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Bacterial soft rots are among the most economically important diseases that cause significant yield losses in various crops. During 2022 and 2023, widespread occurrence of soft rot disease was observed in sugar beet fields in the Firuzan region, Hamedan Province, Iran. From the sugar beet plants with soft rot symptoms, a bacterial isolate was obtained that was obligately aerobic, negative for Gram, oxidase, levan, and arginine dihydrolase tests, and positive for pectinase, urease and hypersensitivity reaction on tobacco. Koch's postulates confirmed its pathogenicity by developing soft rot symptoms in sugar beet plants. The 16SrRNA and rpoD sequences from this isolate were most closely related to Pseudomonas punonensis LMT03 with 99.8% and 94.7% identity, respectively and then to Pseudomonas argentinensis LMG22563 (= CH01) with 99.4% and 94.3% identity values. To our knowledge, this is the first report of a Pseudomonas sp. strain causing soft rot disease in sugar beet worldwide. [ABSTRACT FROM AUTHOR]

The Pseudomonas chlororaphis phylogenetic subgroup of species, within the Pseudomonas fluorescens group, currently includes seven bacterial species, all of which have environmental relevance. Phylogenomic analyses help clarify the taxonomy of strains in the group and allow for precise identification. Thirteen antibiotic-resistant strains isolated in a previous study from nine different sampling sites in the Danube River were suspected to represent a novel species and are investigated taxonomically in the present study, together with four other strains isolated from the Woluwe River (Belgium) that were phylogenetically closely related in their rpoD gene sequences. The strains were characterized phenotypically, chemotaxonomically (fatty acid composition and main protein profiles), and phylogenetically. They could not be assigned to any known Pseudomonas species. Three genomes of representative strains were sequenced and analyzed in the context of the genome sequences of closely related strains available in public databases. The phylogenomic analysis demonstrates the need to differentiate new genomic species within the P. chlororaphis subgroup and that Pseudomonas piscis and Pseudomonas aestus are synonyms. This taxonomic study demonstrates that 14 of the characterized isolates are members of the Pseudomonas_E protegens_A species in the GTDB taxonomy and that they represent a novel species in the genus Pseudomonas, for which we propose the name Pseudomonas danubii sp. nov. with strain JDS02PS016T (=CECT 30214T = CCUG 74756T) as the type strain. The other three strains (JDS08PS003, rDWA16, and rDWA64) are members of the species Pseudomonas_E protegens_B in the GTDB taxonomy and need further investigation for proposal as a new bacterial species. [ABSTRACT FROM AUTHOR]

Pseudomonas plecoglossicida is a Gram-negative pathogenic bacterium that causes visceral white spot disease in several marine fish species, resulting in high mortality and financial loss. Based on previous RNA sequencing (RNA-seq) results, rpoD gene expression is significantly up-regulated in P. plecoglossicida during infection, indicating that rpoD may contribute to bacterial pathogenicity. To investigate the role of this gene, five specific short hairpin RNAs (shRNAs) were designed and synthesized based on the rpoD gene sequence, with all five mutants exhibiting a significant decrease in rpoD gene expression in P. plecoglossicida. The mutant with the highest silencing efficiency (89.2%) was chosen for further study. Compared with the wild-type (WT) P. plecoglossicida strain NZBD9, silencing rpoD in the rpoD -RNA interference (RNAi) strain resulted in a significant decrease in growth, motility, chemotaxis, adhesion, and biofilm formation in P. plecoglossicida. Silencing of rpoD also resulted in a 25% increase in the survival rate, a one-day delay in the onset of death, and a significant decrease in the number of white spots on the spleen surface of infected orange-spotted groupers (Epinephelus coioides). In addition, rpoD expression and pathogen load were significantly lower in the spleens of E. coioides infected with the rpoD -RNAi strain than with the WT strain of P. plecoglossicida. We performed RNA-seq of E. coioides spleens infected with different P. plecoglossicida strains. Results showed that rpoD silencing in P. plecoglossicida led to a significant change in the infected spleen transcriptomes. In addition, comparative transcriptome analysis showed that silencing rpoD caused significant changes in complement and coagulation cascades and the IL-17 signaling pathway. Thus, this study revealed the effects of the rpoD gene on P. plecoglossicida pathogenicity and identified the main pathway involved in the immune response of E. coioides. • Pseudomonas plecoglossicida rpoD gene was silenced by RNA interference with an efficiency of 89.2%. • Silencing rpoD resulted in a 25% increase in the survival rate of E. coioides infected by P. plecoglossicida. • Silencing rpoD significant affected the immune response of infected E. coioides. [ABSTRACT FROM AUTHOR]

Japanese quince trees are grown as ornamental plants in Iran, in parks and in orchards close to stone fruit and pome fruit trees. Shoots of Japanese quince (Chaenomeles japonica) showing sunken brown canker symptoms were observed and collected near Sari, the center of Mazandaran province in the North of Iran, during the 2016 growing season. Gram negative bacteria isolated from symptomatic tissues were similar to Pseudomonas syringae pv. syringae (Pss) were pathogenic on Japanese quince and on quince (Cydonia oblonga) seedlings after artificial inoculation, and were re-isolated from diseased hosts. Phylogenetic tree construction using partial sequences of ITS and rpoD genes showed that the Japanese quince isolates were in the same clade as Pss strains. The isolates had ice nucleation activity, and the InaK gene was amplified successfully. According to the results of phenotypic and genotypic characteristics, genomic DNA fingerprinting using REP-PCR, BOX-PCR and IS50-PCR and isolation of total cell proteins, we conclude that Pss is the causal agent of canker of the Japanese quince trees. Therefore, Japanese quince is a new host for Pss causing bacterial canker on many different host plants. [ABSTRACT FROM AUTHOR]

Japanese quince trees are grown as ornamental plants in Iran, in parks and in orchards close to stone fruit and pome fruit trees. Shoots of Japanese quince (Chaenomeles japonica) showing sunken brown canker symptoms were observed and collected near Sari, the center of Mazandaran province in the North of Iran, during the 2016 growing season. Gram negative bacteria isolated from symptomatic tissues were similar to Pseudomonas syringae pv. syringae (Pss) were pathogenic on Japanese quince and on quince (Cydonia oblonga) seedlings after artificial inoculation, and were re-isolated from diseased hosts. Phylogenetic tree construction using partial sequences of ITS and rpoD genes showed that the Japanese quince isolates were in the same clade as Pss strains. The isolates had ice nucleation activity, and the InaK gene was amplified successfully. According to the results of phenotypic and genotypic characteristics, genomic DNA fingerprinting using REP-PCR, BOX-PCR and IS50-PCR and isolation of total cell proteins, we conclude that Pss is the causal agent of canker of the Japanese quince trees. Therefore, Japanese quince is a new host for Pss causing bacterial canker on many different host plants.

