Pseudomonas protegens Affects Mosquito Survival and Development.

This study investigated the pathogenic potential of Pseudomonas protegens on mosquito larvae of the two species Culex pipiens and Aedes albopictus, representing major threats for disease transmission in the Mediterranean area and worldwide. The bacterium achieved to kill over 90% of the mosquito larvae within 72 h after exposition to a bacterial concentration of 100 million CFU/ml. These lethal effects were concentration dependent and a significantly higher susceptibility was associated with younger larvae of both mosquito species. Significant slowdown of immature (larval and pupal) development and decrease in adult emergence rate after treatment with sub-lethal doses of the bacterium were also detected. This study reports for the first time the insecticidal activity of a root-associated biocontrol bacterium against aquatic mosquito larvae.

Clinical risk factors for admission with Pseudomonas and multidrug-resistant Pseudomonas community-acquired pneumonia.

BACKGROUND: Microbial etiology for community-acquired pneumonia (CAP) is evolving with pathogens known for high CAP mortality e.g., Pseudomonas species. Chronic obstructive pulmonary disease (COPD) patients are at risk for hospitalization for CAP. Understanding regional patterns and risk factors for multidrug-resistant (MDR) Pseudomonas acquisition has implications for antimicrobial stewardship. OBJECTIVES: To evaluate the regional epidemiology of MDR Pseudomonas CAP and its association with COPD. METHODS: We queried the electronic medical records of the University of Alabama at Birmingham Healthcare System to identify patients hospitalized for CAP with Pseudomonas positive respiratory samples between 01/01/2013-12/31/2019. Log binomial regression models were used to examine associations between COPD diagnosis and risk of Pseudomonas/MDR Pseudomonas CAP. RESULTS: Cohort consisted of 913 culture positive CAP cases aged 59-year (IQR:48-68), 61% (560) male, 60% (547) white, 65% (580) current/past smokers, and 42% (384) COPD. Prevalence of Pseudomonas CAP in culture positive CAP was 18% (167), MDR Pseudomonas CAP in Pseudomonas CAP was 22% (36), and yearly incidence of MDR Pseudomonas CAP was stable (p = 0.169). COPD was associated with Pseudomonas CAP (RR 1.39; 95% CI 1.01, 1.91; p = 0.041) but not with MDR Pseudomonas CAP (0.71; 95% CI 0.35, 1.45; p = 0.349). Stroke (RR 2.64; 95% CI 1.51, 4.61; p = 0.0006) and use of supplemental oxygen (RR 2.31; 95% CI 1.30, 4.12; p = 0.005) were associated with MDR Pseudomonas CAP. CONCLUSION: Incidence of MDR Pseudomonas CAP was stable over time. COPD was associated with Pseudomonas CAP but not with MDR Pseudomonas CAP. Larger cohort studies are needed to confirm findings.

Complete genome sequence of the kiwifruit bacterial canker pathogen Pseudomonas savastanoi strain MHT1.

BACKGROUND: Pseudomonas savastanoi is an important plant pathogen that infects and causes symptoms in a variety of economically important crops, causing considerable loss of yield and quality. Because there has been no research reported to date on bacterial canker of kiwifruit (Actinidia chinensis) plants caused by P. savastanoi and, in particular, no in-depth studies of the complete genome sequence or pathogenic mechanism, long-lasting and environmentally friendly control measures against this pathogen in kiwifruit are lacking. This study therefore has both theoretical value and practical significance. RESULTS: We report the complete genome sequence of P. savastanoi strain MHT1, which was first reported as the pathogen causing bacterial canker in kiwifruit plants. The genome consists of a 6.00-Mb chromosome with 58.5% GC content and 5008 predicted genes. Comparative genome analysis of four sequenced genomes of representative P. savastanoi strains revealed that 230 genes are unique to the MHT1 strain and that these genes are enriched in antibiotic metabolic processes and metabolic pathways, which may be associated with the drug resistance and host range observed in this strain. MHT1 showed high syntenic relationships with different P. savastanoi strains. Furthermore, MHT1 has eight conserved effectors that are highly homologous to effectors from P. syringae, Pseudomonas amygdali, and Ralstonia solanacearum strains. The MHT1 genome contains six genomic islands and two prophage sequences. In addition, 380 genes were annotated as antibiotic resistance genes and another 734 as encoding carbohydrate-active enzymes. CONCLUSION: The whole-genome sequence of this kiwifruit bacterial canker pathogen extends our knowledge of the P. savastanoi genome, sets the stage for further studies of the interaction between kiwifruit and P. savastanoi, and provides an important theoretical foundation for the prevention and control of bacterial canker.

