Genomic analysis of a spontaneous unifoliate mutant reveals gene candidates associated with compound leaf development in Vigna unguiculata [L] Walp.

Molecular mechanisms which underpin compound leaf development in some legumes have been reported, but there is no previous study on the molecular genetic control of compound leaf formation in Vigna unguiculata (cowpea), an important dryland legume of African origin. In most studied species with compound leaves, class 1 KNOTTED-LIKE HOMEOBOX genes expressed in developing leaf primordia sustain morphogenetic activity, allowing leaf dissection and the development of leaflets. Other genes, such as, SINGLE LEAFLET1 in Medicago truncatula and Trifoliate in Solanum lycopersicum, are also implicated in regulating compound leaf patterning. To set the pace for an in-depth understanding of the genetics of compound leaf development in cowpea, we applied RNA-seq and whole genome shotgun sequence datasets of a spontaneous cowpea unifoliate mutant and its trifoliate wild-type cultivar to conduct comparative reference-based gene expression, de novo genome-wide isoform switch, and genome variant analyses between the two genotypes. Our results suggest that genomic variants upstream of LATE ELONGATED HYPOCOTYL and down-stream of REVEILLE4, BRASSINOSTERIOD INSENSITIVE1 and LATERAL ORGAN BOUNDARIES result in down-regulation of key components of cowpea circadian rhythm central oscillator and brassinosteroid signaling, resulting in unifoliate leaves and brassinosteroid-deficient-like phenotypes. We have stated hypotheses that will guide follow-up studies expected to provide more insights.

Chemical emasculation in cowpea (Vigna unguiculata (L.) Walp.) and dicotyledonous model species using trifluoromethanesulfonamide (TFMSA).

Hybridization plays an indispensable role in creating the diversity associated with plant evolution and genetic improvement of crops. Production of hybrids requires control of pollination and avoidance of self-pollination for species that are predominantly autogamous. Hand emasculation, male sterility genes or male gametocides have been used in several plant species to induce pollen sterility. However, in cowpea (Vigna unguiculata (L.) Walp), a self-pollinated cleistogamous dryland crop, only hand emasculation is used, but it is tedious and time-consuming. In this study, male sterility was effectively induced in cowpea and two dicotyledonous model species (Arabidopsis thaliana (L.) Heynh. and Nicotiana benthamiana Domin) using trifluoromethanesulfonamide (TFMSA). Pollen viability assays using Alexander staining showed that 30 ml of 1000 mg/l TFMSA with two-time treatments of one-week interval at the early stage of the reproductive phase under field or greenhouse conditions induced 99% pollen sterility in cowpea. TFMSA treatment induced non-functional pollen in diploid A. thaliana at two-time treatment of 10 ml of 125-250 mg/l per plant and N. benthamiana at two-time treatment of 10 ml of 250-1000 mg/l per plant. TFMSA-treated cowpea plants produced hybrid seeds when used as the female parent in crosses with non-treated plants used as male parents, suggesting that TFMSA had no effect on female functionality in cowpea. The ease of TFMSA treatment and its effectiveness to induce pollen sterility in a wide range of cowpea genotypes, and in the two model plant species tested in this study, may expand the scope of techniques for rapid pollination control in self-pollinated species, with potential applications in plant breeding and plant reproduction science.

Exploiting the miniature inverted-repeat transposable elements insertion polymorphisms as an efficient DNA marker system for genome analysis and evolutionary studies in wheat and related species.