The Pseudomonas chlororaphis phylogenetic subgroup of species, within the Pseudomonas fluorescens group, currently includes seven bacterial species, all of which have environmental relevance. Phylogenomic analyses help clarify the taxonomy of strains in the group and allow for precise identification. Thirteen antibiotic-resistant strains isolated in a previous study from nine different sampling sites in the Danube River were suspected to represent a novel species and are investigated taxonomically in the present study, together with four other strains isolated from the Woluwe River (Belgium) that were phylogenetically closely related in their rpoD gene sequences. The strains were characterized phenotypically, chemotaxonomically (fatty acid composition and main protein profiles), and phylogenetically. They could not be assigned to any known Pseudomonas species. Three genomes of representative strains were sequenced and analyzed in the context of the genome sequences of closely related strains available in public databases. The phylogenomic analysis demonstrates the need to differentiate new genomic species within the P. chlororaphis subgroup and that Pseudomonas piscis and Pseudomonas aestus are synonyms. This taxonomic study demonstrates that 14 of the characterized isolates are members of the Pseudomonas_E protegens_A species in the GTDB taxonomy and that they represent a novel species in the genus Pseudomonas, for which we propose the name Pseudomonas danubii sp. nov. with strain JDS02PS016T (=CECT 30214T = CCUG 74756T) as the type strain. The other three strains (JDS08PS003, rDWA16, and rDWA64) are members of the species Pseudomonas_E protegens_B in the GTDB taxonomy and need further investigation for proposal as a new bacterial species.

Cassava (Manihot esculenta Crantz) is one of the most important crops in Vietnam for providing food, starch sources, and raw materials for the production of bio-ethanol and other purposes. Cassava bacterial blight (CBB) disease, caused by Xanthonomas axonopodis pv. manihotis (Xam), is one of the most important factors affecting cassava production in Vietnam. A rapid and sensitive molecular tool is required to support the traditional method, primarily based on biochemical reactions, that was considered less sensitive and time-consuming. In the present study, in order to detect Xam, the Polymerase Chain Reaction (PCR) technique was applied with the two primer pairs, rpoD_17F/rpoD_1005R and XgyrconpcrF1/Xgyrconrpcr1, to amplify partial sequences of rpoD and gyrB genes, respectively. The primers directed the amplification of about 900 bp DNA fragments from CBB-infected leaf and stem tissues. All the amplified DNA sequences were identical in each gene and to the Xam reference strains (accession numbers KP265376 and KP265378 for the rpoD gene, and KP265387 and KP265388 for the gyrB gene) from GenBank. The representative sequences of rpoD and gyrB genes were deposited in GenBank with accession numbers MF774491 and MF774490, respectively. DNA and phylogenetic analysis based on the sequences of rpoD and gyrB genes have confirmed that Xam is the causal agent of CBB in Dong Nai, Vietnam. This is the first confirmed identification of the causal pathogen of CBB disease in Vietnam using molecular tools, and this method is a reliable tool for the detection and identification of other plant bacterial pathogens.

ABSTRACT The genome of the metal-resistant, hydrogen-oxidizing bacterium Cupriavidus metallidurans contains a large number of horizontally acquired plasmids and genomic islands that were integrated into its chromosome or chromid. For the C. metallidurans CH34 wild-type strain growing under nonchallenging conditions, 5,763 transcriptional starting sequences (TSSs) were determined. Using a custom-built motif discovery software based on hidden Markov models, patterns upstream of the TSSs were identified. The pattern TTGACA, −35.6 ± 1.6 bp upstream of the TSSs, in combination with a TATAAT sequence 15.8 ± 1.4 bp upstream occurred frequently, especially upstream of the TSSs for 48 housekeeping genes, and these were assigned to promoters used by RNA polymerase containing the main housekeeping sigma factor RpoD. From patterns upstream of the housekeeping genes, a score for RpoD-dependent promoters in C. metallidurans was derived and applied to all 5,763 TSSs. Among these, 2,572 TSSs could be associated with RpoD with high probability, 373 with low probability, and 2,818 with no probability. In a detailed analysis of horizontally acquired genes involved in metal resistance and not involved in this process, the TSSs responsible for the expression of these genes under nonchallenging conditions were assigned to RpoD- or non-RpoD-dependent promoters. RpoD-dependent promoters occurred frequently in horizontally acquired metal resistance and other determinants, which should allow their initial expression in a new host. However, other sigma factors and sense/antisense effects also contribute—maybe to mold in subsequent adaptation steps the assimilated gene into the regulatory network of the cell. IMPORTANCE In their natural environment, bacteria are constantly acquiring genes by horizontal gene transfer. To be of any benefit, these genes should be expressed. We show here that the main housekeeping sigma factor RpoD plays an important role in the expression of horizontally acquired genes in the metal-resistant hydrogen-oxidizing bacterium C. metallidurans. By conservation of the RpoD recognition consensus sequence, a newly arriving gene has a high probability to be expressed in the new host cell. In addition to integrons and genes travelling together with that for their sigma factor, conservation of the RpoD consensus sequence may be an important contributor to the overall evolutionary success of horizontal gene transfer in bacteria. Using C. metallidurans as an example, this publication sheds some light on the fate and function of horizontally acquired genes in bacteria.

Abstract The damages observed in Tunisian citrus orchards have prompted studies on the Pseudomonas spp. responsible for blast and black pit. Prospective orchards between 2015 and 2017 showed that the diseases rapidly spread geographically and to new cultivars. A screening of Pseudomonas spp. isolated from symptomatic trees revealed their wide diversity according to phylogenetic analysis of their housekeeping rpoD and cts genes. The majority of strains were affiliated to Pseudomonas syringae pv. syringae (Phylogroup PG02b), previously described in Tunisia. However, they exhibited various BOX-PCR fingerprints and were not clonal. This work demonstrated, for the first time in Tunisia, the involvement of Pseudomonas cerasi (PG02a) and Pseudomonas congelans (PG02c). The latter did not show significant pathogenicity on citrus, but was pathogenic on cantaloupe and active for ice nucleation that could play a role in the disease. A comparative phylogenetic study of citrus pathogens from Iran, Montenegro and Tunisia revealed that P. syringae (PG02b) strains are closely related but again not clonal. Interestingly P. cerasi (PG02a) was isolated in two countries and seems to outspread. However, its role in the diseases is not fully understood and it should be monitored in future studies. The diversity of pathogenic Pseudomonas spp. and the extension of the diseases highlight that they have become complex and synergistic. It opens questions about which factors favor diseases and how to fight against them efficiently and with sustainable means.