Immune responses and inorganic ion transport regulations of Epinephelus coioides in response to L321_RS13075 gene of Pseudomonas plecoglossicida.

Pseudomonas plecoglossicida is a well-known pathogen of viscera granulomas disease in fish, which has led to severe economic losses. In our previous study, L321_RS13075 was predicted to be a key virulence gene of P. plecoglossicida during the host-pathogen interaction with Epinephelus coioides. To investigate the role of L321_RS13075 in the regulation of virulence in P. plecoglossicida, a L321_RS13075 knock-down strain was constructed. And a significant reduction in the ability of colonization, intracellular survival, motility, biofilm formation, and adhesion was detected in the L321_RS13075 knock-down strain. Compared with the wild-type strain, the silence of L321_RS13075 in P. plecoglossicida resulted in a significant change in the transcriptome of infected Epinephelus coioides (E. coioides). Results of COG and GO analysis on E. coioides showed that genes related to immune responses and inorganic ion transport were significantly affected by L321_RS13075 of P. plecoglossicida. Meanwhile, the interactions of the genes related to immune responses and inorganic ion transport were predicted, and the important hub genes were identified. Taken together, the results indicated that L321_RS13075 was a virulent gene of P. plecoglossicida, which significantly affected the immune responses and inorganic ion transport in E. coioides.

The contribution of exbB gene to pathogenicity of Pseudomonas plecoglossicida and its interactions with Epinephelus coioides.

To study the roles of the exbB gene in Pseudomonas plecoglossicida during interactions with Epinephelus coioides, five short hairpin RNAs (shRNAs) were designed and synthesized to silence the exbB gene in P. plecoglossicida which resulted in significant reductions in exbB mRNA expression. The mutant with the best silencing efficiency (89.3%) was selected for further study. Silencing exbB in the exbB-RNA interference (RNAi) strain resulted in a 70% increase in the survival rate and a 3-day delay in the onset of infection in E. coioides. Silencing of the exbB gene also resulted in a significant decrease in the number of white spots on the spleen surface and in the spleen pathogen load. The results of dual RNA-seq showed that exbB silencing in P. plecoglossicida also resulted in a significant change in both the pathogen and host transcriptomes in the spleens of infected E. coioides. Comparative transcriptome analysis showed that silencing exbB caused significant changes in multiple signaling molecules and interaction- and immune system-related genes in E. coioides. Gene silencing also resulted in the differential expression of flagellar assembly and the bacterial secretion system in P. plecoglossicida during the infection period, and most of the DEGs were down-regulation. These host-pathogen interactions may make it easier for E. coioides to eliminate the exbB-RNAi strain of P. plecoglossicida, suggesting a significant decrease in the pathogenicity of this strain. These results indicated that exbB was a virulence gene of P. plecoglossicida which contributed a lot in the pathogen-host interactions with E. coioides.

Occurrence of Pseudomonas spp. in Raw Vegetables: Molecular and Phenotypical Analysis of Their Antimicrobial Resistance and Virulence-Related Traits.