Transposable elements (TEs) constitute ~80% of the complex bread wheat genome and contribute significantly to wheat evolution and environmental adaptation. We studied 52 TE insertion polymorphism markers to ascertain their efficiency as a robust DNA marker system for genetic studies in wheat and related species. Significant variation was found in miniature inverted-repeat transposable element (MITE) insertions in relation to ploidy with the highest number of "full site" insertions occurring in the hexaploids (32.6 ± 3.8), while the tetraploid and diploid progenitors had 22.3 ± 0.6 and 15.0 ± 3.5 "full sites," respectively, which suggested a recent rapid activation of these transposons after the formation of wheat. Constructed phylogenetic trees were consistent with the evolutionary history of these species which clustered mainly according to ploidy and genome types (SS, AA, DD, AABB, and AABBDD). The synthetic hexaploids sub-clustered near the tetraploid species from which they were re-synthesized. Preliminary genotyping in 104 recombinant inbred lines (RILs) showed predominantly 1:1 segregation for simplex markers, with four of these markers already integrated into our current DArT-and SNP-based linkage map. The MITE insertions also showed stability with no single excision observed. The MITE insertion site polymorphisms uncovered in this study are very promising as high-potential evolutionary markers for genomic studies in wheat.

Phenotypic and molecular screenings for determination of cassava mosaic disease (CMD) status in farmers' fields in Ebonyi State, Nigeria.

African cassava mosaic virus (ACMV) and East African cassava mosaic virus (EACMV) are among the major constraints to cassava productivity within tropical and sub-tropical regions, including Ebonyi State, Nigeria. Thus, virus indexing has become imperative to determine the status of cassava mosaic disease (CMD) in Ebonyi State, to implement appropriate preventive and control strategies. Seventy-eight cassava accessions obtained from different locations of Ebonyi State were phenotypically scored, using scales 1-5 depending on CMD symptomatic expressions, followed by multiplex-PCR and sequencing for validation. 11% of cassava accessions cultivated in Afikpo were resistant (RE) to ACMV compared to 8% of moderately-resistant (MR) accessions in Izzi and 55% of tolerant (TO) ACMV accessions in Ebonyi. 100% of cassava accessions in Onicha and 66% in Afikpo South were susceptible (SU) and highly susceptible (HS) to ACMV, respectively. With multiplex-PCR, 97.4% (ACMV) and 2.6% (EACMV) were positive. Dunn's multiple comparison tests of CMD mean incidence demonstrated differences (P < 0.05), except between RE and MR, and TO and MR. More transitions (A/G, C/T) compared to transversions (A/T, G/T), were detected, with nonsynonymous mutations (Leucine/Isoleucine; Valine/Isoleucine; Arginine/Lysine; Methionine/Isoleucine), and good bit-scores (91.13-99.07% identites; e-values of 7.00e-148-0.00e+00). Phylogeny resolved the sequences into five major groups. DNA sequencing validated the detected ACMV and EACMV species. This study revealed variants of ACMV and low adoption of RE and MR cassava accessions in the farmers' fields. The findings will guide in getting disease-free and resistant varieties as planting materials to significantly mitigate the CMD spread in Ebonyi State, Nigeria.

Possible association between ABCC8 C49620T polymorphism and type 2 diabetes in a Nigerian population.

BACKGROUND: The association between ABCC8 gene C49620T polymorphism and type 2 diabetes (T2D) in populations of diverse ethnic backgrounds has been reported. However, such occurrence in an African population is yet to be established. This case-control study involving 73 T2D and 75 non-diabetic (ND) patients investigated the occurrence of this polymorphism among T2D patients in Nigeria and assessed its relationship with body lipids of patients. METHODS: Demographic and clinical characteristics of patients were collected and lipid profile indices including total cholesterol (TC), triglyceride (TG), low density lipoprotein (LDL) and high density lipoprotein (HDL) were assayed. Restriction fragment length polymorphism-PCR (RFLP-PCR) was employed to genotype the ABCC8-C49620T polymorphism using PstI restriction enzyme. RESULTS: This study revealed significantly (p < 0.05) higher prevalence of the T allele of the ABCC8 gene in T2D patients (33.1%) compared to ND patients (28.0%). The mutant TT genotype was also higher than the CC and CT genotypes in T2D patients compared to ND patients but did not show any significant risk (p>0.05) of T2D for the unadjusted codominant, dominant and recessive models. Following age adjustment, the mutant genotypes (CT and TT) showed significant (p<0.05) risk of T2D for all the models with the recessive model presenting the greatest risk of T2D (OR: 2.39, 95% CI: 1.16-4.91, p<0.018). The TT genotype significantly (p<0.05) associated with high level of HDL and reduced levels of TC, TG and LDL in non-diabetic patients but was not associated with any of the demographic and clinical characteristics among T2D patients. CONCLUSIONS: ABCC8 C49620T polymorphism showed possible association with T2D marked by predominance of the mutant TT genotype in T2D patients. However, the relationship between TT genotype and lipid abnormalities for possible beneficial effect on people suffering from T2D is unclear.