ABSTRACT Species of the genus Pseudomonas are used for several biotechnological purposes, including plant biocontrol and bioremediation. To exploit the Pseudomonas genus in environmental, agricultural, or industrial settings, the organisms must be profiled at the species level as their bioactivity potential differs markedly between species. Standard 16S rRNA gene amplicon profiling does not allow for accurate species differentiation. Thus, the purpose of this study was to develop an amplicon-based high-resolution method targeting a 760-nucleotide (nt) region of the rpoD gene enabling taxonomic differentiation of Pseudomonas species in soil samples. The method was benchmarked on a 16-member Pseudomonas species mock community. All 16 species were correctly and semiquantitatively identified using rpoD gene amplicons, whereas 16S rRNA V3-V4 amplicon sequencing only correctly identified one species. We analyzed the Pseudomonas profiles in 13 soil samples in northern Zealand, Denmark, where samples were collected from grassland (3 samples) and agriculture soil (10 samples). Pseudomonas species represented up to 0.7% of the 16S rRNA gene abundance, of which each sampling site contained a unique Pseudomonas composition. Thirty culturable Pseudomonas strains were isolated from each grassland site and 10 from each agriculture site and identified by Sanger sequencing of the rpoD gene. In all cases, the rpoD amplicon approach identified more species than were found by cultivation, including hard-to-culture nonfluorescent pseudomonads, as well as more than were found by 16S rRNA V3-V4 amplicon sequencing. Thus, rpoD profiling can be used for species profiling of Pseudomonas, and large-scale prospecting of bioactive Pseudomonas may be guided by initial screening using this method. IMPORTANCE A high-throughput sequencing-based method for profiling of Pseudomonas species in soil microbiomes was developed and identified more species than 16S rRNA gene sequencing or cultivation. Pseudomonas species are used as biocontrol organisms and plant growth-promoting agents, and the method will allow tracing of specific species of Pseudomonas as well as enable screening of environmental samples for further isolation and exploitation.

This paper proposes a comparison of the numerical aspect and efficiency of several low rank approximation techniques for multidimensional data, namely CPD, HOSVD, TT-SVD, RPOD, QTT-SVD and HT. This approach is different from the numerous papers that compare the theoretical aspects of these methods or propose efficient implementation of a single technique. Here, after a brief presentation of the studied methods, they are tested in practical conditions in order to draw hindsight at which one should be preferred. Synthetic data provides sufficient evidence for dismissing CPD, T-HOSVD and RPOD. Then, three examples from mechanics provide data for realistic application of TT-SVD and ST-HOSVD. The obtained low rank approximation provides different levels of compression and accuracy depending on how separable the data is. In all cases, the data layout has significant influence on the analysis of modes and computing time while remaining similarly efficient at compressing information. Both methods provide satisfactory compression, from 0.1% to 20% of the original size within a few percent error in L 2 norm. ST-HOSVD provides an orthonormal basis while TT-SVD doesn’t. QTT is performing well only when one dimension is very large. A final experiment is applied to an order 7 tensor with (4 × 8 × 8 × 64 × 64 × 64 × 64) entries (32 GB) from complex multi-physics experiment. In that case, only HT provides actual compression (50%) due to the low separability of this data. However, it is better suited for higher order d. Finally, these numerical tests have been performed with pydecomp , an open source python library developed by the author. [ABSTRACT FROM AUTHOR]

A collection of 611 Pseudomonas isolated from 14 sampling sites along the Danube River were identified previously by MALDI-TOF MS with the VITEK MS system and were grouped in 53 clusters by their main protein profiles. The strains were identified in the present study at the phylospecies level by rpoD gene sequencing. Partial sequences of the rpoD gene of 190 isolates representatives of all clusters were analyzed. Strains in the same MALDI-TOF cluster were grouped in the same phylospecies when they shared a minimum 95% similarity in their rpoD sequences. The sequenced strains were assigned to 34 known species (108 strains) and to 32 possible new species (82 strains). The 611 strains were identified at the phylospecies level combining both methods. Most strains were assigned to phylospecies in the Pseudomonas putida phylogenetic group of species. Special attention was given to 14 multidrug resistant strains that could not be assigned to any known Pseudomonas species and were considered environmental reservoir of antibiotic resistance genes. Coverage indices and rarefaction curves demonstrated that at least 50% of the Pseudomonas species in the Danube River able to grow in the isolation conditions have been identified at the species level. Main objectives were the confirmation of the correlation between the protein profile clusters detected by MALDI-TOF MS and the phylogeny of Pseudomonas strains based on the rpoD gene sequence, the assessment of the higher species discriminative power of the rpoD gene sequence, as well as the estimation of the high diversity of Pseudomonas ssp. along the Danube river. This study highlights the Pseudomonas species diversity in freshwater ecosystems and the usefulness of the combination of MALDI-TOF mass spectrometry for the dereplication of large sets of strains and the rpoD gene sequences for rapid and accurate identifications at the species level.

Abstract Background The biological production of 2,3-butanediol from xylose-rich raw materials from Klebsiella pneumoniae is a low-cost process. RpoD, an encoding gene of the sigma factor, is the key element in global transcription machinery engineering and has been successfully used to improve the fermentation with Escherichia coli. However, whether it can regulate the tolerance in K. pneumoniae remains unclear. Results In this study, the kpC mutant strain was constructed by altering the expression quantity and genotype of the rpoD gene, and this exhibited high xylose tolerance and 2,3-butanediol production. The xylose tolerance of kpC strain was increased from 75 to 125 g/L, and the yield of 2,3-butanediol increased by 228.5% compared with the parent strain kpG, reaching 38.6 g/L at 62 h. The RNA sequencing results showed an upregulated expression level of 500 genes and downregulated expression level of 174 genes in the kpC mutant strain. The pathway analysis further showed that the differentially expressed genes were mainly related to signal transduction, membrane transport, carbohydrate metabolism, and energy metabolism. The nine most-promising genes were selected based on transcriptome sequencing, and were evaluated for their effects on xylose tolerance. The overexpression of the tktA encoding transketolase, pntA encoding NAD(P) transhydrogenase subunit alpha, and nuoF encoding NADH dehydrogenase subunit F conferred increased xylose consumption and increased 2,3-butanediol production to K. pneumoniae. Conclusions These results suggest that the xylose tolerance and 2,3-butanediol production of K. pneumoniae can be greatly improved by the directed evolution of rpoD. By applying transcriptomic analysis, the upregulation of tktA, pntA, and nuoF that were coded are essential for the xylose consumption and 2,3-butanediol production. This study will provide reference for further research on improving the fermentation abilities by means of other organisms.