Pseudomonas is characterized by its great capacity to colonize different ecological niches, but also by its antimicrobial resistance and pathogenicity, causing human, animal, or plant diseases. Raw and undercooked food is a potential carrier of foodborne disease. The aim of this study was to determine the occurrence of Pseudomonas spp. among raw vegetables, analysing their antimicrobial resistance, virulence, and molecular typing. A total of 163 Pseudomonas spp. isolates (12 different species) were recovered from 77 of the 145 analysed samples (53.1%) and were classified into 139 different pulsed-field gel electrophoresis patterns. Low antimicrobial resistance levels, but one multidrug-resistant isolate, were found. Among the 37 recovered P. aeruginosa strains, 28 sequence-types and nine serotypes were detected. Eleven OprD patterns and an insertion sequence (ISPa1635) truncating the oprD gene of one imipenem-resistant strain were found. Ten virulotypes were observed, including four exoU-positive and thirty-one exoS-positive strains. The lasR gene was absent in three ST155 strains and was truncated by different insertion sequences (ISPre2, IS1411, and ISPst7) in other three strains. High biofilm, motility, pigment, elastase, and rhamnolipid production were detected. Our study demonstrated a low occurrence of P. aeruginosa (18%) and low antimicrobial resistance, but a high number of virulence-related traits in these P. aeruginosa strains, highlighting their pathological importance.

Multilocus sequence analysis reveals different lineages of Pseudomonas anguilliseptica associated with disease in farmed lumpfish (Cyclopterus lumpus L.).

The bacterium Pseudomonas anguilliseptica has in recent years emerged as a serious threat to production of lumpfish in Norway. Little is known about the population structure of this bacterium despite its association with disease in a wide range of different fish species throughout the world. The phylogenetic relationships between 53 isolates, primarily derived from diseased lumpfish, but including a number of reference strains from diverse geographical origins and fish species, were reconstructed by Multi-Locus Sequence Analysis (MLSA) using nine housekeeping genes (rpoB, atpD, gyrB, rpoD, ileS, aroE, carA, glnS and recA). MLSA revealed a high degree of relatedness between the studied isolates, altough the seven genotypes identified formed three main phylogenetic lineages. While four genotypes were identified amongst Norwegian lumpfish isolates, a single genotype dominated, irrespective of geographic origin. This suggests the existence of a dominant genotype associated with disease in production of lumpfish in Norwegian aquaculture. Elucidation of the population structure of the bacterium has provided valuable information for potential future vaccine development.

Pseudomonas donghuensis HYS gtrA/B/II Gene Cluster Contributes to Its Pathogenicity toward Caenorhabditis elegans.

Pseudomonas donghuensis HYS is more virulent than P. aeruginosa toward Caenorhabditis elegans but the mechanism underlying virulence is unclear. This study is the first to report that the specific gene cluster gtrA/B/II in P. donghuensis HYS is involved in the virulence of this strain toward C. elegans, and there are no reports of GtrA, GtrB and GtrII in any Pseudomonas species. The pathogenicity of P. donghuensis HYS was evaluated using C. elegans as a host. Based on the prediction of virulence factors and comparative genomic analysis of P. donghuensis HYS, we identified 42 specific virulence genes in P. donghuensis HYS. Slow-killing assays of these genes showed that the gtrAB mutation had the greatest effect on the virulence of P. donghuensis HYS, and GtrA, GtrB and GtrII all positively affected P. donghuensis HYS virulence. Two critical GtrII residues (Glu47 and Lys480) were identified in P. donghuensis HYS. Transmission electron microscopy (TEM) showed that GtrA, GtrB and GtrII were involved in the glucosylation of lipopolysaccharide (LPS) O-antigen in P. donghuensis HYS. Furthermore, colony-forming unit (CFU) assays showed that GtrA, GtrB and GtrII significantly enhanced P. donghuensis HYS colonization in the gut of C. elegans, and glucosylation of LPS O-antigen and colonization in the host intestine contributed to the pathogenicity of P. donghuensis HYS. In addition, experiments using the worm mutants ZD101, KU4 and KU25 revealed a correlation between P. donghuensis HYS virulence and the TIR-1/SEK-1/PMK-1 pathways of the innate immune p38 MAPK pathway in C. elegans. In conclusion, these results reveal that the specific virulence gene cluster gtrA/B/II contributes to the unique pathogenicity of HYS compared with other pathogenic Pseudomonas, and that this process also involves C. elegans innate immunity. These findings significantly increase the available information about GtrA/GtrB/GtrII-based virulence mechanisms in the genus Pseudomonas.

