FEMS Microbiology Letters

Currently, there are three recognized rhizobial genera belonging to the beta branch of the proteobacteria; Trinickia, Paraburkholderia, and Cupriavidus. These beta-rhizobia have been found associated with legume species mainly within the Mimosoideae and Papillonoideae. Most diversity, evolutionary, and functional studies have focused on Paraburkholderia, whereas few have addressed the diversity and evolution of symbiosis in the Cupriavidus genus. The present work aimed to provide an actual view of the symbiotic Cupriavidus diversity and to analyse the origin and evolution of their symbiotic genes. Using whole-genome information for phylogenetic reconstruction, we showed that the described symbiotic Cupriavidus strains belong to five distinct lineages, although they are intermixed with non-symbiotic species. The high synteny and sequence conservation of symbiotic genes suggest a common origin of acquisition for all rhizobial Cupriavidus described so far. However, we observed very low sequence conservation among (mega)plasmids carrying the symbiotic island, excluding the existence of a conserved symbiotic plasmid within beta-rhizobia. We can conclude that up to now there are five rhizobial species within the Cupriavidus genus, and we predict the description of new symbiotic species in the near future.

Sugar-induced cell death (SICD) remains an intriguing but poorly studied phenomenon in the physiology of Saccharomyces cerevisiae. Recently, it was shown that SICD development largely depends on the redirection of glucose fluxes between glycolysis and the pentose phosphate pathway (PPP). In particular, inhibition of glycolysis by iodoacetamide (IAA) was observed to reduce SICD levels. This study is devoted to further investigation of the relationship between SICD and the functionality of the two PPP branches. It was shown that deletion of the ZWF1 gene does not affect the decrease in SICD levels in IAA-treated cells. This allows us to conclude that the oxidative branch of the PPP is not involved in the suppression of SICD/ROS. Deletion of the GLR1 gene and attenuation of the TRR1 gene also did not restore SICD levels in cells after IAA treatment. The obtained results indicate that the level of reduced glutathione or thioredoxin does not affect SICD genesis. The addition of 5 mM ribose-5-phosphate (R5P) to the incubation medium led to suppression of SICD by 79%. At the same time, the addition of 5 mM ribose + 5 mM Pi suppressed SICD by only 20%. Suppression of SICD by 5 mM R5P in the{Delta} pho3 strain (83%) excludes the mechanism of extracellular dephosphorylation of R5P to ribose, its subsequent transport into the cell, and re-phosphorylation inside the cell. Furthermore, more than 70% suppression of SICD in the{Delta} end3 strain with 5 mM R5P excludes endocytosis as a mechanism of R5P import into the cell. The observed effect of R5P can be explained by the moonlighting function of some unknown protein. Thus, SICD development in S. cerevisiae cells depends on the final product of the non-oxidative PPP--R5P.

Purine moieties are essential for many functions within the eukaryotic cell, including energy, signaling and nucleic acid synthesis. While purine starvation is known to induce stress resistance in eukaryotic model organism budding yeast Saccharomyces cerevisiae, it remains unclear whether the physiological response is related to disruption of synthesis pathway in particular position or it is uniform across all genetic deficiencies within the de novo adenine biosynthesis pathway. It is also not known how purine starved cells perceive purine shortage - weather they share the same signaling elements with nitrogen starvation or not. MethodsWe characterised physiology of strains with deletions in adenine biosynthesis pathway when cultivated in full or purine deficient and compared to cell physiological parameters when cultivated in nitrogen deficient media. We tested stress tolerance, carbon flux, cell cycle arrest and did transcription profiling (RNA-seq). ResultsOur findings demonstrate that purine starvation-induced stress resistance is significantly modulated by the specific step at which the pathway is interrupted. Transcriptional analysis revealed that purine starvation in many aspects phenocopies nitrogen starvation, particularly - in both starvations strong downregulation of ribosome related genes occurs. In the same time several metabolic features which differ from N- and ade- starvations: pentose phosphate pathway is specifically upregulated within ade4{Delta}-ade2{Delta} and downregulated in N-cells. Notably, the expression of stress-responsive genes such as HSP12, HSP26, and GRE1 varied between mutants, suggesting that the accumulation of pathway intermediates (e.g., AIR in ade2{Delta}) or the absence of downstream precursors (AICAR) alters the perception of starvation especially in the case of ade16{Delta}ade17{Delta} strain. ConclusionsMetabolic and stress-tolerance phenotypes of purine auxotrophs are not merely a result of purine depletion but seems that the response is signalled via the same pathways, like TOR1. The results suggest that strains having mutations within various positions of the purine pathway "perceive" purine limitation a bit differently - especially when we compare the end of the pathway with the other mutants. Different phenotypic outcomes of the occasional purine depletion might give preferences for organisms which have mutations in the beginning rather at the end of the pathway. Besides, our findings might have implications in the design of synthetic pathways and the use of auxotrophic markers in yeast research.