A collection of 611 Pseudomonas isolated from 14 sampling sites along the Danube River were identified previously by MALDI-TOF MS with the VITEK MS system and were grouped in 53 clusters by their main protein profiles. The strains were identified in the present study at the phylospecies level by rpoD gene sequencing. Partial sequences of the rpoD gene of 190 isolates representatives of all clusters were analyzed. Strains in the same MALDI-TOF cluster were grouped in the same phylospecies when they shared a minimum 95% similarity in their rpoD sequences. The sequenced strains were assigned to 34 known species (108 strains) and to 32 possible new species (82 strains). The 611 strains were identified at the phylospecies level combining both methods. Most strains were assigned to phylospecies in the Pseudomonas putida phylogenetic group of species. Special attention was given to 14 multidrug resistant strains that could not be assigned to any known Pseudomonas species and were considered environmental reservoir of antibiotic resistance genes. Coverage indices and rarefaction curves demonstrated that at least 50% of the Pseudomonas species in the Danube River able to grow in the isolation conditions have been identified at the species level. Main objectives were the confirmation of the correlation between the protein profile clusters detected by MALDI-TOF MS and the phylogeny of Pseudomonas strains based on the rpoD gene sequence, the assessment of the higher species discriminative power of the rpoD gene sequence, as well as the estimation of the high diversity of Pseudomonas ssp. along the Danube river. This study highlights the Pseudomonas species diversity in freshwater ecosystems and the usefulness of the combination of MALDI-TOF mass spectrometry for the dereplication of large sets of strains and the rpoD gene sequences for rapid and accurate identifications at the species level. [ABSTRACT FROM AUTHOR]

Abstract Escherichia coli (E. coli) is a promising platform for expression of full-length antibodies owing to its several advantages as a production host (fast growth, well characterized genetics, low manufacturing cost), however, low titers from shake flask (typically < 5 mg/L) has limited its use for production of research-grade material in antibody discovery programs. In this work, we used global transcriptional machinery engineering (gTME) with high throughput screening to increase the expression of full-length antibodies in E. coli. A library of E. coli mutants carrying mutations in the global sigma factor RpoD were generated and screened using the Bacterial Antibody Display (BAD) method for enhanced expression. RpoD mutants were isolated that resulted in full-length antibody titers of up to 130.7 ± 6.6 mg/L of shake flask culture with chaperone co-expression. These results could be useful for production of several antibodies quickly in shake flasks for characterization (e.g. antigen binding, biological function) during the early discovery phase. Highlights • Sigma factor RpoD was engineered to increase full-length IgG expression in E. coli. • Three RpoD mutants were isolated using a screen for enhanced IgG expression. • A truncated RpoD mutant with only a partial region 4 resulted in highest expression. • RpoD mutant + chaperone expression resulted in > 100 mg/L IgG titer in shake flasks. [ABSTRACT FROM AUTHOR]

Abstract In E. coli , a single oligomeric enzyme transcribes the genomic DNA, while multiple auxiliary proteins and regulatory RNA interact with the core RNA polymerase (RP) during different stages of the transcription cycle to influence its function. In this work, using fast protein isolation techniques combined with mass spectrometry (MS) and immuno-analyses, we studied growth phase-specific changes in the composition of E. coli transcription complexes. We show that RP isolated from actively growing cells is represented by prevalent double copy assemblies and single copy RP–RNA and RP–RNA–RapA complexes. We demonstrate that RpoD/σ70 obtained in fast purification protocols carries tightly associated RNA and show evidence pointing to a role of sigma-associated RNA in the formation of native RP–(RNA)–RpoD/σ70 (holoenzyme) complexes. We report that enzymes linked functionally to the metabolism of lipopolysaccharides co-purify with RP–RNA complexes and describe two classes of RP-associated molecules (phospholipids and putative phospholipid–rNT species). We hypothesize that these modifications could enable anchoring of RP–RNA and RNA in cell membranes. We also report that proteins loosely associated with ribosomes and degradosomes (S1, Hfq) co-purify with RP–RNA complexes isolated from actively growing cells – a result consistent with their proposed roles as adaptor-proteins. In contrast, GroEL, SecB, and SecA co-purified with RP obtained from cells harvested in early stationary phase. Our results demonstrate that fast, affinity chromatography-based isolation of large multi-protein assemblies in combination with MS can be used as a tool for analysis of their composition and the profiling of small protein-associated molecules (SPAM). Highlights • Growth phase-specific changes in the makeup of E. coli RP complexes were analyzed. • LC-MS and immunoassays were used to identify RP-associated proteins and SPAM. • RP-associated lipids and small accessory nucleic acid fragments were studied. • MS can be used to profile SPAM in multi-protein complexes obtained via fast methods. [ABSTRACT FROM AUTHOR]

Quorum sensing is a well-studied cell-to-cell communication method that involves a cell-density dependent regulation of genes expression mediated by signalling molecules. In this study, a bacterium isolated from a plant material compost pile was found to possess quorum sensing activity based on bioassay screening. Isolate YL12 was identified using matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) mass spectrometry and molecular typing using rpoD gene which identified the isolate as Aeromonas caviae. High resolution tandem mass spectrometry was subsequently employed to identify the N-acyl homoserine lactone profile of Aeromonas caviae YL12 and confirmed that this isolate produced two short chain N-acyl homoserine lactones, namely C4-HSL and C6, and the production was observed to be cell density-dependent. Using the thin layer chromatography (TLC) bioassay, both AHLs were found to activate C. violaceum CV026, whereas only C6-HSL was revealed to induce bioluminescence expression of E. coli [pSB401]. The data presented in this study will be the leading steps in understanding the role of quorum sensing in Aeromonas caviae strain YL12.

