Biology subjects

Reverberi, M.

Publications and source records attributed to Reverberi, M..

6 recordsLinked to original sources

bioRxiv2026

A structurally unique effector shared between Verticillium dahliae and Fusarium oxysporum is involved in cotton defoliation and virulence on other hosts

Defoliating (D) strains of the vascular wilt fungus Verticillium dahliae cause severe yield losses in cotton and olive worldwide, yet the genetic basis underlying this pathotype has remained unknown. Here, we combined comparative genomics, functional genetics, structural analysis, and phylogenomics to uncover the molecular determinant of defoliation. We identified a small, D pathotype-specific genomic region encoding two duplicated secreted effector genes. Simultaneous deletion of both gene copies abolished pathogenicity and defoliation in cotton, olive, Nicotiana benthamiana, and Arabidopsis thaliana, whereas single deletions reduced virulence and genetic complementation restored disease symptoms. Conversely, expression of the D effector in non-defoliating strains was sufficient to induce defoliation. Moreover, or exogenous application of the purified protein, induced wilting and defoliation as well. Structural analyses revealed that D homologs share a conserved but previously uncharacterized protein fold and are distributed across Verticillium and Fusarium species, exhibiting functional diversification and host-specific activity. Phylogenomic and genomic context analyses indicate repeated horizontal transfer events mediated by giant transposable elements known as Starships. Together, our findings identify the D effector as a central pathogenicity factor that drives defoliation and virulence, and demonstrate how Starship-mediated horizontal gene transfer shapes the emergence and dissemination of an agriculturally devastating fungal trait.

plant biology↗

bioRxiv2022

ANTARCTIC FUNGI: A BIO-SOURCE ALTERNATIVE TO PRODUCE POLYUNSATURATED FATTY ACIDS (PUFAs)

The Antarctic ecosystem is a combination of conditions including extremely low values of temperature. The environmental temperature is one of the parameters thoroughly affecting the structure and composition of fungal membranes lipids. The psychrophilic fungi generally increase the disorder within macromolecules to maintain membrane fluidity at low temperatures. The strategy adopted by Antarctic fungi is to increase the proportion of unsaturated fatty acid that allows maintaining a semi-fluid state of the membranes. This ecological feature might be exploited for using Antarctic fungi as potential alternative source of polyunsaturated fatty acids (PUFAs) for human diet. This study provides both the characterization of fungal strains isolated from Antarctica by lipidomic analysis and the laboratory/large-scale production of fungal biomass with high content of beneficial PUFAs. In detail, three fungal species isolated from environmental matrices from Antarctica were tested and identified at genome level. Growth experiments to evaluate the influence of temperature and substrate in the yield in biomass and unsaturated fatty acid (UFA) were conducted. The results showed that the selected fungi have a high percentage of UFA compared to saturated ones; low growth temperatures increase the yield in linolenic fatty acid (C18:3); the biomass yield depends on the composition of the growth substrate and a satisfying qualitative-quantitative yield has also been obtained by using an agri-food chain waste product as growth substrate. IMPORTANCEThe presence of polyunsaturated fatty acids (PUFAs) in human and animal diet is gaining attention because PUFAs have several recognized functional properties: they modulate immune response, have anti-allergic and anti-inflammatory activity, cardio-protective effect and reduce blood LDL cholesterol levels. Human diets typically do not contain sufficient PUFAs because foods rich in PUFAs are few and it is therefore necessary to supplement this diet. Food supplements with these types of fatty acid currently commercially available come from marine fish oils and this source is no longer sustainable. It is necessary to develop efficient industrial processes capable of producing good quality PUFAs and in quantity, even using as carbon and nitrogen sources agro-industrial chains (in our case spent yeast from brewing and whey waste) waste products. Like microorganisms we used Antarctic fungi because they are adapted at very low temperature increasing the proportion of unsaturated fatty acid that allows maintaining a semi-fluid state of the membranes.

ecology↗

bioRxiv2021

Mechanistic Insight from Full Quantum Mechanical Modeling: Laccase as a Detoxifier of Aflatoxins

We demonstrate a path towards full Quantum Mechanics (QM) characterization of enzymatic activity. As a case-study, we investigate the detoxification of aflatoxin, a carcinogenic food contaminant, by laccase, a versatile oxidase capable of--but not efficient for--degrading aflatoxin. We use a combination of quantitative experimentation and QM modeling to show that low enzymatic steric affinity for aflatoxin is the main bottleneck, rather that the oxidative activity of laccase. To identify the structural elements responsible for low reaction rates, we perform a density functional theory (DFT) based modeling of both the substrate and the enzyme in a full QM simulation of more than 7,000 atoms. Thanks to our approach we point to amino acid residues that determine the affinity of laccase for aflatoxin. We show that these residues are substrate-dependent, making a full QM approach necessary for enzyme optimization. Altogether, we establish a roadmap for rational enzyme engineering applicable beyond our case study.

biophysics↗

bioRxiv2020

Full quantum mechanical modeling of the enzyme-substrate system: how laccase detoxifies aflatoxin