Cyanobacteria utilize type IV pili for many behavioural responses, such as phototaxis, aggregation, floating, and DNA uptake. Type IV pilus-dependent functions are regulated by the nucleotide second messengers, c-di-GMP and cAMP. In this study, we investigated the role of a recently identified c-di-GMP receptor (CdgR) in cyanobacteria that harbours a ComFB domain. ComFB-domain proteins are widespread in cyanobacteria and are also present in heterotrophic bacteria. We demonstrated that the CdgR homolog from the cyanobacterium Synechocystis sp. PCC 6803, a model organism for studying type IV pilus-dependent functions, specifically binds to c-di-GMP. Genetic and phenotypic analyses revealed that Synechocystis CdgR is involved in phototactic motility and natural competence. Inactivation of cdgR resulted in altered expression of specific sets of minor pilins, which are essential for motility or natural competence. We identified interactions between CdgR and the CRP-family transcription factors, SyCRP1 and SyCRP2. Disruption of these CdgR-SyCRP1 and CdgR/SyCRP2 complexes is initiated by elevated c-di-GMP levels. Moreover, the assembly and stability of these complexes are influenced by other cyclic nucleotides, such as cAMP and c-di-AMP. These observed interactions imply a complex regulatory mechanism by which CdgR influences gene expression in response to cyclic nucleotide messenger signalling, particularly c-di-GMP. The present findings highlight the importance of CdgR in c-di-GMP signalling and its role in regulating type IV pilus-dependent functions in Synechocystis. The modulation of the expression of specific minor pilin genes by CdgR, through interactions with the transcription factors SyCRP1 and SyCRP2, contributes to the establishment of multiple type IV pilus functions and adaptive behaviours of cyanobacteria.

Currently, the fungal class Archaeosporomycetes consists of one order, Archaeosporales with four families: Archaeosporaceae, Ambisporaceae, Geosiphonaceae, and Polonosporaceae. In the present study, the objective was to re-analyze the phylogeny and morphology of the Archaeosporomycetes from order to genus level. The different ecological strategies and, consequently, distinct evolutionary patterns of these taxa, as well as their morphological characters and other data updated here, suggest the need to divide Archaeosporales into four orders: (i) the type order Archaeosporales, (ii) Ambisporales ord. nov., both with four genera, (iii) Geosiphonales and (iv) Polonosporales ord. nov., both with single families and genera. Remarkably, the order Geosiphonales was described in the past, but was not considered in the Archaeosporomycetes until now. Phylogenetically, the four main clades (orders here proposed) of Archaeosporomycetes are well supported, with bootstrap values higher than 95% in all analyses, except Ambisporales/Ambisporaceae for RAxML-NG FBP analysis in the SSU tree (75%). Ecologically, this class includes three orders of arbuscular mycorrhizal fungi (AMF) forming symbiotic associations with plants, while Geosiphonales form an endocytobiosis with the cyanobacterium Nostoc. Morphologically, there are at least two AMF orders with spore bimorphism, which has not (yet) been described for Polonosporales. The only known species of Polonosporales, Polonospora polonica, forms spores directly on the neck of sporiferous saccules and the spores can morphologically be differentiated from all other taxa in Archaeosporomycetes by the formation of three permanent, rather thick spore walls, of which two form de novo during spore formation. The outer spore wall of Archaeosporales and Ambisporales are semi-permanent, evanescent or even short-lived, or show multiple fissures during aging, when it is more resistant. Ambisporales can easily be differentiated from Archaeosporales for instance by larger spores of the acaulosporoid morph and thicker spore walls. Our phylogenetic analyses suggested that Archaeosporales can be divided into two families: Antiquisporaceae that was described to form intraradical hyphae, vesicles and spores, staining darkly in Trypan blue, and Archaeosporaceae whose hyphae generally do not or only faintly stain in this reagent, and vesicles and intraradical spores have been rarely, if ever reported.