Pseudomonas fluorescens are known bacterial pathogens in fish. The P. fluorescens group contains at least nine different bacterial species, although species from fish have rarely been differentiated. Two isolated fish kills affecting wild bluegills, Lepomis macrochirus Rafinesque, and pumpkinseed sunfish, Lepomis gibbosus (Linnaeus), occurred in the spring of 2015 during cool water temperatures (12.5°C-15.5°C). Disease signs included severe bacteraemia with rare gross external signs. Pure bacterial cultures isolated from kidneys of all affected fish were identified as P. fluorescens using the API 20 NE system, while no bacteria were isolated from asymptomatic fish. To further identify the species of bacterium within the P. fluorescens complex, genetic analysis of the 16S rRNA, rpoD and gyrB genes was conducted. DNA sequences of bacterial isolates from both mortality events were identical and had close identity (≥99.7%) to Pseudomonas mandelii. Although likely widespread in the aquatic environment, this is the first report of a bacterium closely resembling P. mandelii infecting and causing disease in fish. The bacterium grew at temperatures between 5°C and 30°C, but not at 37°C. It is possible that infections in fish were a result of immunosuppression associated with spring conditions combined with the psychrotrophic nature of the bacterium. [ABSTRACT FROM AUTHOR]

Pseudomonas species isolated from rainbow trout (Oncorhynchus mykiss) in the Eastern Black Sea Region of Turkey were identified on the basis of three different genes (16S rRNA, rpoD and rpoB). Different Pseudomonas strains were analyzed using biochemical tests and PCR asssays designed to for Pseudomonas species. The rpoD gene was show to be more discriminatory than the 16S rRNA and rpoB genes. Phylogenetic analyses discerned two main groups as Pseudomonas fluorescens and Pseudomonas putida. The Pseudomonas strains used in this study were mainly identified as Pseudomonas fluorescens or Pseudomonas putida although other Pseudomonas spp. were characterised. [ABSTRACT FROM AUTHOR]

In the late summer of 2015, severe leaf blight occurred on Miscanthus sinensis grown on natural riverside lands of the Han River in Seoul, South Korea. Symptoms included chlorosis and necrotic lesions on the leaves. Gram-negative, non-spore-forming, and rod-shaped Pseudomonas syringae were consistently recovered from the infected leaves. We provide ample evidence for P. syringae pv. syringae ( Pss) as the pathogen, based on the results of LOPAT, Biolog, API tests, MALDI-TOF MS, and multilocus sequence analysis. Symptoms after artificial inoculation were mild compared to symptoms following natural infection, whereas the control plants were asymptomatic. Bacteria re-isolated from leaves with mild symptoms were identical to the original strain, as determined by the rpoD gene sequence. This is the first report of Pss causing bacterial leaf blight on M. sinensis. [ABSTRACT FROM AUTHOR]

Rice is the dominant cereal crop of North India. It is regularly affected by grain discoloration and chaffiness of panicles during maturity of the crop. Lack of effective management practices for this problem has been a major concern among farmers in the region. Farmers give many sprays of fungicides due to the already reported association of fungal pathogens but fail to achieve the required control of this malady. It is necessary to know the correct etiology of this disease to successfully devise management options against this disease. We report the existence of Pseudomonas oryzihabitans associated with grain discoloration, panicle blight, and chaffiness of rice from the Punjab region of North India for the first time. During Kharif 2021, panicle blight symptoms (disease incidence 20–25%, cultivar PR 114) were observed in rice fields of three districts. These blighted panicles were partially filled or grains were aborted causing chaffiness in many panicles. Yellow-colored smooth, circular colonies of bacteria were obtained from the diseased panicles. The associated bacterium was identified as a rod-shaped, gram-negative P. oryzihabitans based on biochemical characterization, Scanning Electron Microscope imaging, and sequencing of the 16S rDNA region along with housekeeping genes rpoD and gyrB. The pathogenicity of the bacterium was tested in the fields during Kharif 2021 and Kharif 2022.The study demonstrated that the bacterial pathogen, P. oryzihabitans should be considered as one of the cause of grain discoloration and chaffiness of rice in Punjab region of India and management options for bacterial grain discoloration, panicle blight and chaffiness should be worked out for effective management of this disease. • Rice is affected by grain discoloration and chaffiness of panicles during maturity of the crop. • Lack of effective management practices is a major concern among farmers. • Knowledge of correct etiology of this disease is the prerequisite to devise management options for this disease. • Pseudomonas oryzihabitans was found associated with grain discoloration, panicle blight, and chaffiness of rice from the Punjab region of North India. [ABSTRACT FROM AUTHOR]

Sigma factors are RNA polymerase subunits engaged in promoter recognition and DNA strand separation during transcription initiation in bacteria. Primary-sigma factors are responsible for the expression of housekeeping genes and are essential for survival. RpoD, the primary-sigma factor of Escherichia coli, a γ-proteobacteria, recognizes consensus promoter sequences highly similar to those of some α-proteobacteria species. Despite this resemblance, RpoD is unable to sustain transcription from most of the α-proteobacterial promoters tested so far. In contrast, we have found that SigA, the primary-sigma factor of Rhizobium etli, an α-proteobacteria, is able to transcribe E. coli promoters, although it exhibits only 48% identity (98% coverage) to RpoD. We have called this the transcriptional laxity phenomenon. Here, we show that SigA partially complements the thermo-sensitive deficiency of RpoD285 from E. coli strain UQ285 and that the SigA region σ4 is responsible for this phenotype. Sixteen out of 74 residues (21.6%) within region σ4 are variable between RpoD and SigA. Mutating these residues significantly improves SigA ability to complement E. coli UQ285. Only six of these residues fall into positions already known to interact with promoter DNA and to comprise a helix-turn-helix motif. The remaining variable positions are located on previously unexplored sites inside region σ4, specifically into the first two α-helices of the region. Neither of the variable positions confined to these helices seem to interact directly with promoter sequence; instead, we adduce that these residues participate allosterically by contributing to correct region folding and/or positioning of the HTH motif. We propose that transcriptional laxity is a mechanism for ensuring transcription in spite of naturally occurring mutations from endogenous promoters and/or horizontally transferred DNA sequences, allowing survival and fast environmental adaptation of α-proteobacteria.

DNA-based approaches were used to characterize a strain (Salento-1) of Xylella fastidiosa obtained from an olive plant suffering from the syndrome of quick decline in Apulia (South Italy). Salento-1 was indistinguishable from strain CoDiRO previously isolated from olive in Apulia and assigned to X. fastidiosa subsp. pauca. Based on our results and comparative analysis with reported data, the subspecies pauca, multiplex, and fastidiosa may invade olive throughout the world (California, Italy, Argentina and Brazil). The strain Salento-1 has been deposited in the National Collection of Plant Pathogenic Bacteria (NCPPB), England, and in the Belgian Coordinated Collections of Microorganisms (BCCM), Belgium.