Assessment of the Pro12Ala Polymorphism in the PPAR-γ2 Gene among Type 2 Diabetes Patients in a Nigerian Population.

The association between the Pro12Ala polymorphism of the PPARγ2 gene, type 2 diabetes (T2D), and obesity in certain ethnic populations has been reported. However, this relationship has not yet been described among diabetes patients in Nigeria. This study investigated the relationship between the Pro12Ala polymorphism in the PPARγ2 gene, obesity, and lipid abnormalities characterizing T2D among patients in Nigeria. This case-control study recruited 73 T2D and 75 non-diabetic (ND) patients. Demographic and clinical data were collected and blood glucose levels together with serum lipid profile for patients were measured. Pro12Ala polymorphism in the PPARγ2 gene was genotyped by restriction fragment length-Polymerase Chain Reaction (RFLP-PCR). The PPAR-γ2 gene (amplicon size = 270 base pair) was successfully amplified for all samples. Following restriction enzyme digestion and analysis by agarose gel electrophoresis, amplicons from samples showed a band of size 270 bp and were of the wild homozygous Pro/Pro genotype. Ala12 variant was totally absent from the study population. Obesity, estimated using Body Mass Index (BMI) and waist circumference (WC), was significantly higher (p < 0.05) in T2D patients compared to the non-diabetic patients. More so, the prevalence of lipid abnormalities; hypercholesterolaemia (TC > 200 mg/dL), hypertriglyceridaemia (TG > 150 mg/dL), high HDL (>100 mg/dL), and low HDL (<50 mg/dL) was significantly greater (p < 0.001) in T2D patients compared to non-diabetic patients. Results obtained further indicated lack of significant association between PPAR-γ2 gene polymorphism, T2D, and obesity. However, obesity and dyslipidaemia were strongly associated in T2D patients.

Assessment of genetic diversity in Vigna unguiculata L. (Walp) accessions using inter-simple sequence repeat (ISSR) and start codon targeted (SCoT) polymorphic markers.

BACKGROUND: Assessment of genetic diversity of Vigna unguiculata (L.) Walp (cowpea) accessions using informative molecular markers is imperative for their genetic improvement and conservation. Use of efficacious molecular markers to obtain the required knowledge of the genetic diversity within the local and regional germplasm collections can enhance the overall effectiveness of cowpea improvement programs, hence, the comparative assessment of Inter-simple sequence repeat (ISSR) and Start codon targeted (SCoT) markers in genetic diversity of V. unguiculata accessions from different regions in Nigeria. Comparative analysis of the genetic diversity of eighteen accessions from different locations in Nigeria was investigated using ISSR and SCoT markers. DNA extraction was done using Zymogen Kit according to its manufacturer's instructions followed by amplifications with ISSR and SCoT and agarose gel electrophoresis. The reproducible bands were scored for analyses of dendrograms, principal component analysis, genetic diversity, allele frequency, polymorphic information content, and population structure. RESULTS: Both ISSR and SCoT markers resolved the accessions into five major clusters based on dendrogram and principal component analyses. Alleles of 32 and 52 were obtained with ISSR and SCoT, respectively. Numbers of alleles, gene diversity and polymorphic information content detected with ISSR were 9.4000, 0.7358 and 0.7192, while SCoT yielded 11.1667, 0.8158 and 0.8009, respectively. Polymorphic loci were 70 and 80 in ISSR and SCoT, respectively. Both markers produced high polymorphism (94.44-100%). The ranges of effective number of alleles (Ne) were 1.2887 ± 0.1797-1.7831 ± 0.2944 and 1.7416 ± 0.0776-1.9181 ± 0.2426 in ISSR and SCoT, respectively. The Nei's genetic diversity (H) ranged from 0.2112 ± 0.0600-0.4335 ± 0.1371 and 0.4111 ± 0.0226-0.4778 ± 0.1168 in ISSR and SCoT, respectively. Shannon's information index (I) from ISSR and SCoT were 0.3583 ± 0.0639-0.6237 ± 0.1759 and 0.5911 ± 0.0233-0.6706 ± 0.1604. Total gene diversity (Ht), gene diversity within population (Hs), coefficient of gene differentiation (Gst) and level of gene flow (Nm) revealed by ISSR were 0.4498, 0.3203, 0.2878 and 1.2371 respectively, while SCoT had 0.4808, 0.4522, 0.0594 and 7.9245. CONCLUSIONS: Both markers showed highest genetic diversity in accessions from Ebonyi. Our study demonstrated that SCoT markers were more efficient than ISSR for genetic diversity studies in V. unguiculata and can be integrated in the exploration of their genetic diversity for improvement and germplasm utilization.

