Female self-incompatibility type in heterostylous Primula is determined by the brassinosteroid-inactivating cytochrome P450 CYP734A50.

Most flowering plants are hermaphrodites, with flowers having both male and female reproductive organs. One widespread adaptation to limit self-fertilization is self-incompatibility (SI), where self-pollen fails to fertilize ovules.1,2 In homomorphic SI, many morphologically indistinguishable mating types are found, although in heteromorphic SI, the two or three mating types are associated with different floral morphologies.3-6 In heterostylous Primula, a hemizygous supergene determines a short-styled S-morph and a long-styled L-morph, corresponding to two different mating types, and full seed set only results from intermorph crosses.7-9 Style length is controlled by the brassinosteroid (BR)-inactivating cytochrome P450 CYP734A50,10 yet it remains unclear what defines the male and female incompatibility types. Here, we show that CYP734A50 also determines the female incompatibility type. Inactivating CYP734A50 converts short S-morph styles into long styles with the same incompatibility behavior as L-morph styles, and this effect can be mimicked by exogenous BR treatment. In vitro responses of S- and L-morph pollen grains and pollen tubes to increasing BR levels could only partly explain their different in vivo behavior, suggesting both direct and indirect effects of the different BR levels in S- versus L-morph stigmas and styles in controlling pollen performance. This BR-mediated SI provides a novel mechanism for preventing self-fertilization. The joint control of morphology and SI by CYP734A50 has important implications for the evolutionary buildup of the heterostylous syndrome and provides a straightforward explanation for why essentially all of the derived self-compatible homostylous Primula species are long homostyles.11.

Screening of antimicrobial synergism between phenolic acids derivatives and UV-A light radiation.

The objective of this study was to investigate synergistic antibacterial activity based on a combination of UV-A light and three classes of food grade compounds: benzoic acid derivatives, cinnamic acid derivatives, and gallates. By using Escherichia coli O157:H7 as the model strain, it was observed that three cinnamic acid derivatives (ferulic acid, coumaric acid, and caffeic acid) and one benzoic acid derivative (2,5-dihydroxybenzoic acid) presented strong synergistic antibacterial activity with UV-A light radiation, where 1 mM levels of these compounds plus with 15 min of UV-A light (total light dose of 6.1 cm-2) led to more than 7-log CFU mL-1 of bacterial inactivation. In contrast, synergistic antibacterial activity between UV-A light and most benzoic acid derivatives (benzoic acid, gallic acid, vanillic acid, and 2,5-dimethoxybenzoic acid) were only observed after higher concentrations of these compounds were applied (10 mM). Lastly, from the three gallates tested (methyl gallate, ethyl gallate, and propyl gallate), only propyl gallate showed strong antibacterial synergism with UV-A light, where 10 mM of propyl gallate plus 15 min of UV-A light led to approximately 6.5-log of bacterial reduction. Presence of antioxidant compounds mitigated the light-mediated antibacterial activity of gallic acid, 2,5-dihydroxybenzoic acid, and propyl gallate. Similarly, the light-mediated antibacterial activity of these compounds was significantly (P < 0.05) reduced against metabolic-inhibited bacterial cells (sodium azide pretreatment). On the other hand, the antibacterial synergism between ferulic acid and UV-A light was not affected by the presence of antioxidants or the metabolic state of the bacterial cells. Due to the increasing concerns of antimicrobial resistant (AMR) pathogens, the study also investigated the proposed synergistic treatment on AMR Salmonella. Combinations of 1 mM of ferulic acid or 1 mM of 2,5-dihydroxybenzoic acid with UV-A light radiation was able to inactivate more than 6-log of a multi-drug resistant Salmonella Typhimurium strain.

Fairy circles in Namibia are assembled from genetically distinct grasses.