Forty-five Xanthomonas arboricola pv. juglandis (Xaj) strains originating from Juglans regia cultivation in different countries were molecularly typed by means of MultiLocus Sequence Typing (MLST), using acnB, gapA, gyrB and rpoD gene fragments. A total of 2.5 kilobases was used to infer the phylogenetic relationship among the strains and possible recombination events. Haplotype diversity, linkage disequilibrium analysis, selection tests, gene flow estimates and codon adaptation index were also assessed. The dendrograms built by maximum likelihood with concatenated nucleotide and amino acid sequences revealed two major and two minor phylotypes. The same haplotype was found in strains originating from different continents, and different haplotypes were found in strains isolated in the same year from the same location. A recombination breakpoint was detected within the rpoD gene fragment. At the pathovar level, the Xaj populations studied here are clonal and under neutral selection. However, four Xaj strains isolated from walnut fruits with apical necrosis are under diversifying selection, suggesting a possible new adaptation. Gene flow estimates do not support the hypothesis of geographic isolation of the strains, even though the genetic diversity between the strains increases as the geographic distance between them increases. A triplet deletion, causing the absence of valine, was found in the rpoD fragment of all 45 Xaj strains when compared with X. axonopodis pv. citri strain 306. The codon adaptation index was high in all four genes studied, indicating a relevant metabolic activity.

[Display omitted] • Current gold standard in bacterial taxonomy is phylogenomics. • Fast, cheap, and taxonomically competent options, like MLSA, are recommendable. • We did an unbiased search of core gene regions with highest taxonomic value for Pseudomonas. • Internal regions of rpoD , pepN , and gltX are the most informative for the Pseudomonas genus. • The rpoD-pepN-gltX MLSA is a robust tool with phylogenetic power comparable to ANI. In the field of prokaryotic taxonomy, there has been a recent transition towards phylogenomics as the gold standard approach. However, genome-based phylogenetics is still restrictive for its cost when managing large amounts of isolates. Fast, cheap, and taxonomically competent alternatives, like multilocus sequence analysis (MLSA) are thus recommendable. Nevertheless, the criteria for selecting the conserved genes for MLSA have not been explicit for different bacterial taxa, including the broadly diverse Pseudomonas genus. Here, we have carried out an unbiased and rational workflow to select internal sequence regions of Pseudomonas core genes (CG) for a MLSA with the best phylogenetic power, and with a resolution comparable to the genome-based ANI approach. A computational workflow was established to inspect 126 complete genomes of representatives from over 60 Pseudomonas species and subspecies, in order to identify the most informative CG internal regions and determine which combinations in sets of three partial CG sequences have comparable phylogenetic resolution to that of the current ANI standard. We found that the rpoD 346-1196- pepN 1711-2571- gltX 86-909 concatenated sequences were the best performing in terms of phylogenetic robustness and resulted highly sensitive and specific when contrasted with ANI. The rpoD - pepN - gltX MLSA was validated in silico and in vitro. Altogether, the results presented here supports the proposal of the rpoD - pepN - gltX MLSA as a fast, affordable, and robust phylogenetic tool for members of the Pseudomonas genus. [ABSTRACT FROM AUTHOR]

Sigma factors are RNA polymerase subunits engaged in promoter recognition and DNA strand separation during transcription initiation in bacteria. Primary sigma factors are responsible for the expression of housekeeping genes and are essential for survival. RpoD, the primary sigma factor of Escherichia coli, a γ-proteobacteria, recognizes consensus promoter sequences highly similar to those of some α-proteobacteria species. Despite this resemblance, RpoD is unable to sustain transcription from most of the α-proteobacterial promoters tested so far. In contrast, we have found that SigA, the primary sigma factor of Rhizobium etli, an α-proteobacteria, is able to transcribe E. coli promoters, although it exhibits only 48% identity (98% coverage) to RpoD. We have called this the transcriptional laxity phenomenon. Here, we show that SigA partially complements the thermo-sensitive deficiency of RpoD285 from E. coli strain UQ285 and that the SigA region s4 is responsible for this phenotype. Sixteen out of 74 residues (21.6%) within region s4 are variable between RpoD and SigA. Mutating these residues significantly improves SigA ability to complement E. coli UQ285. Only six of these residues fall into positions already known to interact with promoter DNA and to comprise a helix-turn-helix motif. The remaining variable positions are located on previously unexplored sites inside region s4, specifically into the first two α-helices of the region. Neither of the variable positions confined to these helices seem to interact directly with promoter sequence; instead, we adduce that these residues participate allosterically by contributing to correct region folding and/or positioning of the HTH motif. We propose that transcriptional laxity is a mechanism for ensuring transcription in spite of naturally occurring mutations from endogenous promoters and/or horizontally transferred DNA sequences, allowing survival and fast environmental adaptation of α-proteobacteria. [ABSTRACT FROM AUTHOR]

Bioconversion processes of organic acid or acid hydrolysis of raw material for microbial metabolism often suffer limitations as a result of microbial sensitivity in low-pH conditions. We adopted a three-step method called RAndom Insertional-deletional Strand Exchange mutagenesis (RAISE) to engineer the components of global regulator Sigma D factor (RpoD) of Escherichia coli to improve its acid tolerance. The best strain Mutant VII was identified from random mutagenesis libraries based on the growth performance, which exhibited much higher growth rate than the control (0.22 h −1 vs. 0.15 h −1 ) at pH as low as 3.17. Combined transcriptome and phenome analysis of E. coli was carried out to better understand the global effects of RpoD on the regulatory networks. Our analysis showed that 95 (2.1%) of all E. coli genes were induced and 178 (4.0%) genes were repressed, including those for trehalose biosynthesis, nucleotides biosynthesis, carbon metabolism, amino acid utilization, except for acid resistance. Also regulated were the master regulators (ArcA, EvgA, H-NS and RpoS) and gene/operon-specific transcription factors (GadX, GadW, AppY, YdeO, KdgR). These results demonstrated that RpoD acts as global regulator in the growth phase of E. coli and consequently improves acid tolerances. [ABSTRACT FROM AUTHOR]

DNA-based approaches were used to characterize a strain (Salento-1) of Xylella fastidiosa obtained from an olive plant suffering from the syndrome of quick decline in Apulia (South Italy). Salento-1 was indistinguishable from strain CoDiRO previously isolated from olive in Apulia and assigned to X. fastidiosa subsp. pauca. Based on our results and comparative analysis with reported data, the subspecies pauca, multiplex, and fastidiosa may invade olive throughout the world (California, Italy, Argentina and Brazil). The strain Salento-1 has been deposited in the National Collection of Plant Pathogenic Bacteria (NCPPB), England, and in the Belgian Coordinated Collections of Microorganisms (BCCM), Belgium. [ABSTRACT FROM AUTHOR]

