An app for apples: Citizen-led mapping of fire blight in Central Asia.

Fire blight, caused by the bacterial pathogen Erwinia amylovora, is a severe bacterial disease of apple and pear that can quickly destroy whole plants. In the last decade, it was also detected in Central Asia, where wild pomaceous fruit plants represent the dominant species in mid-altitude forests and constitute a critical foundation for the entire ecosystem. Efficiently informing farmers, forestry services and private persons about the instances and dangers of fire blight, the correct way to recognize the symptoms, and the methods of disease control is thus of paramount importance in a vast and fragmented natural landscape like the one characterizing countries like Kyrgyzstan, Kazakhstan and Tajikistan. For that purpose, we have developed an app for smartphones and mobile devices that can inform stakeholders about fire blight, simultaneously allowing a citizen science approach for mapping the spread of the disease in Central Asia. The app is available in the three national languages as well as in Russian, English, and German, and can easily be adapted to new countries, languages or even diseases.

Fire blight cases in Almaty Region of Kazakhstan in the proximity of wild apple distribution area.

Fire blight caused by Erwinia amylovora reached Kazakhstan in 2008. Here, the disease poses a threat not only to agricultural production of apples and pears, but also to the forests of wild Malus sieversii, the progenitor of most domesticated apple varieties worldwide. In the period 2019-2021, the spread of fire blight in the growth area of wild apples was limited by the weather conditions. In 2022, late spring and early summer were characterized by increased rainfall and moderate temperatures favorable for the disease. The goal of this study was to monitor the distribution of fire blight in private households and small orchards in the zones adjacent to wild apple distribution areas. A total of 91 samples with fire blight-compatible symptoms were collected from cultural apples (68), wild apple (10), pear (5), hawthorn (7), and quince (1) in south-eastern and eastern Kazakhstan, resulting in 21 isolates (one from pear, one from quince, and 19 from apple) of E. amylovora. All isolates belonged to the archetypal CRISPR genotype A. Considering the relative proximity of the infections to the forests of wild M. sieversii, additional measures for fire blight control and prevention will have to be implemented, including state monitoring of the wild apple forests for disease symptoms and awareness campaigns for specially protected natural territories that safeguard M. sieversii, as well as for local pomaceous-fruit growing communities.

Development of two quantitative PCR assays for the detection of emerging opportunistic human pathogens belonging to the genus Phytobacter in routine diagnostics.

The genus Phytobacter (fam. Enterobacteriaceae) includes species like Phytobacter diazotrophicus and Phytobacter ursingii, which have emerged as opportunistic human pathogens, particularly affecting vulnerable populations such as pre-term infants and immunocompromised patients. Traditional biochemical and molecular methods have struggled to accurately identify Phytobacter species in clinical diagnostics. This study addresses the issue by developing and validating two quantitative PCR (qPCR) assays using SYBR® Green I and TaqMan® technologies, targeting the nitrogen fixation regulatory gene (nifL) of Phytobacter spp. The SYBR® Green I assay showed a detection limit of a single cell per reaction, while the TaqMan® assay was easier to interpret due to the absence of background noise. These assays, validated with clinical isolates from Brazil, identified multiple new Phytobacter isolates, including a potentially novel species, providing improved diagnostic tools for detecting Phytobacter spp. and aiding in better clinical management.

Dissemination of NDM-producing bacteria in Southern Brazil.

BACKGROUND: The dissemination of NDM-1 carbapenemases (New Delhi Metallo-ß-lactamase) is a global public health problem, mainly in developing countries. The aim of this study was to characterize the spread of NDM-producing bacteria in the Southern Brazilian states analyzing epidemiological, molecular, and antimicrobial susceptibility aspects. METHODS: A total of 10,684 carbapenem-resistant isolates of Enterobacterales, Pseudomonas spp. and Acinetobacter spp. obtained from several hospitals in eight cities in Southern Brazil were screened, and 486 NDM-producing bacteria were selected. RESULTS: The incidence varied from 0.5 to 77 cases/100.000 habitants. ST11, ST15, ST340 and ST674 were the most common in K. pneumoniae. A total of 5 plasmids were identified in one K. pneumoniae strain: Col440I, Col440II, IncFIA(HI1), IncFIB(K), IncFIB(pQil)/ IncFII(K), and IncR. CONCLUSIONS: The number of patients with NDM-producing bacteria has increased in Southern Brazil, whose gene is present in different plasmids, explaining the expansion of this enzyme.