Integration of RNA-seq and RNAi reveals the contribution of znuA gene to the pathogenicity of Pseudomonas plecoglossicida and to the immune response of Epinephelus coioides.

Pseudomonas plecoglossicida is an important pathogen in aquaculture and causes serious economic losses. Our previous study indicated that znuA gene might play an important role in the pathogenicity of P. plecoglossicida. Five shRNAs were designed and synthesized to silence the znuA gene of P. plecoglossicida. Two of the five mutants of P. plecoglossicida exhibited significant reduction in the expression level of znuA mRNA with different efficiencies. The mutant with the highest silencing efficiency of 89.2% was chosen for further studies. Intrapleural injection of the znuA-RNAi strain at a dose of 105  cfu/fish did not cause the death of Epinephelus coioides, and no significant signs were observed at the spleen surface of infected E. coioides, while the counterpart E. coioides infected by the same dose of wild-type strain of P. plecoglossicida all died in 5 days post-infection (dpi). The expression of znuA gene of znuA-RNAi strain in E. coioides was always lower than that in wild-type strain of P. plecoglossicida. The pathogen load in the early stage of infection was higher than that in the later stage of infection. Although the infection of the znuA-RNAi strain of P. plecoglossicida could induce the production of antibodies in E. coioides, it failed to produce a good immune protection against the infection of wild-type strain of P. plecoglossicida. Compared with the transcriptome data of E. coioides infected by the wild-type strain of P. plecoglossicida, the transcriptome data of E. coioides infected by the znuA-RNAi strain of P. plecoglossicida have altered significantly. Among them, KEGG enrichment analysis showed that the focal adhesion pathway was significantly enriched and exhibited the largest number of 302 DEMs (differentially expressed mRNAs). These results showed that the immune response of E. coioides to P. plecoglossicida infection was significantly affected by the RNAi of znuA gene.

Preliminary studies on the different roles of T6SSs in pathogenicity of Pseudomonas plecoglossicida NB2011.

Pseudomonas plecoglossicida, the causative agent of visceral granulomas in the large yellow croaker (Larimichthys crocea) in China, encodes three sets of type Ⅵ secretion systems (T6SS1-3). The purpose of this study was to characterize the different roles of T6SSs involved in infection. In-frame deletion of T6SSs was constructed, which resulted in 8 mutants. Competition against E. coli DH5α, virulence against the croaker and in vivo survival ability of the mutants were tested. The expression and secretion of Hcp by P. plecoglossicida NB2011 were investigated. The results showed T6SS2 mutant failed to inhibit the growth of E. coli, which is an indication of T6SS2 acting against environmental bacteria. The LD50 value of T6SS1 mutant strongly increased; T6SS2 and T6SS3 mutants were similar to that of the wild type; and the virulence of double deletion or triple deletion mutant was drastically alleviated, indicating that T6SS1 being one of the major virulence factors, and T6SS2 and T6SS3 directly or indirectly being involved in the pathogenicity. T6SS1 mutant disappeared in the fish spleen in 3 days, while other strains kept increasing, indicating the T6SS1 stimulation bacteria replication in vivo. Hcp1 secreted at 12-28°C and Hcp2 secreted at 12-35°C, while Hcp3 secretion not detected in vitro. This study has thrown some insights on the understanding of pathogenicity mechanisms of this pathogen.

Characterization and functional study of a chimera galectin from yellow drum Nibea albiflora.