ASS1 was isolated as a motile Stenotrophomonas strain from crude oil-contaminated soils in Tabasco, Mexico. We characterized this strain using physiological and biochemical traits. ASS1 grew at temperature 25-37 (optimally at 37 {degrees} C) and at pH 6 to 8 (optimally at pH 7 to 8). The assembled genome has a total length of 4.56MB with a G + C content of 66.6%. The 16S rRNA gene sequence analysis confirmed that this strain belongs to the genus Stenotrophomonas. Based on the 16S rRNA analysis, Stenotrophomonas geniculata ATCC 19374 is the closest species, and it shares 99.86% similarity with ASS1. Similarly, a phylogenomic tree based on core genome sequence revealed that the closest species to ASS1 is Stenotrophomonas geniculata ATCC 19374. The major fatty acids in ASS1 are C16:0, antesio C15:0, iso C12:0, iso C15:0, iso C17:0 and C18:0. The genome of ASS1 consists of 4,373,402 bp. The Average Nucleotide Identity (ANI) values for ASS1 which it shared with its closest phylogenetic neighbors, are Stenotrophomonas geniculata ATCC 19374 = JCM 13324 [T] 92.66 %, Stenotrophomonas maltophilia 13637[T] 92.15%, Stenotrophomonas maltophilia K279a 92.13% Stenotrophomonas maltophilia R551-3 92.15% Stenotrophomonas maltophilia MTCC 434 [T] 92.08% and Pseudomonas hibisicicola ATCC [T] 91.66%. ASS1 possesses genes that are essential for the degradation of Polycyclic Aromatic hydrocarbon. Genes such as 1, 2 dihydroxyl 1, 2 dihydronaphthalene dehydrogenase; MG068 17425, homologous to 2 hydroxyl chromene 2 carboxylate isomerases; MG 18055, homologous to salicylaldehyde dehydrogenase and MG068 20095, homologous to naphthalene 1, 2 dioxygenases were identified in ASS1. The dDDH value between ASS1 and its closest neighbor Stenotrophomonas geniculata ATCC 19374 = JCM 13324 [T] is 50%, which is the highest for all the typed species and as such we proposed that ASS1 is a novel species with the name Stenotrophomonas oleivorans sp. nov. sp. nov. and ASS1T as the typed strain

Climate change and increasing drought conditions significantly impedes citrus productivity in subtropical and tropical regions. This study explores the potential of combining arbuscular mycorrhizal fungi (AMF) Funneliformis mosseae and plant growth-promoting rhizobacteria (PGPR) Pseudomonas putida to mitigate drought resilience in Citrus reticulata (Red tangerine). AMF-mediated drought tolerance has been extensively documented; however, the collegial influence of PGPR and AMF on phytohormone signaling, photosynthetic efficiency, nutrient acquisition, and gene expression remains largely unexplored in citrus. We conducted a greenhouse experiment under both well water and drought stress conditions to assess the physiological and molecular responses to individual and co-inoculation with PGPR and AMF. Drought-stressed citrus plants, inoculated with AMF and PGPR, demonstrated significantly improved leaf water potential, stomatal conductance, carbon assimilation, and antioxidant defense. PGPR-AMF co-inoculation enhanced chlorophyll stability, osmotic adjustment, and nutrient uptake, while significantly reducing lipid peroxidation and ROS accumulation. The turquoise module emerged from transcriptomic and gene co-expression network analysis (WGCNA) as a potential key regulator of stress adaptation, revealed key regulatory transcription factors, e.g., CrMYB4, CrZFP8, CrSOS5, CrRGFR2, and CrQUA1, that were upregulated under combined inoculation, highlighting their potential role in stress adaptation. Our findings demonstrate that the synergistic PGPR-AMF interaction improves antioxidant enzyme activities and modulates gene expression to promote drought tolerance, providing new insights into the microbiomes role in plant resilience. These results offer a potential strategy to boost citrus growth and yield under water scarcity, with broad implications for agricultural resilience to climate change.