This work focuses on: 1) the development of a methodology to perform a full Quantum Mechanics (QM) characterization of enzymatic activity; 2) the development of a rational approach to laccase engineering as a food bioremediator. Aflatoxins are among the most dangerous natural carcinogens, and regularly contaminate reserves of staple crops worldwide. Decontamination of aflatoxin-polluted food is of great interest for ensuring food safety, and bioremediation is regarded as the most promising solution. The fungal isoforms of laccase display the rare potential to detoxify aflatoxin by tackling its aromatic moieties.. Yet, because of a generally low efficiency, large-scale application of naturally occurring isoforms has so far been unfeasible. We perform a combination of quantitative experimentation and quantum mechanical modeling on aflatoxin and reveal that: (1) detoxification efficiency is limited by the low enzymatic affinity for the substrate; and (2) aflatoxin is not detoxified by oxidative activity of laccase alone, but requires additional stimulation from the environment. QM modeling also allowed identification of the residues in the laccase tertiary structure that determine affinity of the enzymatic pocket for aflatoxin. We conclude that, for our case-study, a full QM approach is mandatory as a first step towards rational optimization. We detail a feasible approach towards this endeavor and argue that our full QM characterization can serve as a roadmap for enzyme development in other applications pertaining laccase as well as other enzymes.

biophysics↗

bioRxiv2019

Bacterial and plant produced lipids can exacerbate the Olive Quick Decline Syndrome caused by Xylella

Xylella fastidiosa is an insect vector-transmitted bacterial plant pathogen associated with severe diseases in a wide range of plants. In last decades, X. fastidiosa was detected in several European countries. Among X. fastidiosa subspecies, here we study X. fastidiosa subsp. pauca associated with the Olive Quick Decline Syndrome (OQDS) causing severe losses in Southern Italy. First, we collected Olea europaea L. (cv. Ogliarola salentina) samples in groves located in infected zones and uninfected zones. Secondly, the untargeted LC-TOF analysis of the lipid profiles of OQDS positive (+) and negative (-) plants showed a significant clustering of OQDS+ samples apart from OQDS-ones. Thirdly, using HPLC-MS/MS targeted methods and chemometric analysis, we identified a shortlist of 10 lipids significantly different in the infected versus healthy samples. Last, we observed a clear impact on X. fastidiosa subsp. pauca growth and biofilm formation in vitro liquid cultures supplemented with these compounds. Considering that growth and biofilm formation are primary ways by which X. fastidiosa causes disease, our results demonstrate that lipids produced as part of the plants immune response can exacerbate the disease. This is reminiscent of an allergic reaction in animal systems, offering the depression of plant immune response as a potential strategy for OQDS treatment. Author summaryGlobal trade and climate change are re-shaping the distribution map of pandemic pathogens. One major emerging concern is Xylella fastidiosa, a tropical bacterium recently introduced into Europe from America. Its impact has been dramatic: in the last 5-years only, Olive Quick Decline Syndrome (OQDS) has caused thousands of 200 years old olive trees to be felled in the southern Italy. Xylella fastidiosa through a tight coordination of the adherent biofilm and the planktonic states, invades the host systemically. The planktonic phase is correlated to low cell density and vessel colonization. Increase in cell density triggers a quorum sensing system based on cis 2-enoic fatty acids--diffusible signalling factors (DSF) that promote stickiness and biofilm. Xylem vessels are occluded by the combined effect of bacterial biofilm and plant defences (e.g. tyloses). This study provides novel insight on how X. fastidiosa subsp. pauca biology relates to the Olive Quick Decline Syndrome. We found that some class of lipids increase their amount in the infected olive tree. These lipid entities, provided to X. fastidiosa subsp. pauca behave as hormone-like molecules: modulating the dual phase, e.g. planktonic versus biofilm. Probably, part of these lipids represents a reaction of the plant to the bacterial contamination.

microbiology↗

bioRxiv2019

Nanopore sequencing for the detection and the identification of Xylella fastidiosa subspecies and sequence types from naturally infected plant material

Xylella fastidiosa (Xf) is a polyphagous gram-negative bacterial plant pathogen that can infect more than 300 plant species. It is endemic in America while, in 2013, Xf subsp. pauca was for the first time reported in Europe on olive tree in the Southern Italy. The availability of fast and reliable diagnostic tools is indispensable for managing current and future outbreaks of Xf. In this work, we used the Oxford Nanopore Technologies (ONT) device MinION platform for detecting and identifying Xf at species, subspecies and Sequence Type (ST) level straight from infected plant material. The study showed the possibility to detect Xf by direct DNA sequencing and identify the subspecies in highly infected samples. In order to improve sensitivity, Nanopore amplicon sequencing was assessed. Using primers within the set of the seven MLST officially adopted for identifying Xf at type strain level, we developed a workflow consisting in a multiple PCR and an ad hoc pipeline to generate MLST consensus after Nanopore-sequencing of the amplicons. The here-developed combined approach achieved a sensitivity higher than real-time PCR allowing within few hours, the detection and identification of Xf at ST level in infected plant material, also at low level of contamination. Originality Significance StatementIn this work we developed a methodology that allows the detection and identification of Xylella fastidiosa in plant using the Nanopore technology portable device MinION. The approach that we develop resulted more sensitive than methods currently used for detecting X. fastidiosa, like real-time PCR. This approach can be extensively used for X. fastidiosa detection and it may pave the road for the detection of other tedious vascular pathogens.

pathology↗