Galectins are protein that participates in a variety of immune responses in the process of pathogenic infections. In the present study, a chimera galectin gene was screened from the transcriptome database of Nibea albiflora, which was named as YdGal-3. The results of qRT-PCR showed that the mRNA transcripts of YdGal-3 were ubiquitously distributed in all the detected tissues. After infection with Vibrio harveyi, the expression of YdGal-3 in liver, spleen, and head kidney increased significantly. Immunohistochemistry showed that YdGal-3 protein was widely expressed in the head kidney. The purified YdGal-3 protein by prokaryotic expression agglutinated red blood cells. Sugar inhibition assay showed that the agglutinating activity of YdGal-3 protein was inhibited by different sugars including lactose, D-galactose, and lipopolysaccharide. In addition, we mutated YdGal-3 His 294 into proline (P), alanine (A), glycine (G), and aspartic acid (D), it was further proved that the residue plays a key role in agglutination. YdGal-3 agglutinated some gram-negative bacteria including Pseudomonas plecoglossicida, Vibrio parahemolyticus, V. harveyi, and Aeromonas hydrophila, and exhibited antibacterial activity. These results suggested that YdGal-3 protein played an important role in the innate immunity of N. albiflora.

mSphere of Influence: Forgotten Questions.

Laura-Isobel McCall studies the relationship between location and disease pathogenesis, with a focus on infectious diseases and neglected diseases of poverty. In this mSphere of Influence article, she reflects on how three papers, "Opposing effects of fasting metabolism on tissue tolerance in bacterial and viral inflammation" (A. Wang, S. C. Huen, H. H. Luan, S. Yu, et al., Cell 166:1512-1525.e12, 2016, https://doi.org/10.1016/j.cell.2016.07.026), "Three-dimensional microbiome and metabolome cartography of a diseased human lung" (N. Garg, M. Wang, E. Hyde, R. R. da Silva, et al., Cell Host Microbe 22:705-716.e4, 2017, https://doi.org/10.1016/j.chom.2017.10.001), and "'It's like a phantom disease': patient perspectives on access to treatment for Chagas disease in the United States" (C. J. Forsyth, S. Hernandez, C. A. Flores, M. F. Roman, et al., Am J Trop Med Hyg 98:735-741, 2018, https://doi.org/10.4269/ajtmh.17-0691), shaped her spatial approach to infectious disease pathogenesis and helped her broaden her perspective from a pathogen-centric focus to a holistic view that include diseases tolerance mechanisms and barriers to health care access.

Neuronal mitochondrial dynamics coordinate systemic mitochondrial morphology and stress response to confer pathogen resistance in C. elegans.

Mitochondrial functions across different tissues are regulated in a coordinated fashion to optimize the fitness of an organism. Mitochondrial unfolded protein response (UPRmt) can be nonautonomously elicited by mitochondrial perturbation in neurons, but neuronal signals that propagate such response and its physiological significance remain incompletely understood. Here, we show that in C. elegans, loss of neuronal fzo-1/mitofusin induces nonautonomous UPRmt through multiple neurotransmitters and neurohormones, including acetylcholine, serotonin, glutamate, tyramine, and insulin-like peptides. Neuronal fzo-1 depletion also triggers nonautonomous mitochondrial fragmentation, which requires autophagy and mitophagy genes. Systemic activation of UPRmt and mitochondrial fragmentation in C. elegans via perturbing neuronal mitochondrial dynamics improves resistance to pathogenic Pseudomonas infection, which is supported by transcriptomic signatures of immunity and stress-response genes. We propose that C. elegans surveils neuronal mitochondrial dynamics to coordinate systemic UPRmt and mitochondrial connectivity for pathogen defense and optimized survival under bacterial infection.

The Zinc Nutritional Immunity of Epinephelus coioides Contributes to the Importance of znuC During Pseudomonas plecoglossicida Infection.