Comparative genomics to examine the endophytic potential of Pantoea agglomerans DAPP-PG 734.

Pantoea agglomerans DAPP-PG 734 was isolated as endophyte from knots (tumors) caused by Pseudomonas savastanoi pv. savastanoi DAPP-PG 722 in olive trees. To understand the plant pathogen-endophyte interaction on a genomic level, the whole genome of P. agglomerans DAPP-PG 734 was sequenced and annotated. The complete genome had a total size of 5'396'424 bp, containing one circular chromosome and four large circular plasmids. The aim of this study was to identify genomic features that could play a potential role in the interaction between P. agglomerans DAPP-PG 734 and P. savastanoi pv. savastanoi DAPP-PG 722. For this purpose, a comparative genomic analysis between the genome of P. agglomerans DAPP-PG 734 and those of related Pantoea spp. was carried out. In P. agglomerans DAPP-PG 734, gene clusters for the synthesis of the Hrp-1 type III secretion system (T3SS), type VI secretion systems (T6SS) and autoinducer, which could play an important role in a plant-pathogenic community enhancing knot formation in olive trees, were identified. Additional gene clusters for the biosynthesis of two different antibiotics, namely dapdiamide E and antibiotic B025670, which were found in regions between integrative conjugative elements (ICE), were observed. The in-depth analysis of the whole genome suggested a characterization of the P. agglomerans DAPP-PG 734 isolate as endophytic bacterium with biocontrol activity rather than as a plant pathogen.

Resolving taxonomic confusion: establishing the genus Phytobacter on the list of clinically relevant Enterobacteriaceae.

Although many clinically significant strains belonging to the family Enterobacteriaceae fall into a restricted number of genera and species, there is still a substantial number of isolates that elude this classification and for which proper identification remains challenging. With the current improvements in the field of genomics, it is not only possible to generate high-quality data to accurately identify individual nosocomial isolates at the species level and understand their pathogenic potential but also to analyse retrospectively the genome sequence databases to identify past recurrences of a specific organism, particularly those originally published under an incorrect or outdated taxonomy. We propose a general use of this approach to classify further clinically relevant taxa, i.e., Phytobacter spp., that have so far gone unrecognised due to unsatisfactory identification procedures in clinical diagnostics. Here, we present a genomics and literature-based approach to establish the importance of the genus Phytobacter as a clinically relevant member of the Enterobacteriaceae family.

Phytobacter diazotrophicus from Intestine of Caenorhabditis elegans Confers Colonization-Resistance against Bacillus nematocida Using Flagellin (FliC) as an Inhibition Factor.

Symbiotic microorganisms in the intestinal tract can influence the general fitness of their hosts and contribute to protecting them against invading pathogens. In this study, we obtained isolate Phytobacter diazotrophicus SCO41 from the gut of free-living nematode Caenorhabditis elegans that displayed strong colonization-resistance against invading biocontrol bacterium Bacillus nematocida B16. The colonization-resistance phenotype was found to be mediated by a 37-kDa extracellular protein that was identified as flagellin (FliC). With the help of genome information, the fliC gene was cloned and heterologously expressed in E. coli. It could be shown that the B. nematocida B16 grows in chains rather than in planktonic form in the presence of FliC. Scanning Electronic Microscopy results showed that protein FliC-treated B16 bacterial cells are thinner and longer than normal cells. Localization experiments confirmed that the protein FliC is localized in both the cytoplasm and the cell membrane of B16 strain, in the latter especially at the position of cell division. ZDOCK analysis showed that FliC could bind with serine/threonine protein kinase, membrane protein insertase YidC and redox membrane protein CydB. It was inferred that FliC interferes with cell division of B. nematocidal B16, therefore inhibiting its colonization of C. elegans intestines in vivo. The isolation of P. diazotrophicus as part of the gut microbiome of C. elegans not only provides interesting insights about the lifestyle of this nitrogen-fixing bacterium, but also reveals how the composition of the natural gut microbiota of nematodes can affect biological control efforts by protecting the host from its natural enemies.

Complete Genome and Plasmid Sequence Data of Three Strains of Xanthomonas arboricola pv. corylina, the Bacterium Responsible for Bacterial Blight of Hazelnut.

