Cristiano Lacorte

The plant cytoskeleton undergoes extensive remodelling during the formation of Meloidogyne incognita ‐induced giant cells and plays a crucial role in their ontogenesis. Fimbrins (FIMs) are a conserved family of actin‐bundling proteins that regulate cytoskeletal architecture and dynamics during diverse cellular processes. However, their functional contribution to giant cell development remains unknown. Here, we show that all five Arabidopsis thaliana FIM genes ( FIM1 ‐ FIM5 ) are expressed in M. incognita ‐induced galls, with expression levels declining as giant cells mature. Loss of function of FIM2 or FIM3 , as well as FIM2 overexpression, altered actin organisation and giant cell morphology, resulting in reduced cytoplasmic density and cell wall stubs. These defects were associated with changes in cell wall composition and vacuolar organisation. FIM2 overexpression increased the accumulation of low‐methyl‐esterified homogalacturonan in giant cell walls, whereas fim2 and fim3 mutants showed reduced accumulation, consistent with altered cell wall thickness. Increased vacuolar sizes in INT1‐eGFP Arabidopsis galls following latrunculin B treatment further indicates that actin organisation is required to maintain vacuolar architecture in giant cells. Moreover, fim2 , fim3 and FIM2 ^OE lines showed enhanced resistance to M. incognita , with reduced gall formation and egg mass production. Together, our findings identify FIM2 and FIM3 as critical regulators of actin organisation, cell wall remodelling and vacuolar architecture during gall development, highlighting the importance of a balanced cytoskeleton dynamics for successful nematode parasitism.

Cowpea mild mottle virus (CPMMV) is a carlavirus that is transmitted by whiteflies and can also be spread through seeds in cowpea, soybean, and common bean. Seed transmission of CPMMV has been described in various countries; however, it was only recently reported in Brazil as a seed-transmitted virus in soybean. CPMMV has spread widely in bean-growing areas in recent years. Most Brazilian farmers use harvested grains as seed for reseeding, increasing the risk of seedborne infection. In this study, we examine seed transmission of CPMMV in two bean cultivars using RT-PCR in combination with nucleic acid hybridization. The plants evaluated were obtained from seeds harvested in commercial and experimental fields and from seeds collected in transmission experiments using either mechanical or whitefly inoculation. The observed seed transmission was estimated between 10 to 45% for ‘BRS FC 401 RMD’ and 13 to 22% for ‘Pérola’. In addition, evidence of secondary transmission (23.3%) by Bemisia tabaci MEAM1 was observed. These results suggest that CPMMV could be spread in bean fields by sowing infected seeds and that germinated plants could serve as an inoculum for whitefly transmission. This is the first report of seed transmission of CPMMV in common bean in Brazil.

This study aimed to explore the RNA viruses affecting Passiflora species in Brazil. Our results enhance the understanding of the viruses that infect Passiflora plants by identifying and characterizing three previously unrecognized viruses: Passiflora cytorhabdovirus (PFCV), Passiflora nucleorhabdovirus 1 (PaNV1), and Passiflora nucleorhabdovirus 2 (PaNV2). These rhabdoviruses were identified through high-throughput sequencing and validated by reverse transcription-polymerase chain reaction (RT-PCR) in various Passiflora species. PFCV has a genome organization 3′-N-P-P3-P4-M-G-P7-L-5′ and was classified as a novel member of the Gammacytorhabdovirus genus. A particularly noteworthy feature of PFCV is its glycoprotein, as the genomes of other gammarhabdoviruses do not contain this gene. PFCV has a high incidence across multiple locations and was identified in plants from Northeastern, Central, and Southeastern Brazil. PaNV1 with genome structure 3′-N-P-P3-M-G-L-5′ and PaNV2 with genome organization 3′-N-X-P-Y-M-G-L-5′ are new members of the Alphanucleorhabdovirus genus and have a more restricted occurrence. Importantly, all three viruses were found in mixed infections alongside at least one other virus. In situ observations confirmed mixed infections, with PaNV2 particles co-located in tissues with a potyvirus and a carlavirus. Phylogenetic and glycoprotein sequence similarity network analysis provided insights into their evolutionary placement and potential vector associations. These findings expand the known diversity of rhabdoviruses in Passiflora and contribute to the understanding of their evolution and epidemiology.

Aim: The aim of this study was to evaluate the antibacterial and antibiofilm capacity against Enterococcus faecalis of endodontic sealers AH Plus Jet and the calcium silicate-based sealers, Bio-C Sealer and Sealer Plus BC. Methods: All experiments were performed with fresh endodontic sealers without setting. For this, antibacterial agar diffusion test, analysis of antibiofilm activity evaluated by confocal microscopy, assessment of cell viability and pH evaluation were used. Results: In the agar diffusion test, all sealers showed an ability to inhibit bacterial growth. In the antibiofilm analysis by confocal microscopy, all the evaluated sealers showed the ability to reduce the mature bacterial biofilm. A pH analysis was also performed in the period from 1min to 72h; calcium silicate-based sealers presented a more alkaline pH than AH Plus Jet, and Bio-C Sealer presented a pH greater than 11 in all the analyzed periods. In the assessment of cell viability AH Plus Jet demonstrated a lower cell viability after 24h. However, after 72h, all sealers led to a reduction in cell viability when compared to control. Conclusions: Bio-C Sealer and Sealer Plus BC and AH Plus Jet showed similar antibiofilm capacity against mature E. faecalis biofilms. Furthermore, all evaluated sealers showed antibacterial capacity in the agar diffusion test, and AH Plus Jet and Bio-C Sealer showed significantly greater inhibition of bacterial growth than Sealer Plus BC. Thus, evaluated sealers demonstrated antibacterial and antibiofilm activity against E. faecalis. Knowledge about the antibiofilm and antibacterial activity of endodontic sealers for E. faecalis is relevant, as it is currently one of the main causes of failure in endodontic treatment.

The comprehension of virus-host interactions has allowed numerous advances in developing biotechnological methodologies for plant genome editions, constituting a promising path for plant genetic engineering. Among these advancements, phage- encoded large serine-integrases have emerged as noteworthy tools to modulate plant metabolic pathways by inserting, excising, or inverting DNA stretches in a reversible and specific way. The present work shows the foundation of the Int-Plex@ (INTegrase PLant EXpression) binary memory switch system, which consists of the application of four distinct orthogonal prophage large serine-integrases (Int) (BxB1, phiC31, Int13, and Int9) as an input trigger mechanism for the inversion or excision of genomic DNA. The memory genetic switch is divided into the excision module and the inversion module. The excision module is activated by BxB1 or phiC31 enzymes (input). In this case, the DNA sequence flanked by its attachment sites is excised from the genome (output). The inversion module is activated by Int9 or Int13 (input). The inverted mgf gene sequence is flipped to its functional coding sequence, and the switch output is mGFP. Moreover, prokaryotic-based cell-free in vitro transcription-translation reactions (TxTl) were used as a fast platform for testing Ints attB/P in tandem site activity. Furthermore different plasmid delivery strategies for plant cell Int heterologous expression were tested: leaf tissue agroinfiltration of Agrobacterium tumefaciens transformed with binary plasmids and a biolistic system. After each treatment, the edited genomic DNA sequences were amplified and verified by Sanger and Nanopore sequencing. Despite the challenges of using Ints, the potential benefits are significant and deserve deeper exploration and development. The Int-Plex@ binary genome memory switch system can be applied to produce genetic circuits combined with omics tools and sgRNAs to engineer and modulate plant metabolic pathways temporally and reversibly.