Previously, the dual RNA-seq was carried out in a Pseudomonas plecoglossicida- Epinephelus coioides infection model to investigate the dynamics of pathogen-host interplay in vivo. ZnuC, a member of ZnuCBA Zn importer, was found transcriptionally up-regulated during infection. Thus, this study aimed to assess its role during the trade-off for Zn between host and P. plecoglossicida. ICP-MS analysis and fluorescent staining showed that Zn was withheld from serum and accumulated in the spleen, with increased Zn uptake in the Golgi apparatus of macrophages after infection. Additionally, growth assay, macrophage infection and animal infection after gene knockout / silencing revealed that znuC was necessary for growth in Zn-limiting conditions, colonization, intracellular viability, immune escape and virulence of P. plecoglossicida. Further analysis with dual RNA-seq revealed associations of host's Zn nutritional immunity genes with bacterial Zn assimilation genes. IL6 and ZIP4 played key roles in this network, and markedly affected znuB expression, intracellular viability and immune escape, as revealed by gene silencing. Moreover, EMSA and GFP reporter gene analysis showed that Fur sensed changes in Fe concentration to regulate znuCBA in P. plecoglossicida. Jointly, these findings suggest a trade-off for Zn between host and P. plecoglossicida, while ZnuC is important for P. plecoglossicida Zn acquisition.

A Breach in Plant Defences: Pseudomonas syringae pv. actinidiae Targets Ethylene Signalling to Overcome Actinidia chinensis Pathogen Responses.

Ethylene interacts with other plant hormones to modulate many aspects of plant metabolism, including defence and stomata regulation. Therefore, its manipulation may allow plant pathogens to overcome the host's immune responses. This work investigates the role of ethylene as a virulence factor for Pseudomonas syringae pv. actinidiae (Psa), the aetiological agent of the bacterial canker of kiwifruit. The pandemic, highly virulent biovar of this pathogen produces ethylene, whereas the biovars isolated in Japan and Korea do not. Ethylene production is modulated in planta by light/dark cycle. Exogenous ethylene application stimulates bacterial virulence, and restricts or increases host colonisation if performed before or after inoculation, respectively. The deletion of a gene, unrelated to known bacterial biosynthetic pathways and putatively encoding for an oxidoreductase, abolishes ethylene production and reduces the pathogen growth rate in planta. Ethylene production by Psa may be a recently and independently evolved virulence trait in the arms race against the host. Plant- and pathogen-derived ethylene may concur in the activation/suppression of immune responses, in the chemotaxis toward a suitable entry point, or in the endophytic colonisation.

Evolution of pathogen-specific improved survivorship post-infection in populations of Drosophila melanogaster adapted to larval crowding.

The environment experienced by individuals during their juvenile stages has an impact on their adult stages. In holometabolous insects like Drosophila melanogaster, most of the resource acquisition for adult stages happens during the larval stages. Larval-crowding is a stressful environment, which exposes the larvae to scarcity of food and accumulation of toxic waste. Since adult traits are contingent upon larval stages, in larval-crowding like conditions, adult traits are prone to get affected. While the effect of resource limited, poor-developmental environment on adult immune response has been widely studied, the effect of adaptation to resource-limited developmental environment has not been studied, therefore in this study we assayed the evolution of ability to survive infection in adult stages as a correlated response to adaptation to larval crowding environments. Using four populations of Drosophila melanogaster adapted to larval crowding for 240 generations and their respective control populations, we show that populations adapted to larval crowding show an improved and evolved post-infection survivorship against a gram-negative bacteria Pseudomonas entomophila. Whereas, against a gram-positive bacteria Enterococcus faecalis, no difference in post-infection survivorship was observed across control and selected populations. In this study, we report the co-related evolution of pathogen-specific increased survivorship post-infection in populations of Drosophila melanogaster as a result of adaptation to larval crowding environment.

Causative bacteria associated with a clinically relevant postoperative pancreatic fistula infection after distal pancreatectomy.