Xanthomonas arboricola pv. corylina is the causal agent of bacterial blight of hazelnut. The bacterium has been listed as an A2 quarantine pathogen in Europe since 1978 and on the regulated non-quarantine pest list since 2019. Three isolates from various geographic regions and isolated at different times were sequenced using a hybrid approach with short- and long-read technologies to generate closed genome and plasmid sequences in order to better understand the biology of this pathogen.

Synergistic interaction between the type III secretion system of the endophytic bacterium Pantoea agglomerans DAPP-PG 734 and the virulence of the causal agent of olive knot Pseudomonas savastanoi pv. savastanoi DAPP-PG 722.

The endophytic bacterium Pantoea agglomerans DAPP-PG 734 was previously isolated from olive knots caused by infection with Pseudomonas savastanoi pv. savastanoi DAPP-PG 722. Whole-genome analysis of this P. agglomerans strain revealed the presence of a Hypersensitive response and pathogenicity (Hrp) type III secretion system (T3SS). To assess the role of the P. agglomerans T3SS in the interaction with P. savastanoi pv. savastanoi, we generated independent knockout mutants in three Hrp genes of the P. agglomerans DAPP-PG 734 T3SS (hrpJ, hrpN, and hrpY). In contrast to the wildtype control, all three mutants failed to cause a hypersensitive response when infiltrated in tobacco leaves, suggesting that P. agglomerans T3SS is functional and injects effector proteins in plant cells. In contrast to P. savastanoi pv. savastanoi DAPP-PG 722, the wildtype strain P. agglomerans DAPP-PG 734 and its Hrp T3SS mutants did not cause olive knot disease in 1-year-old olive plants. Coinoculation of P. savastanoi pv. savastanoi with P. agglomerans wildtype strains did not significantly change the knot size, while the DAPP-PG 734 hrpY mutant induced a significant decrease in knot size, which could be complemented by providing hrpY on a plasmid. By epifluorescence microscopy and confocal laser scanning microscopy, we found that the localization patterns in knots were nonoverlapping for P. savastanoi pv. savastanoi and P. agglomerans when coinoculated. Our results suggest that suppression of olive plant defences mediated by the Hrp T3SS of P. agglomerans DAPP-PG 734 positively impacts the virulence of P. savastanoi pv. savastanoi DAPP-PG 722.

Microbial communities in floodplain ecosystems in relation to altered flow regimes and experimental flooding.

River floodplains are spatially diverse ecosystems that respond quickly to flow variations and disturbance. However, it remains unclear how flow alteration and hydrological disturbance impacts the structure and biodiversity of complex microbial communities in these ecosystems. Here, we examined the spatial and seasonal dynamics of microbial communities in aquatic (benthic) and terrestrial habitats of three hydrologically contrasting (natural flow, residual flow, hydropeaking flow) floodplain systems. Microbial communities (alpha and beta diversity) differed more among floodplain habitats than between riverine floodplains. Microbial communities in all systems displayed congruent seasonal effects. In the residual and hydropeaking systems, an experimental flood was released from a reservoir to mimic a natural high flow event causing hydromorphological disturbance. The experimental flood caused a temporary shift in microbial communities by releasing microbes from the reservoir as well as redistributing communities among floodplain habitats. The flood-mediated shift in community structures had only a transient impact as pelagic bacteria did not persist within floodplain habitats over time after the flood. More frequent pulse disturbances might lead to an alternate structure of bacterial communities in floodplains over time.

Broad diversity of bacteria degrading 17ß-estradiol-3-sulfate isolated from river sediment and biofilm at a wastewater treatment plant discharge.

Conjugated estrogens, such as 17ß-estradiol-3-sulfate (E2-3S), can be released into aquatic environments through wastewater treatment plants (WWTP). There, they are microbiologically degraded into free estrogens, which can have harmful effects on aquatic wildlife. Here, the degradation of E2-3S in environmental samples taken upstream, downstream and at the effluent of a WWTP was assessed. Sediment and biofilm samples were enriched for E2-3S-degrading microorganisms, yielding a broad diversity of bacterial isolates, including known and novel degraders of estrogens. Since E2-3S-degrading bacteria were also isolated in the sample upstream of the WWTP, the WWTP does not influence the ability of the microbial community to degrade E2-3S.

Environmental parameters and microbial community profiles as indication towards microbial activities and diversity in aquaponic system compartments.