Repression of TERMINAL FLOWER1 primarily mediates floral induction in pear (Pyrus pyrifolia Nakai) concomitant with change in gene expression of plant hormone-related genes and transcription factors.

Floral induction is an important event in the annual growth cycle of perennial fruit trees. For pear, this event directly affects fruit production in the following year. The flower buds in many species are induced by FLOWERING LOCUS T (FT), whose effect is repressed by the meristem-expressed gene TERMINAL FLOWER1 (TFL1). In this study, we investigated the functions of pear FT and TFL1 genes during floral development. Expression of pear FTs (PpFT1a and PpFT2a) in reproductive meristems was not obviously induced prior to floral initiation, while expression of TFL1s (PpTFL1-1a and PpTFL1-2a) rapidly decreased. The induction of the productive meristem identity MADS-box gene AP1 after repression of PpTFL1s suggested a primary role for PpTFL1 in floral induction. RNA-seq analysis suggested that plant hormone-related genes and several transcription factors that were coexpressed with PpTFL1 were potentially involved in the PpTFL1-mediated floral induction. Our data indicate the essential function of TFL1 in pear floral induction and add another species in the family Rosaceae in addition to strawberry and rose that shows a role for TFL1 in floral induction.

Macroalgae culture to treat anaerobic digestion piggery effluent (ADPE).

Environmental consequences of high productivity piggeries are significant and can result in negative environmental impacts, hence bioremediation techniques (in particular using macroalgae) are therefore of great interest. Here, the growth potential of several freshwater macroalgae in anaerobic digestion piggery effluent (ADPE), their nutrient removal rates and biochemical composition of the biomass were investigated under outdoor climatic conditions. A consortium of two macroalgae, Rhizoclonium sp. and Ulothrix sp. was isolated and could efficiently grow in the ADPE. Maximum ammonium removal rate (30.6±6.50mg NH4+-NL-1d-1) was achieved at ADPE concentration equivalent to 248mgNH4+-NL-1. Mean biomass productivity of 31.1±1.14g ash-free dry weight (AFDW) m-2d-1 was achieved. Total carbohydrate and protein contents ranged between 42.8-54.8 and 43.4-45.0% AFDW, respectively, while total lipid content was very low. The study indicates the potential use of this macroalgal consortium for treating ADPE as well as source of animal feed production.

Phenolic compounds in Rosaceae fruit and nut crops.