PURPOSE: Clinically relevant postoperative pancreatic fistulas (CR-POPF) occurring after distal pancreatectomy often cause intra-abdominal infections. We monitored the presence of bacterial contamination in the ascitic fluid after distal pancreatectomy to clarify the bacterial origin of intra-abdominal infections associated with CR-POPF. METHODS: In 176 patients who underwent distal pancreatectomy, ascitic fluid bacterial cultures were performed on postoperative days (POD) 1-4 and when the drainage fluid became turbid. The association between postoperative ascitic bacterial contamination and CR-POPF incidence was investigated. RESULTS: CR-POPF occurred in 18 cases (10.2%). Among the patients with CR-POPF, bacterial contamination was detected in 0% on POD 1, in 38.9% on POD 4, and in 72.2% on the day (median, day 9.5) when the drainage fluid became turbid. A univariate analysis revealed a significant difference in ascitic bacterial contamination on POD 4 (p < 0.001) and amylase level on POD 3-4 (p < 0.001). A multivariate analysis revealed the amylase level and ascitic bacterial contamination on POD 4 to be independent risk factors. CONCLUSIONS: In the CR-POPF group, ascitic bacterial contamination was not observed in the early postoperative stage, but the bacterial contamination rate increased after pancreatic juice leakage occurred. Therefore, CR-POPF-related infections in distal pancreatectomy may be caused by a retrograde infection of pancreatic juice.

High fluoride resistance and virulence profile of environmental Pseudomonas isolated from water sources.

In our previous study, all Pseudomonas strains THP6, THP41, and OHP5 were identified as fluoride-resistant bacteria isolated from Dindigul district, Tamilnadu, India. The selected strains exhibiting a high level of fluoride resistance was determined in Luria broth (LB) medium and LB agar plates. In a further effort, fluoride-resistant organisms were tested for hemolytic activity and showed ß-hemolysis on blood agar plates. The virulence factors such as gyrB, toxA, algD and lasB, plcH, rhlC and biofilm response genes (pslA, pelA, ppyR) were detected by PCR analysis. The putative genus-specific and species-specific PCR also confirmed that the selected fluoride-resistant strains were belonging to Pseudomonas aeruginosa species. Fluoride-resistance gene crcB was amplified by gene-specific primers. The crcB gene was cloned in TA vector and transformed into E. coli DH5α. Comparative and blast analysis of THP6, THP41, and OHP5 strains crcB gene sequences were high homology with P. aeruginosa fluoride efflux transporter crcB and P. aeruginosa putative fluoride ion transporter crcB. The recombinants were efficiently growing in the NaF containing LB agar plates. The fluoride tolerance of these strains was also associated with resistance to multiple antibiotics. These results can lead to the use of the fluoride resistance gene of P. aeruginosa for the development of a biosensor for fluoride detection.

Pseudomonas savastanoi pv. mandevillae pv. nov., a Clonal Pathogen Causing an Emerging, Devastating Disease of the Ornamental Plant Mandevilla spp.

Commercial production of the ornamental plant dipladenia (Mandevilla spp.) is threatened by dipladenia leaf and stem spot disease, caused by the bacterium Pseudomonas savastanoi. P. savastanoi includes four pathovars of woody hosts differentiated by a characteristic host range in olive, oleander, ash, and broom plants. However, isolates from dipladenia have not been ascribed to any particular lineage or P. savastanoi pathovar. Here we report that isolates from dipladenia represent a distinct, clonal lineage. First, dipladenia isolates display very similar plasmid profiles, including a plasmid encoding the iaaM gene for biosynthesis of indole-3-acetic acid. Second, multilocus sequence analysis and core genome single-nucleotide polymorphisms phylogenies showed a monophyletic origin for dipladenia isolates, which cluster with isolates from oleander (pathovar nerii) in a distinct clade well separated from other P. savastanoi strains. Metabolic profiling and cross-pathogenicity tests in olive, oleander, ash, broom, and dipladenia clearly distinguished dipladenia isolates from the four P. savastanoi pathovars. Comparative genomics of the draft genome sequence of the dipladenia strain Ph3 with the other four pathovars showed that Ph3 encodes very few strain-specific genes and a similar set of virulence genes to pv. nerii, including its repertoire of type III secretion system effectors. However, hierarchical clustering based on the catalog of effectors and their allelic variants clearly separated Ph3 from pv. nerii strains. Based on their distinctive pathogenicity profile, we propose a de novo pathovar for P. savastanoi isolates from dipladenia, P. savastanoi pv. mandevillae pv. nov., for which strain Ph3 (CFBP 8832PT) has been designated as the pathotype strain.