BACKGROUND: An aquaponic system couples cultivation of plants and fish in the same aqueous medium. The system consists of interconnected compartments for fish rearing and plant production, as well as for water filtration, with all compartments hosting diverse microbial communities, which interact within the system. Due to the design, function and operation mode of the individual compartments, each of them exhibits unique biotic and abiotic conditions. Elucidating how these conditions shape microbial communities is useful in understanding how these compartments may affect the quality of the water, in which plants and fish are cultured. RESULTS: We investigated the possible relationships between microbial communities from biofilms and water quality parameters in different compartments of the aquaponic system. Biofilm samples were analyzed by total community profiling for bacterial and archaeal communities. The results implied that the oxygen levels could largely explain the main differences in abiotic parameters and microbial communities in each compartment of the system. Aerobic system compartments are highly biodiverse and work mostly as a nitrifying biofilter, whereas biofilms in the anaerobic compartments contain a less diverse community. Finally, the part of the system connecting the aerobic and anaerobic processes showed common conditions where both aerobic and anaerobic processes were observed. CONCLUSION: Different predicted microbial activities for each compartment were found to be supported by the abiotic parameters, of which the oxygen saturation, total organic carbon and total nitrogen differentiated clearly between samples from the main aerobic loop and the anaerobic compartments. The latter was also confirmed using microbial community profile analysis.

Complete or High-Quality Draft Genome Sequences of Six Xanthomonas hortorum Strains Sequenced with Short- and Long-Read Technologies.

We report the genome sequences of six Xanthomonas hortorum species-level clade members, X. hortorum pathovars taraxaci, pelargonii, cynarae, and gardneri (complete genome sequences) and X. hortorum pathovars carotae and vitians (high-quality draft genome sequences). Both short- and long-read sequencing technologies were used.

Isolation and Identification of Actinomycetes Strains from Switzerland and their Biotechnological Potential.

Actinomycetes strains isolated from different habitats in Switzerland were investigated for production of antibacterial and antitumoral compounds. Based on partial 16S rRNA gene sequences, the isolated strains were identified to genus level. Streptomyces as the largest genus of Actinobacteriawas isolated the most frequently. A screening assay using the OmniLog instrument was established to facilitate the detection of active compounds from actinomycetes. Extracts prepared from the cultivated strains able to inhibit Staphylococcus aureusand Escherichia coliwere further analysed by HPLC and MALDI-TOF MS to identify the produced antibiotics. In this study, the bioactive compound echinomycin was identified from two isolated Streptomycesstrains. Natural compounds similar to TPU-0037-C, azalomycin F4a 2-ethylpentyl ester, a derivative of bafilomycin A1, milbemycin-α8 and dihydropicromycin were detected from different isolated Streptomyces strains. Milbemycin-α8 showed cytotoxic activity against HT-29 colon cancer cells. The rare actinomycete,Micromonospora sp. Stup16_C148 produced a compound that matches with the antibiotic bottromycin A2. The draft genome sequence from Actinokineospora strain B136.1 was determined using Illumina and nanopore-based technologies. The isolated strain was not able to produce antibacterial compounds under standard cultivation conditions. The antiSMASH bioinformatics analyses of the genome from strain B136.1 identified biosynthetic gene clusters with identity values between 4% to 90% to known gene clusters encoding antibiotics. The combinations of cultivation conditions, screening assays, analytical methods and genome mining are important tools to characterize strains of actinomycetes for the identification of their potential to produce natural compounds with antimicrobial activity.

Phytobacter palmae sp. nov., a novel endophytic, N2 fixing, plant growth promoting Gammaproteobacterium isolated from oil palm (Elaeis guineensis Jacq.).

A novel strain S29T with high nitrogen fixing ability was isolated from surface-sterilized leaf tissues of oil palm (Elaeis guineensis) growing in Science Park II, Singapore. On the basis of 16S rRNA gene analysis and multilocus sequence typing with the rpoB, gyrB, infB and atpD genes, strain S29T was a member of the genus Phytobacter, with Phytobacter ursingii ATCC 27989T and Phytobacter diazotrophicus LS 8T as its closest relatives. Unique biochemical features that differentiated strain S29T from its closest relatives were the ability to utilize melibiose, α-cyclodextrin, glycogen, adonital, d-arabitol, m-inositol and xylitol. The major fatty acids were C14 : 0, C16 : 0, C17 : 0, C16 : 1 ω5c and summed feature 2 containing C16 : 1 ω7c and/or C16 : 1 ω6c. The polar fatty acid profile consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, an unidentified aminophospholipid and aminolipids. The draft genome of strain S29T comprised 5, 284, 330 bp with a G + C content of 52.6 %. The average nucleotide identity and digital DNA-DNA hybridization values between strain S29T and the phylogenetically related Enterobacterales species were lower than 95 % and 70 %, respectively. Thus, the polyphasic evidences generated through the phenotypic, chemotaxonomic and genomic methods confirmed that strain S29T represents a novel species of the genus Phytobacter, for which we propose the name Phytobacter palmae sp. nov. with the type strain of S29T (=DSM 27342T=KACC 17598T).