The phosphodiesterase (PDE) NbdA (NO-induced biofilm dispersion locus A) consists of a membrane-integrated MHYT domain, a degenerated diguanylate cyclase (DGC) AGDEF domain and an EAL domain. The integral membrane domain MHYT is proposed to sense a so far unknown extracellular signal and transfers the information to the cytosolic enzyme domains to modulate cellular c-di-GMP level. Here, we show that full length NbdA from Pseudomonas aeruginosa PAO1 is an active PDE in vivo. In line with its PDE activity, overexpression leads to slightly reduced global c-di-GMP levels, and reduced twitching motility. Surprisingly, overexpression of truncated cytosolic NbdA variants exhibited increased c-diGMP levels, suggesting previously uncharacterized DGC activity despite lacking a canonical GGDEF motif. While full-length NbdA overexpression resulted in only slight c-di-GMP reduction, cytosolic variants induced a significant increase, indicating a potential for nonenzymatic effects like protein-protein interactions. Further investigation revealed a connection between NbdA and type IV pilus (T4P) function. Overexpression of NbdA conferred resistance to the T4P-dependent phage DMS3vir, suggesting interference with T4P assembly or function. Microscopic analysis demonstrated dynamic localization of NbdA, partially co-localizing with T4P components, supporting a role in T4P regulation. However, no clear link was re-established with flagellar motor switching or chemotaxis signaling. These findings position NbdA in the complex signaling network of c-diGMP and T4P-mediated surface behavior in P. aeruginosa. Future work will focus on elucidating the precise mechanisms of NbdAs PDE activity and its interplay with other DGC/PDE networks. ImportanceIn this work, we show the in vivo activity of the membrane-bound phosphodiesterase NbdA of Pseudomonas aeruginosa, its role in c-di-GMP homeostasis, cellular localization and implications in surface behavior. Using strains overexpressing NbdA and truncated protein variants, we detected a strong defect in growth on solid surfaces and an altered phage susceptibility. Co-localization experiments supported further the hypothesis of interaction with the type IV pilus apparatus. We propose for NbdA to be part of the protein network responsible for c-di-GMP level modulation at the cell pole and thereby regulating the function of type IV pilus apparatus.

The transcription factor Pho4 is crucial for the response to phosphate starvation in many fungi, and it has been linked to tolerance to alkalinization of the medium and to pathogenicity. It is widely accepted that it is encoded by a single gene. However, the industrially relevant yeast Komagataella phaffii might contain two Pho4-encoding genes (PAS_chr1-1_0265 and PAS_chr2-1_0177, designated here PHO4(A) and PHO4(B), respectively), which have never been functionally characterized. The phenotypic analysis of single and double mutants suggests that Pho4(B) plays a major role in the adaptation to Pi scarcity. While single mutants exhibited limited and non-overlapping phenotypic defects, the pho4(A) pho4(B) strain was sensitive to multiple types of stress, including phosphate starvation and alkaline pH. Transcriptomic analysis confirms that Pho4(B) is crucial for the transcriptional response to phosphate starvation, including induction of typical gene markers (PHO5, PHO89, VTC1, etc.). However, by using a GFP reporter we found that PHO4(A) also participates in the induction of PHO89 under high pH stress. Expression of both PHO4(A) and PHO4(B) in S. cerevisiae complemented the pho4 mutation under phosphate limitation by restoring growth, expression of the Pho84 transporter and secreted phosphatase activity. These results indicate that both transcription factors display partially overlapping functions, responding differently to diverse stimuli, and that together they constitute a key component in the adaptation to a variety of stresses. Therefore, K. phaffii is an exceptional example among fungi that encodes two Pho4 functional transcription factors.

