguously identified. Collinearity breaks involved with chromosomes 2 and 3 were revealed. We identified minor differences in the painting pattern among the subgenera, in addition to multiple intra- and interblock inversions and intrachromosomal translocations. Other rearrangements included a pericentric inversion in chromosome 4 ( V. subg. Vigna ), a reciprocal translocation between chromosomes 1 and 5 ( V. subg. Ceratotropis ), a potential deletion in chromosome 11 of V. radiata (L.) Wilczek, as well as multiple intrablock inversions and centromere repositioning via genomic blocks. Our study allowed the visualization of karyotypic patterns in each subgenus, revealing important information for understanding intrageneric karyotypic evolution, and suggesting V. vexillata as the most karyotypically divergent species. Graphical abstract

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