Fairy circles are striking regularly sized and spaced, bare circles surrounded by Stipagrostis grasses that occur over thousands of square kilometres in Namibia. The mechanisms explaining their origin, shape, persistence and regularity remain controversial. One hypothesis for the formation of vegetation rings is based on the centrifugal expansion of a single individual grass plant, via clonal growth and die-back in the centre. Clonality could explain FC origin, shape and long-term persistence as well as their regularity, if one clone competes with adjacent clones. Here, we show that for virtually all tested fairy circles the periphery is not exclusively made up of genetically identical grasses, but these peripheral grasses belong to more than one unrelated genet. These results do not support a clonal explanation for fairy circles. Lack of clonality implies that a biological reason for their origin, shape and regularity must emerge from competition between near neighbor individuals within each fairy circle. Such lack of clonality also suggests a mismatch between longevity of fairy circles versus their constituent plants. Furthermore, our findings of lack of clonality have implications for some models of spatial patterning of fairy circles that are based on self-organization.

Supergene evolution via stepwise duplications and neofunctionalization of a floral-organ identity gene.

Heterostyly represents a fascinating adaptation to promote outbreeding in plants that evolved multiple times independently. While l-morph individuals form flowers with long styles, short anthers, and small pollen grains, S-morph individuals have flowers with short styles, long anthers, and large pollen grains. The difference between the morphs is controlled by an S-locus "supergene" consisting of several distinct genes that determine different traits of the syndrome and are held together, because recombination between them is suppressed. In Primula, the S locus is a roughly 300-kb hemizygous region containing five predicted genes. However, with one exception, their roles remain unclear, as does the evolutionary buildup of the S locus. Here we demonstrate that the MADS-box GLOBOSA2 (GLO2) gene at the S locus determines anther position. In Primula forbesii S-morph plants, GLO2 promotes growth by cell expansion in the fused tube of petals and stamen filaments beneath the anther insertion point; by contrast, neither pollen size nor male incompatibility is affected by GLO2 activity. The paralogue GLO1, from which GLO2 arose by duplication, has maintained the ancestral B-class function in specifying petal and stamen identity, indicating that GLO2 underwent neofunctionalization, likely at the level of the encoded protein. Genetic mapping and phylogenetic analysis indicate that the duplications giving rise to the style-length-determining gene CYP734A50 and to GLO2 occurred sequentially, with the CYP734A50 duplication likely the first. Together these results provide the most detailed insight into the assembly of a plant supergene yet and have important implications for the evolution of heterostyly.

Synergistic Antimicrobial Activity by Light or Thermal Treatment and Lauric Arginate: Membrane Damage and Oxidative Stress.

The need for more effective antimicrobials is critical for the food industry to improve food safety and reduce spoilage of minimally processed foods. The present study was initiated to develop an efficient and novel antimicrobial approach which combines physical treatments (UV-A or mild heat) and generally recognized as safe lauroyl arginate ethyl (LAE) to inactivate surrogate strains, including Escherichia coli and Listeria innocua Synergistic inactivation of bacteria resulted in an ∼6-log reduction of target bacteria, while individual treatments resulted in <1.5-log inactivation under the same set of conditions. In addition, the synergistic mechanism between LAE and UV-A/mild heat was evaluated by supplementing with a variety of antioxidants for suppressing oxidative stress and measurement of cell membrane damage by nucleic acid release. These results demonstrate that the synergistic antimicrobial activity of LAE and mild physical stresses was suppressed by supplementation with antioxidants. The research also compared LAE with another membrane-targeting lipopeptide antimicrobial agent, polymyxin B, to understand the uniqueness of LAE-induced synergy. Briefly, differences in modes of action between LAE and polymyxin B were characterized by comparing the MIC, damage to liposomes, and oxidative stress generation. These differences in the mode of action between LAE and polymyxin B suggested that both compounds target cell membrane but significantly differ in mechanisms, including membrane disruption and oxidative stress generation. Overall, this study illustrates synergistic antimicrobial activity of LAE with light or mild heat and indicates a novel oxidative stress pathway that enhances the activity of LAE beyond membrane damage.IMPORTANCE This study highlights an effective antimicrobial processing approach using a novel combination of lauroyl arginate ethyl (LAE) and two different physical treatments, light (UV-A) and mild heat. Both combinations demonstrated synergistic inactivation against a model Gram-negative bacterium or a Gram-positive bacterium or both by a >5-log reduction. Further mechanistic study revealed that oxidative stress is responsible for synergistic inactivation between LAE and UV-A, while both membrane damage and oxidative stress are responsible for the synergistic combination between LAE and mild heat. The mode of action of LAE was further compared to that of polymyxin B and analyzed using artificial membrane model systems and the addition of antioxidants. The proposed combination of LAE and common physical treatments may improve food preservation, food safety, and current sanitation processes for the food industry and the inactivation of pathogenic strains in biomedical environments.