Phytophthora species are globally significant soilborne oomycetes responsible for widespread ecosystem decline. Standard soil sampling protocols, originally developed for qualitative baiting assays, typically require collecting substantial soil volumes in order to capture viable propagules. While effective for culture-based detection, these protocols are labour-intensive and can damage the shallow root systems of sensitive host species such as New Zealand kauri (Agathis australis). Phytophthora agathidicida (PA), the pathogen associated with kauri dieback disease, is routinely surveyed using these methods. However, quantitative data describing the vertical distribution of PA in natural forest soils are lacking. Consequently, it remains unclear whether extensive depth sampling is necessary to ensure consistent molecular detection. In this study, we applied a quantitative oospore DNA (oDNA) qPCR assay to characterise the fine-scale vertical distribution of PA across four soil depth increments (0-5, 5-10, 10-15, 15-20 cm) from 12 kauri trees representing a range of disease stages. Results revealed distinct vertical stratification, with PA DNA concentrations peaking within the upper 0-10 cm of soil in non-symptomatic and possibly symptomatic trees. In symptomatic trees, the absolute peak occasionally reached 10-15 cm, while pathogen signals remained consistently detectable within the top 10 cm. Field validation from an additional eight trees confirmed that targeted 0-10 cm "shallow" sampling yielded higher PA concentrations than deeper sampling protocols. These findings provide a data-driven basis for refining soil sampling strategies, enabling more sensitive molecular detection while minimising disturbance and logistical effort in fragile ecosystems. IMPORTANCEPhytophthora species are among the most destructive soilborne pathogens globally, requiring robust diagnostic protocols for both agricultural and conservation settings. Traditional sampling frameworks were established to meet the biological requirements of baiting assays, which often necessitate collecting large soil volumes from broad depth profiles to ensure the capture of viable, infectious propagules. However, these extensive requirements are labour-intensive and can cause significant soil disturbance in sensitive forest ecosystems. Using P. agathidicida as a model, this study provides a high-resolution quantitative assessment of how pathogen DNA is distributed vertically across different disease stages. We demonstrate that while absolute peak abundance can shift within the 0-15 cm range as infection progresses, the pathogen signal remains consistently detectable within the top 10 cm. This evidence-based approach suggests that targeted, shallow sampling enhances sensitivity by reducing signal dilution, offering a lower-impact path for monitoring soilborne oomycetes worldwide.

Coordination of metabolism, cell growth and cell division is essential to life. Recent single-cell measurements in S. cerevisiae have shown that metabolic processes and the cellular redox state are dynamic along the cell cycle. However, it is unknown whether similar metabolic oscillations also occur in other organisms. Until now, the dynamics of metabolism in other eukaryotes have predominantly been studied in cell cycle synchronised populations. Since cell cycle synchronisation methods can perturb metabolism, they may also introduce artefacts in the recorded dynamics. Here, we performed time-lapse microscopy analyses of exponentially growing single cells of the budding yeast S. cerevisiae, the fission yeast S. pombe and murine leukaemia L1210 cells. Measuring the NAD(P)H autofluorescence and the cell surface area growth rate in unsynchronised cells, we discovered oscillations along the cell cycle of the cellular redox state and lipid metabolism, respectively. Thus, our work shows that metabolism is dynamic along the cell cycle of these three evolutionarily distant eukaryotic organisms. This finding suggests that such metabolic oscillations could be a conserved characteristic among eukaryotes.

The cryptococcal Amt family of ammonium transporters have been identified from our previous studies as one of the most highly upregulated proteins during transmigration in an in vitro blood-brain barrier (BBB) model, however, the role of this gene family has never been reported. Therefore, this study aimed to investigate the role of the Amt2 gene in the transmigration of C. neoformans across the BBB, examine its association with other common virulence factors, and assess its relevance to morphological changes in C. neoformans. The results showed that the C. neoformans mutant strain lacking the Amt2 gene (amt2{Delta}) exhibited a significantly reduced ability to transmigrate across the BBB in an in vitro model. Our findings suggest that C. neoformans primarily utilizes a transcellular mechanism for invasion, as indicated by the FITC-dextran permeability assays. Additionally, the size of polysaccharide capsule was significantly smaller in the mutant strain compared to the wild-type. In conclusion, our study proposed that the Amt2 gene plays a crucial role in both the transmigration process and capsule production in C. neoformans, without affecting morphological changes. Our study provides a foundation for future research into the underlying mechanisms of the Amt2 gene in C. neoformans pathogenesis. Author summaryCryptococcus neoformans transmigrates the blood-brain barrier through various mechanisms, with transcellular migration being the major route leading to cryptococcal meningitis. In this study, we identified the Amt2 gene, a member of the Amt family of ammonium transporters, as playing a crucial role in the funguss transmigration process. Our findings indicate that the Amt2 gene promotes capsule production and facilitates the transmigration of C. neoformans, all while not causing damage to human endothelial cells.