A major source of pseudomonad-specialized metabolites is the nonribosomal peptide synthetases (NRPSs) assembling siderophores and lipopeptides. Cyclic lipopeptides (CLPs) of the Mycin and Peptin families are frequently associated with, but not restricted to, phytopathogenic species. We conducted an in silico analysis of the NRPSs encoded by lipopeptide biosynthetic gene clusters in nonpathogenic Pseudomonas genomes, covering 13 chemically diversified families. This global assessment of lipopeptide production capacity revealed it to be confined to the Pseudomonas fluorescens lineage, with most strains synthesizing a single type of CLP. Whereas certain lipopeptide families are specific for a taxonomic subgroup, others are found in distant groups. NRPS activation domain-guided peptide predictions enabled reliable family assignments, including identification of novel members. Focusing on the two most abundant lipopeptide families (Viscosin and Amphisin), a portion of their uncharted diversity was mapped, including characterization of two novel Amphisin family members (nepenthesin and oakridgin). Using NMR fingerprint matching, known Viscosin-family lipopeptides were identified in 15 (type) species spread across different taxonomic groups. A bifurcate genomic organization predominates among Viscosin-family producers and typifies Xantholysin-, Entolysin-, and Poaeamide-family producers but most families feature a single NRPS gene cluster embedded between cognate regulator and transporter genes. The strong correlation observed between NRPS system phylogeny and rpoD -based taxonomic affiliation indicates that much of the structural diversity is linked to speciation, providing few indications of horizontal gene transfer. The grouping of most NRPS systems in four superfamilies based on activation domain homology suggests extensive module dynamics driven by domain deletions, duplications, and exchanges. IMPORTANCE Pseudomonas species are prominent producers of lipopeptides that support proliferation in a multitude of environments and foster varied lifestyles. By genome mining of biosynthetic gene clusters (BGCs) with lipopeptide-specific organization, we mapped the global Pseudomonas lipopeptidome and linked its staggering diversity to taxonomy of the producers, belonging to different groups within the major Pseudomonas fluorescens lineage. Activation domain phylogeny of newly mined lipopeptide synthetases combined with previously characterized enzymes enabled assignment of predicted BGC products to specific lipopeptide families. In addition, novel peptide sequences were detected, showing the value of substrate specificity analysis for prioritization of BGCs for further characterization. NMR fingerprint matching proved an excellent tool to unequivocally identify multiple lipopeptides bioinformatically assigned to the Viscosin family, by far the most abundant one in Pseudomonas and with stereochemistry of all its current members elucidated. In-depth analysis of activation domains provided insight into mechanisms driving lipopeptide structural diversification.

A Multilocus Sequence Analysis (MLSA) was performed on members of the Scophthalmi clade in the genus Vibrio , including type and reference strains of the species V. scophthalmi , V. ichthyoenteri , and 39 strains phenotypically identified as Vibrio ichthyoenteri-like , with the aim of better defining boundaries between these two closely related, fish-associated species. The type strain of V. ponticus , recently added to the clade Scophthalmi, was also included. The study was based on partial sequences of the protein-coding housekeeping genes rpoD , mreB , recA , ftsZ , and gyrB , and the 16S rRNA. While the 16S rRNA gene-based trees were unable to pull apart members of V. scophthalmi or V. ichthyoenteri , both the other individual gene trees and the trees obtained from the five-genes concatenated sequences were able to consistently differentiate four subclades within the main clade, corresponding to the bona fide V. scophthalmi , V. ichthyoenteri , and two small ones that may represent a new species each. The best genes to differentiate V. scophthalmi from V. ichthyoenteri were rpoD , recA , and mreB. Vibrio ponticus failed to associate to the clade in the MLSA and in most single gene trees for which it should not be considered part of it. In this study we also confirm using genomic indexes that V. ichthyoenteri and V. scophthalmi are two separate species. [ABSTRACT FROM AUTHOR]

Storage in ice is a common way of preserving commercial fish species but some microorganisms can still contaminate and participate in the spoilage of the product; therefore, identification of potential harmful microbes is important. Thirteen colonies were isolated from common carp (Cyprinus carpio) that had been stored in ice, whose phenotypic identification revealed that they belonged to the genera Aeromonas (n = 5) and Shewanella (n = 8). Molecular genotyping with ERIC-PCR showed clonality only among two of the five Aeromonas isolates and for two groups (n = 3; n = 2) of the eight Shewanella isolates. Sequencing the rpoD gene showed that four Aeromonas isolates belonged to the species Aeromonas salmonicida and one to A. sobria. Of the eight Shewanella, seven isolates cluster with Shewanella putrefaciens and one with Shewanella profunda in the 16S rRNA phylogenetic tree. However, analysis of the gyrB gene showed that these eight isolates could constitute a new species closely related to S. baltica. The Shewanella and A. salmonicida isolates produce off-odours and reduce trimethylamine oxide, indicating that they might contribute to the spoilage of the fish. [ABSTRACT FROM AUTHOR]

Background Aeromonas salmonicida is a major fish pathogen associated with mass mortalities in salmonid fish. In the present study, we applied In Vivo Induced Antigen Technology (IVIAT), a technique that relies on antibodies adsorbed against in vitro cultures of the pathogen, to a clinical isolate of A. salmonicida subsp. salmonicida. Results The results from IVIAT allowed identification of four proteins that were upregulated in the fish samples: A UDP-3-O-acyl-N-acetylglucosamine deacetylase, an RNA polymerase sigma factor D as well as TonB and a hypothetical protein. Subsequent investigations were performed using real-time PCR and cDNA synthesised from infected spleen, liver and anterior kidneys. These confirmed that the transcription level of each of these genes was significantly upregulated during the infection process compared to bacteria in vitro. Conclusions The present studied identified four genes that were upregulated during the infectious process and are likely to play a role in the virulence of A. salmonicida. Because these are antigenic they might constitute potential targets for the development of new vaccine as well as therapeutic agents. [ABSTRACT FROM AUTHOR]