A short story gets longer: recent insights into the molecular basis of heterostyly.

Heterostyly is a fascinating adaptation to promote outbreeding and a classical paradigm of botany. In the most common type of heterostyly, plants either form flowers with long styles and short stamens, or short styles and long stamens. This reciprocal organ positioning reduces pollen wastage and promotes cross-pollination, thus increasing male fitness. In addition, in many heterostylous species selfing and the generation of unfit progeny due to inbreeding depression is limited by a self-incompatibility system, thus promoting female fitness. The two floral forms are genetically determined by the S locus as a complex supergene, namely a chromosomal region containing several individual genes that control the different traits, such as style or stamen length, and are held together by very tight linkage due to suppressed recombination. Recent molecular-genetic studies in several systems, including Turnera, Fagopyrum, Linum, and Primula have begun to identify and characterize the causal heterostyly genes residing at the S locus. An emerging theme from several families is that the dominant S haplotype represents a hemizygous region not present on the recessive s haplotype. This provides an explanation for the suppressed recombination and suggests a scenario for the chromosomal evolution of the S locus. In this review, we discuss the results from recent molecular-genetic analyses in light of the classical models on the genetics and evolution of heterostyly.

Presence versus absence of CYP734A50 underlies the style-length dimorphism in primroses.

Heterostyly is a wide-spread floral adaptation to promote outbreeding, yet its genetic basis and evolutionary origin remain poorly understood. In Primula (primroses), heterostyly is controlled by the S-locus supergene that determines the reciprocal arrangement of reproductive organs and incompatibility between the two morphs. However, the identities of the component genes remain unknown. Here, we identify the Primula CYP734A50 gene, encoding a putative brassinosteroid-degrading enzyme, as the G locus that determines the style-length dimorphism. CYP734A50 is only present on the short-styled S-morph haplotype, it is specifically expressed in S-morph styles, and its loss or inactivation leads to long styles. The gene arose by a duplication specific to the Primulaceae lineage and shows an accelerated rate of molecular evolution. Thus, our results provide a mechanistic explanation for the Primula style-length dimorphism and begin to shed light on the evolution of the S-locus as a prime model for a complex plant supergene.

The draft genome of Primula veris yields insights into the molecular basis of heterostyly.

BACKGROUND: The flowering plant Primula veris is a common spring blooming perennial that is widely cultivated throughout Europe. This species is an established model system in the study of the genetics, evolution, and ecology of heterostylous floral polymorphisms. Despite the long history of research focused on this and related species, the continued development of this system has been restricted due the absence of genomic and transcriptomic resources. RESULTS: We present here a de novo draft genome assembly of P. veris covering 301.8 Mb, or approximately 63% of the estimated 479.22 Mb genome, with an N50 contig size of 9.5 Kb, an N50 scaffold size of 164 Kb, and containing an estimated 19,507 genes. The results of a RADseq bulk segregant analysis allow for the confident identification of four genome scaffolds that are linked to the P. veris S-locus. RNAseq data from both P. veris and the closely related species P. vulgaris allow for the characterization of 113 candidate heterostyly genes that show significant floral morph-specific differential expression. One candidate gene of particular interest is a duplicated GLOBOSA homolog that may be unique to Primula (PveGLO2), and is completely silenced in L-morph flowers. CONCLUSIONS: The P. veris genome represents the first genome assembled from a heterostylous species, and thus provides an immensely important resource for future studies focused on the evolution and genetic dissection of heterostyly. As the first genome assembled from the Primulaceae, the P. veris genome will also facilitate the expanded application of phylogenomic methods in this diverse family and the eudicots as a whole.