Updated Genome Sequence and Annotation for the Full Genome of Pseudomonas protegens CHA0.

Minor differences in the previously obtained genome of Pseudomonas protegens CHA0 were detected after resequencing the strain. Based on this, the genome size slightly increased. Additionally, we performed a manual annotation of genes involved in biocontrol and insect pathogenicity. This annotation version will be the basis for upcoming genome studies.

High-Quality Draft Genome Sequence of Pseudomonas reidholzensis Strain CCOS 865T.

We have sequenced and assembled the genome of Pseudomonas reidholzensis CCOS 865T, which was isolated in 2014 from forest soil. Members of the genus Pseudomonas play important roles in environmental systems and are utilized in many biotechnological processes. The genome of this species may provide an important resource for the discovery of novel enzyme activities.

High-Quality Draft Genome Sequence of Pseudomonas wadenswilerensis CCOS 864T.

Pseudomonas wadenswilerensis CCOS 864T was isolated in 2014 from forest soil. The organism belongs taxonomically to the Pseudomonas putida group, members of which have been well studied for their potential in biotechnological applications. We present here the draft genome sequence of P. wadenswilerensis CCOS 864T.

Molecular investigation of isolates from a multistate polymicrobial outbreak associated with contaminated total parenteral nutrition in Brazil.

BACKGROUND: Between November 2013 and June 2014, 56 cases of bacteremia (15 deaths) associated with the use of Total Parenteral Nutrition (TPN) and/or calcium gluconate (CG) were reported in four Brazilian states. METHODS: We analyzed 73 bacterial isolates from four states: 45 from blood, 25 from TPN and three from CG, originally identified as Acinetobacter baumannii, Rhizobium radiobacter, Pantoea sp. or Enterobacteriaceae using molecular methods. RESULTS: The first two bacterial species were confirmed while the third group of species could not be identified using standard identification protocols. These isolates were subsequently identified by Multi-Locus Sequence Analysis as Phytobacter diazotrophicus, a species related to strains from similar outbreaks in the United States in the 1970's. Within each species, TPN and blood isolates proved to be clonal, whereas the R. radiobacter isolates retrieved from CG were found to be unrelated. CONCLUSION: This is the first report of a three-species outbreak caused by TPN contaminated with A. baumannii, R. radiobacter and P. diazotrophicus. The concomitant presence of clonal A. baumannii and P. diazotrophicus isolates in several TPN and blood samples, as well as the case of one patient, where all three different species were isolated simultaneously, suggest that the outbreak may be ascribed to a discrete contamination of TPN. In addition, this study highlights the clinical relevance of P. diazotrophicus, which has been involved in outbreaks in the past, but was often misidentified as P. agglomerans.

Genome-based population structure analysis of the strawberry plant pathogen Xanthomonas fragariae reveals two distinct groups that evolved independently before its species description.

Xanthomonas fragariae is a quarantine organism in Europe, causing angular leaf spots on strawberry plants. It is spreading worldwide in strawberry-producing regions due to import of plant material through trade and human activities. In order to resolve the population structure at the strain level, we have employed high-resolution molecular typing tools on a comprehensive strain collection representing global and temporal distribution of the pathogen. Clustered regularly interspaced short palindromic repeat regions (CRISPRs) and variable number of tandem repeats (VNTRs) were identified within the reference genome of X. fragariae LMG 25863 as a potential source of variation. Strains from our collection were whole-genome sequenced and used in order to identify variable spacers and repeats for discriminative purpose. CRISPR spacer analysis and multiple-locus VNTR analysis (MLVA) displayed a congruent population structure, in which two major groups and a total of four subgroups were revealed. The two main groups were genetically separated before the first X. fragariae isolate was described and are potentially responsible for the worldwide expansion of the bacterial disease. Three primer sets were designed for discriminating CRISPR-associated markers in order to streamline group determination of novel isolates. Overall, this study describes typing methods to discriminate strains and monitor the pathogen population structure, more especially in the view of a new outbreak of the pathogen.