In Summer and Autumn of 2010, blighted cucurbits suspected of bacterial infection were observed in the East Azarbayejan province of Iran. Infected plants exhibited spots on the leaves which, in the early stages of the disease, appeared as small, yellow and water-soaked with chlorotic haloes. As time went by, the spots enlarged, necrotized and coalesced so as to cover a large area of the blade, while the leaves rolled along their longitudinal axis. Such symptoms had not previously been observed in cucurbits in East Azarbayejan. To investigate the etiology of this disorder, symptomatic leaves were collected from affected plants and the putative agent was isolated and identified as Pseudomonas syringae pv. aptata based on morphological, physiological, biochemical, molecular and pathogenicity tests. In BOX-PCR, four bacterial isolates from cucurbits formed a coherent group and their sequences showed a high similarity (99%) with P. syringae pv. aptata GSPB1067, as confirmed by phylogenetic analysis based on gyrB and rpoD. To the best of our knowledge, this is the first report of leaf blight and necrosis of Cucurbitaceae caused by P. syringae pv. aptata in Iran. [ABSTRACT FROM AUTHOR]

Forty-eight Aeromonas isolates from rainbow trout previously identified by the 16S rDNA-RFLP technique were re-identified using 2 housekeeping genes (gyrB and rpoD). After sequencing the prevalences of the species were A. veronii (29.2%), A. bestiarum (20.8%), A. hydrophila (16.7%), A. sobria (10.4%), A. media (8.3%), A. popoffii (6.2%), A. allosaccharophila (2.1%), A. caviae (2.1%), A. salmonicida (2.1%) and one isolate (2.1%) belongs to a candidate new species "Aeromonas lusitana". Coincident identification results to the 16S rDNA-RFLP technique were only obtained for 68.8% of the isolates. PCR amplification of the enterobacterial repetitive intergenic consensus (ERIC-PCR) indicated that the 48 isolates belonged to 33 different ERIC genotypes. Several genotypes were isolated from different farms and organs in the same fish, indicating a systemic dissemination of the bacteria. The presence of genes (blaIMP, blaCphA/IMIS, blaTEM, blaSHV and intI1) that encode extended-spectrum beta-lactamases (ESBLs), metallo-beta-lactamases (MBLs) and class 1 integrons were studied by PCR. Only 39.6% (19/48) of the strains showed the presence of one or more resistance genes. The gene blaCphA/IMIS was detected in 29.2% of the isolates, followed by the intI1 (6.2%) and blaSHV (4.2%) genes. The variable region of class 1 integrons of the 3 positive isolates was sequenced revealing the presence of the gene cassette aadA1 (aminoglycoside transferase) that plays a role in streptomycin/spectinomycin resistance. [ABSTRACT FROM AUTHOR]

During Czech expeditions at James Ross Island, Antarctica, in the years 2007-2009, the bacterial diversity of the genus Pseudomonas was studied. Twelve fluorescent Pseudomonas strains were isolated from various samples and were subjected to a detailed taxonomic study. A polyphasic approach included genotypic and phenotypic analyses. The genotypic analysis involved sequencing of rrs, rpoB and rpoD genes, DNA-DNA hybridization (DDH) studies as well as manual ribotyping using HindIII endonuclease. The phenotypic characterization included conventional tests as well as biotyping using the Biolog system, protein profiling by SDS-PAGE, and MALDI-TOF MS analysis. Our taxonomic study revealed that all isolates belonged to the same Pseudomonas species with psychrotrophic growth not exceeding 37 °C. The cultures showed a unique position among the phylogenetically related pseudomonads. DDH experiment between the proposed type strain of the antarctic isolates and the closest neighbour P. arsenicoxydans CCM 8423 showed only 40.9-50.1 % similarity, thus confirming that the characterized strains do not belong to the P. arsenicoxydans species. According to the results obtained we propose the name P. prosekii sp. nov. for this novel Pseudomonas taxon with type strain AN/28/1 (=CCM 7990 and LMG 26867). [ABSTRACT FROM AUTHOR]

Recent taxonomic advances, based on biochemical and genotypic processes demonstrate that the plant pathogenic species Pseudomonas cichorii consists of a cluster of closely related genomic groups. Prior to this study, three morphotype groups had been described (C1-C3), all sharing various phenotypic and biochemical characters but partially differing in their DNA content. All entities of the complex could cause disease among a variety of hosts, including lettuce, celery, chrysanthemum and others. In this study, we present the biochemical and molecular characterization of P. cichorii isolates as the causal agent of pith necrosis of tomato plants. A detailed characterization of the genetic variability among strains belonging to P. cichorii was achieved using BOX-PCR and Multi Locus Sequence Analysis utilizing three housekeeping genes ( gyrB, rpoD, rpoB). In addition, a number of biochemical and physiological tests were also used for the identification of the tomato P. cichorii isolates. To our knowledge, this is the first complete biochemical, molecular and phylogenetic study of P. cichorii strains isolated from tomato plants affected by pith necrosis disease. Our findings demonstrate the emergence of a new genomovar of P. cichorii, yet another indication for the genetic heterogeneity of the species. [ABSTRACT FROM AUTHOR]

Abstract: Seventy Aeromonas strains were identified by phylogenetic analysis using housekeeping genes (gyrB and rpoD) in order to investigate etiological agents for aeromoniasis in farmed eels (Anguilla japonica). The phylogenetic analysis showed that Aeromonas aquariorum (n =22, 31.4%) was the predominant species among the investigated eel strains, followed by Aeromonas caviae (n =16, 22.9%), A. veronii (n =13, 18.6%), A. hydrophila (n =12, 17.1%), A. jandaei (n =4, 5.7%), A. media (n =2, 2.9%), and A. trota (n =1, 1.4%). The potential virulence of the present strains was estimated by performing PCR assays using the following seven virulence genes: cytotoxic enterotoxin (act), two cytotonic enterotoxins (alt and ast), glycerophospholipid:cholesterol acyltransferase (gcaT), DNase (exu), lipase (lip), and flagellin (fla). The detection rates of act, alt, ast, gcaT, exu, lip, and fla among all 70 strains were 91.4%, 55.7%, 27.1%, 97.1%, 95.7%, 100%, and 98.6%, respectively. In genotyping of enterotoxin genes, act +/alt +/ast +, act +/alt +/ast −, and act +/alt −/ast − genotypes were prevalent in A. hydrophila (8/12 strains), A. aquariorum (13/22 strains), and A. caviae (14/16 strains), respectively, suggesting a high heterogeneity among Aeromonas species. In this study, A. aquariorum, which has been an unrecorded species in Korea, can be an etiological agent for aeromoniasis of eel. [Copyright &y& Elsevier]

Abstract: A genetic reidentification of 7 Aeromonas blood isolates revealed that phenotype-based identification systems misidentified 5 (71.4%) isolates. In Aeromonas strains, A. aquariorum was the most common misidentified organism and showed the most potent cytotoxic activities against human blood cell lines, suggesting that the correct identification of A. aquariorum is important. [Copyright &y& Elsevier]