AbstractBeneficial associations between bobtail squids (Cephalopoda: Sepiolidae) and Vibrio bacteria encompass a unique association where symbionts are obtained environmentally from the surrounding environment. Vibrio symbionts are susceptible to a number of ecological pressures such as protozoan grazing whilst in their free-living state. Impacts of grazing have several consequences for symbiosis characteristics such as biofilm formation, a trait crucial for survival both in and outside the squid. Therefore, in order to ascertain how biotic factors such as grazing in the environment effect symbiotic success, two V. fischeri strains, ES114 and ETBB1-C were experimentally evolved in separate biofilm grazing experiments with the amoeba, Acanthamoeba castellanii and ciliate Tetrahymena pyriformis. Both ES114 and ETBB1-C biofilms were evolved up to 50 generations through serial passaging. At 50 generations, ES114 biofilms displayed variability in response to predation by both predators, whereas ETBB1-C biofilms were more stable across generations of grazing. A. castellanii decreased in population numbers when co-inoculated with ETBB1-C, whereas T. pyriformis increased in numbers with biofilm growth. Growth of V. fischeri biofilms in the presence of grazers such as T. pyriformis has an important role in inducing biofilm growth by acting as a chaperone for recycling nutrients back into the environment. Additionally, V. fischeri colonization fitness in the host was dependent on which grazer was used to evolve the biofilms. Such variation in response by V. fischeri to different types of predation demonstrates the versatility of this symbiont in its free living state and has subsequent impacts on the eventual association with squids. ImportanceThis manuscript demonstrates the importance of biotic factors (such as protozoan grazing) in the environment that effect host colonization in a beneficial symbiosis. Using an experimental evolution approach, this work demonstrates how symbiotic biofilms can adapt to pressures such as grazing that subsequently influences the ability to colonize its invertebrate host.

The fastidious basidiomycete Rhizoctonia (Ceratobasidium) theobromae is a biotrophic pathogen that causes Vascular-Streak Dieback (VSD) of Theobroma cacao (cocoa). The fungus has also been identified as the cause of an emergent disease known as Cassava Witches Broom Disease (CWBD) raising concerns that the pathogen is spreading to alternative hosts and to new regions. Interestingly, while VSD of cocoa and CWBD are reported as co-present in several countries, there is currently no evidence for cross-infection between species. The fungus is difficult to culture in vitro due its slow growth and Kochs postulates have not been definitive on either host. The complete fungus life cycle therefore remains enigmatic, though studies have progressed knowledge on pathology within the both the cocoa and cassava hosts. We have conducted limited field trials and sequenced mating (MAT) and ITS loci of isolates from various infected hosts and regions. We hypothesize that (i) genetic variation at MAT loci correlates with region or host (ii) long amplicon ITS sequences between isolates are more definitive for polymorphisms (iii) life-cycle traits of R. theobromae may be inferred from MAT loci (iv) cassava grown under VSD infected cocoa will be infected and develop symptoms of CWBD. We did not find any cross-infection in field trials, and we show that the pathogen is highly homozygous, despite undergoing meiosis, indicating a predominantly homothallic life cycle. Our data indicate that the pathogen is likely host specific and regionally divergent and suggests that host specificity on cocoa and cassava evolved by selection from a common ancestor rather than a host jump.

The strain LEGE 10371, isolated from the surface of a marine sponge at Praia da Memoria, Portugal, was characterized as a new Thalassoporum species (Pseudanabaenales) using a polyphasic approach that included 16S rRNA gene phylogenetic analysis (Maximum Likelihood and Bayesian Inference), 16S-23S ITS secondary structures, p-distance calculations, MALDI-TOF MS profiling, and morphological analysis by optical and scanning electron microscopy, as well as ecological and biochemical characterization. Phylogenetically, LEGE 10371 clustered within the Thalassoporum clade, however distant from the other existent species of the genus. The p-distance analysis revealed low sequence identity with other Thalassoporum species, with a maximum value of 97.2% to Th. komareki. The MALDI-TOF profile displayed high-intensity peaks at approximately 3,000, 4,000, 6,000 and 8,000 m/z, representing strong candidates for diagnostic markers of the new species. Morphologically, the new species differ from the other species of the genus by presenting trichomes with more than 10 cells and lack of aerotopes. Biocompatibility of the fractions was evaluated in HaCaT keratinocytes, showing no cytotoxic effects at most tested concentrations. PCR screening targeting mcyE, sxtG, anaC, and cyrA confirmed the absence of the genetic potential for the production of major cyanotoxins. Chemical characterization revealed a pigment-rich profile dominated by chlorophyll-a and carotenoids, including {beta}-carotene, zeaxanthin, lutein, and mixoxanthophyll. Bioactivity assays showed superoxide anion radical scavenging by the aqueous fraction (IC2 {approx} 0.042-0.045 mg mL-{superscript 1}), strong nitric oxide radical scavenging by the acetonic fraction (IC = 0.045 mg mL-{superscript 1}), and lipoxygenase inhibition ([~]41%, for a fraction concentration of 0.25 mg mL-), suggesting a potential contribution of these fractions to modulate inflammation-related pathways. Additionally to this results, the polyphasic analysis permitted to confirm previous data that Pseudanabaena and Limnothrix represent the same generic entity. Both genera clustered together, presented high 16S rRNA gene identity (up to 99.9%) and share the same morphological and ecological features. Consequently, we formally proposed the synonimization of Limnothrix into Pseudanabaena.