The demand for new fruit cultivars with high levels of phytochemicals, in particular phenolic compounds, has received increasing attention from biochemists, pharmaceutical companies, plant breeders, and the general public due to their health benefits. This review focuses on the economically important Rosaceae, which contains varying proportions and concentrations of these compounds. The paper discusses the common phenolics in the Rosaceae including phenolic acids, flavonols, flavanols, anthocyanins, and dihydrochalcones. The nonextractable phenolics are also presented but not discussed in detail. The metabolism and bioavailability of phenolics, as well as human and environmental factors that affect their concentration and composition, are highlighted. Furthermore, the paper presents different approaches for biofortification and posits that breeding may be the most viable and sustainable option as it improves other fruit quality traits simultaneously and increases confidence in adoption of new cultivars with enhanced consumer appeal.

Characterization of 10 MADS-box genes from Pyrus pyrifolia and their differential expression during fruit development and ripening.

We cloned 10 Japanese pear (Pyrus pyrifolia) MIKC-type II MADS-box genes, and analyzed their expression during fruit development and ripening. PpMADS2-1 was APETALA (AP)1-like; PpMADS3-1 was FRUITFULL (FUL)/SQUAMOSA (SQUA)-like; PpMADS4-1 was AGAMOUS-like (AGL)6; PpMADS5-1 and PpMADS8-1 were SUPPRESSOR OF OVEREXPRESSION OF CONSTANS (SOC)-like; PpMADS9-1, PpMADS12-1, PpMADS14-1 and PpMADS16-1 were SEPALLATA (SEP)-like; while PpMADS15-1 was AGL/SHATTERPROOF (SHP)-like. Phylogenetic analysis showed their grouping into five major clades (and 10 sub-clades) that was consistent with their diverse functional types. Expression analysis in flower tissue revealed their distinct putative homeotic functional classes: A-class (PpMADS2-1, PpMADS3-1, PpMADS4-1, and PpMADS14-1), C-class (PpMADS15-1), E-class (PpMADS9-1, PpMADS12-1, and PpMADS16-1) and E (F)-class (PpMADS5-1 and PpMADS8-1). Differential gene expression was observed in different fruit tissues (skin, cortex and core) as well as in the cortex during the course of fruit development and ripening. Collectively, our results suggest their involvement in the diverse aspects of plant development including flower development and the course of fruit development and ripening.

UDP-sugar biosynthetic pathway: contribution to cyanidin 3-galactoside biosynthesis in apple skin.

UDP-galactose:flavonoid 3-O-galactosyltransferase (UFGalT) is responsible for cyanidin 3-galactoside (cy3-gal) synthesis from cyanidin (cy) and UDP-galactose (UDP-gal) which are, respectively, catalyzed by anthocyanidin synthase (ANS) and UDP-glucose 4-epimerase (UGE). To clarify the contribution of UDP-galactose pathway to cy3-gal accumulation in apple skin, we analyzed the contents of UDP-gal and UDP-glucose (UDP-glu), cy, and, cy3-gal contents along with UGE activity. We confirmed that transcript levels for apple ANS and UDP-glucose: flavonoid 3-O-glucosyltransferase (UFGT) coincided with anthocyanin accumulation in three apple cultivars differing in their skin colors. During fruit development, changes in level of cy coincided with that of cy3-gal, whereas UDP-gal and UGE activity showed no similar trend with cy3-gal. Significant correlation was not observed between the changes in UGE activity and UDP-sugar contents. The effect of temperature and UV-B radiation (different environmental conditions) on the accumulation of UDP-sugars, cy and cy3-gal, and UGE activity were also investigated in a pale-red cultivar. High temperature tended to depress the accumulation of both UDP-sugars and cy concomitant with the decrease in cy3-gal content irrespective of UV-B radiation. Although there was no high inhibition of both cy and UDP-sugars at low-temperature without UV-B, cy3-gal accumulation was highly depressed. UGE activity was highest at low temperature with UV-B, but not much different under other conditions. Most of the parameters under different environmental conditions were significantly correlated with each other. Based on these results, contribution of UDP-sugar biosynthetic pathway to anthocyanin biosynthesis under different environmental conditions as well as during fruit development is discussed.