Leaf diseases pose a serious threat to faba bean production. Leaf blotch of faba bean, caused by Alternaria spp., has become increasingly widespread and destructive in several countries. Leaf diseases pose a serious threat to faba bean production. The infection of plant by pathogens can be influenced by various factors associated with the host plant, environmental conditions and presence of other microorganisms. The phyllosphere and endosphere play a critical role in plant health and disease development. This study aimed to evaluate the factors shaping the structure and diversity of fungal communities associated with faba beans. Plant samples were collected in 2004 from two intensively managed faba bean production fields in the central region of Latvia. Fungal assemblages were characterized using an ITS region metabarcoding approach based on Illumina MiSeq sequencing. Among the assigned amplicon sequence variant (AVS), 65% belonged to the phylum Ascomycota, while approximately 4% were classified as Basidiomycota. Alternaria and Cladosporium were the dominant genera across samples. The alfa and beta diversities of fungal communities was higher during flowering of faba beans to compare with ripening. The higher abundance of Basidiomycota yeasts were observed during flowering, in contrast, Cladosporium genus was significantly more abundant during ripening. Alternaria DNA was found on leaves that showed no symptoms of the disease. The diversity and composition of fungal communities were significantly influenced by sampling time and presence of leaf blotch, caused by Alternaria spp.

Non-pathogenic Xanthomonas (NPX) from a diverse plant hosts are being reported on an increasing basis. There are also reports of multiple species forming communities on a single host plant, such as rice, and, given their role as core endophytes in protecting plants from pathogens, it is essential to isolate and characterization of more NPX species from diverse host plants. Using phylogenomic analysis of publicly available Xanthomonas genome sequences, we identified a novel clade comprising NPX strains from diverse hosts. One of the strains previously reported from our lab is from healthy rice seeds and was reported to be non-pathogenic, with bio-protection function against the bacterial leaf blight pathogen. Genomic investigation confirmed the lack of type III secretion system and its effectors, consistent with their non-pathogenic nature. These strains also harbour core and unique biosynthetic loci identified in other non-pathogenic Xanthomonas (NPX) strains. Further investigation using multiple genomic-based taxonomic indices indicates that these strains represent a potential new species. Hence, we propose Xanthomonas imtechensis sp. nov. as a new species of the genus Xanthomonas, with the type strain being PPL568 = MTCC 13186 = CFBP 9040 = ICMP 24395.

The pELF-type linear plasmid is a unique transconjugative plasmid that disseminates various antimicrobial resistance (AMR) genes (e.g., vancomycin resistance in vancomycin-resistant Enterococcus faecium, [VRE]), leading to hospital outbreaks worldwide. Despite evidence of its role in AMR gene expansion, the molecular mechanisms underlying conjugative transfer of pELF-type plasmids have been poorly understood due to their unique linear structure and the lack of convenient genetic engineering tools adapted to E. faecium strain. Herein, we focused on a putative transconjugation-associated gene cluster, named the trapELF cluster, encoded on pELF2, a vanA-type vancomycin resistance gene-harboring plasmid. RNA-seq analysis of the pELF2 plasmid identified a putative operon containing an ftsK-like ATPase-encoding gene and an additional 10 genes as candidates for conjugative transfer of pELF-type linear plasmids. We constructed isogenic deletion mutants for these 10 genes and revealed that two genes, designated traC and traD, are essential for the conjugative transfer of pELF2 to a recipient strain. Inducible overexpression of the tra genes in trans from an ectopic shuttle vector confirmed the essentiality of traC and traD for conjugative transfer. Promoter assay using a luciferase reporter plasmid revealed that the upstream region of the putative tra operon possesses functional promoter activity characterized by constitutive strong transcriptional expression. Furthermore, we observed membrane localization for a fluorescent protein fusion of TraC, but not for that of TraD. Based on these findings, we propose a working model for the unique molecular machinery for the transconjugation of pELF-type linear plasmids. Author SummaryEnterococci are among the opportunistic pathogens of greatest global concern due to their multidrug resistance, and it is well established that various plasmids mediate horizontal transfer of resistance genes in this species, converting susceptible strains into resistant ones. While most enterococcal plasmids have a circular structure, a linear pELF-type plasmid was recently discovered. Since then, pELF-type plasmids have been reported worldwide as important vehicles for antimicrobial resistance genes, but their mechanism of transfer has remained unknown. In this study, we have, for the first time, successfully identified the genes required for the conjugative transfer of pELF-type plasmids. We confirmed that these genes are absent from known conjugative plasmids, yet are highly conserved among pELF-type plasmid sequences deposited in public databases. Taken together, these findings suggest that pELF-type plasmids possess a unique transfer mechanism that reflects their distinctive molecular architecture.

It is well established that Staphylococcus aureus can incorporate straight-chain unsaturated fatty acids (SCUFAs) into its lipids in addition to the normally biosynthesized branched-chain and straight-chain saturated fatty acids. Incorporation of oleic acid into S. aureus lipids has recently been shown to significantly enhance S. aureus growth at low temperatures due to the greater fluidity imparted to the membrane. Here, we show that low-temperature growth of S. aureus is not limited to oleic acid but enhanced also by various antimicrobial SCUFAs when present at low concentrations. A fakA-deficient strain did not show SCUFA-induced growth stimulation, which indicates that the fatty acid kinase is necessary for SCUFA incorporation into membrane lipids to promote low-temperature growth. Determination of total lipid fatty acid composition showed that incorporated SCUFAs make up [~]12% or less of the total fatty acids. Lipidomic investigations revealed elevated synthesis of diglucosyldiglyceride in the absence or presence of SCUFAs. SCUFAs were incorporated into diglucosyldiglyceride to a greater extent than phosphatidyglycerol at both 12 {degrees}C and 37 {degrees}C. The presence of SCUFAs at low temperatures also enhanced production of the carotenoid staphyloxanthin. The results suggest that multiple strategies are at play in the membrane adaptation of S. aureus to low temperatures. Inclusion of oleic acid in media decreased the minimum growth temperature of S. aureus, suggesting that the presence of SCUFAs in food may facilitate the growth of S. aureus at low temperature. Also, incorporation of SCUFAs into lipids may promote the disruption of the membrane by SCUFAs.

Bacteria of the genus Achromatium are known for their large cell sizes and intracellular calcium carbonate deposits. Achromatium inhabit freshwater, brackish, and marine sediments where they accumulate to high abundances at the oxic-anoxic interface. These bacteria alter their vertical position in the sediment along with daily fluctuations in oxygen concentrations. Yet, the mechanism behind their migration in the sediment remains unknown. In this study, we used chemotaxis assays and time-lapse microphotography to analyze the motility and chemotactic behavior of Achromatium oxaliferum. Microscopic observations revealed that rolling and gliding were the main forms of locomotion exhibited by Achromatium. In absence of any stimulant, the movement appeared to be mostly random and changes in direction frequently occurred. Chemotaxis assays showed a negative chemotaxis of Achromatium to oxygen, sulfide, and nitrate, as evidenced by the change from undirected to directed locomotion against the respective chemical gradient. For periods of more than 1 hour, Achromatium cells moved continuously towards regions of low concentration. We further investigated whether the genetic repertoire of Achromatium corresponds to our observations. Based on lab experiments and bioinformatic analyses we conclude that Achomatium motility is propelled by type IV pili guided by a plethora of chemo- and photoreceptors. We conclude that Achromatium uses negative chemo- and phototaxis to confine their distribution in aquatic sediments between opposing oxygen and sulfide gradients. This allows Achromatium to dynamically adjust its position in redox gradients, and thus is likely to have a major contribution to its success in the global colonization of diverse aquatic sediments.