Huanglongbing (HLB), also known as citrus greening, is a lethal disease of citrus caused by several species of 'Candidatus Liberibacter', a psyllid-transmitted, phloem-limited, alpha proteobacteria. 'Ca. Liberibacter asiaticus' is widespread in Florida citrus. The recently published 'Ca. L. asiaticus' psy62 genome, derived from a psyllid, revealed a prophage-like region of DNA in the genome, but phage have not been associated with 'Ca. L. asiaticus' to date. In the present study, shotgun sequencing and a fosmid DNA library of curated 'Ca. L. asiaticus' UF506, originally derived from citrus symptomatic for HLB, revealed two largely homologous, circular phage genomes, SC1 and SC2. SC2 encoded putative adhesin and peroxidase genes that had not previously been identified in 'Ca. L. asiaticus' and which may be involved in lysogenic conversion. SC2 also appeared to lack lytic cycle genes and replicated as a prophage excision plasmid, in addition to being found integrated in tandem with SC1 in the UF506 chromosome. By contrast, SC1 carried suspected lytic cycle genes and was found in nonintegrated, lytic cycle forms only in planta. Phage particles associated with 'Ca. L. asiaticus' were found in the phloem of infected periwinkles by transmission electron microscopy. In psyllids, both SC1 and SC2 were found only as prophage.

An 8x draft genome was obtained and annotated for Ralstonia solanacearum race 3 biovar 2 (R3B2) strain UW551, a United States Department of Agriculture Select Agent isolated from geranium. The draft UW551 genome consisted of 80,169 reads resulting in 582 contigs containing 5,925,491 base pairs, with an average 64.5% GC content. Annotation revealed a predicted 4,454 protein coding open reading frames (ORFs), 43 tRNAs, and 5 rRNAs; 2,793 (or 62%) of the ORFs had a functional assignment. The UW551 genome was compared with the published genome of R. solanacearum race 1 biovar 3 tropical tomato strain GMI1000. The two phylogenetically distinct strains were at least 71% syntenic in gene organization. Most genes encoding known pathogenicity determinants, including predicted type III secreted effectors, appeared to be common to both strains. A total of 402 unique UW551 ORFs were identified, none of which had a best hit or >45% amino acid sequence identity with any R. solanacearum predicted protein; 16 had strong (E < 10(-13)) best hits to ORFs found in other bacterial plant pathogens. Many of the 402 unique genes were clustered, including 5 found in the hrp region and 38 contiguous, potential prophage genes. Conservation of some UW551 unique genes among R3B2 strains was examined by polymerase chain reaction among a group of 58 strains from different races and biovars, resulting in the identification of genes that may be potentially useful for diagnostic detection and identification of R3B2 strains. One 22-kb region that appears to be present in GMI1000 as a result of horizontal gene transfer is absent from UW551 and encodes enzymes that likely are essential for utilization of the three sugar alcohols that distinguish biovars 3 and 4 from biovars 1 and 2.

Ralstonia solanacearum genes that are induced during tomato infection suggested that this pathogen encounters reactive oxygen species (ROS) during bacterial wilt pathogenesis. The genomes of R. solanacearum contain multiple redundant ROS-scavenging enzymes, indirect evidence that this pathogen experiences intense oxidative stress during its life cycle. Over 9% of the bacterium's plant-induced genes were also upregulated by hydrogen peroxide in culture, suggesting that oxidative stress may be linked to life in the plant host. Tomato leaves infected by R. solanacearum contained hydrogen peroxide, and concentrations of this ROS increased as pathogen populations increased. Mutagenesis of a plant-induced predicted peroxidase gene, bcp, resulted in an R. solanacearum strain with reduced ability to detoxify ROS in culture. The bcp mutant caused slightly delayed bacterial wilt disease onset in tomato. Moreover, its virulence was significantly reduced on tobacco plants engineered to overproduce hydrogen peroxide, demonstrating that Bcp is necessary for detoxification of plant-derived hydrogen peroxide and providing evidence that host ROS can limit the success of this pathogen. These results reveal that R. solanacearum is exposed to ROS during pathogenesis and that it has evolved a redundant and efficient oxidative stress response to adapt to the host environment and cause disease.

The plant pathogen Ralstonia solanacearum, which causes bacterial wilt disease, is exposed to reactive oxygen species (ROS) during tomato infection and expresses diverse oxidative stress response (OSR) genes during midstage disease on tomato. The R. solanacearum genome predicts that the bacterium produces multiple and redundant ROS-scavenging enzymes but only one known oxidative stress response regulator, OxyR. An R. solanacearum oxyR mutant had no detectable catalase activity, did not grow in the presence of 250 M hydrogen peroxide, and grew poorly in the oxidative environment of solid rich media. This phenotype was rescued by the addition of exogenous catalase, suggesting that oxyR is essential for the hydrogen peroxide stress response. Unexpectedly, the oxyR mutant strain grew better than the wild type in the presence of the superoxide generator paraquat. Gene expression studies indicated that katE, kaG, ahpC1, grxC, and oxyR itself were each differentially expressed in the oxyR mutant background and in response to hydrogen peroxide, suggesting that oxyR is necessary for hydrogen peroxideinducible gene expression. Additional OSR genes were differentially regulated in response to hydrogen peroxide alone. The virulence of the oxyR mutant strain was significantly reduced in both tomato and tobacco host plants, demonstrating that R. solanacearum is exposed to inhibitory concentrations of ROS in planta and that OxyRmediated responses to ROS during plant pathogenesis are important for R. solanacearum host adaptation and virulence.Plants produce reactive oxygen species (ROS) in response to invading pathogens, and an effective oxidative stress response (OSR) contributes to the fitness of phytopathogenic bacteria (18,39,40,51). However, research on the role of regulators of the OSR in the virulence of plant-pathogenic bacteria is limited and contradictory. In Agrobacterium tumefaciens, oxyR is necessary for tumorigenesis (34), but Erwinia chrysanthemi does not need oxyR for soft-rot virulence (32). OxyR is a redox-sensing LysR family transcriptional regulator that has been well characterized in several bacteria (23,43,52). In the absence of oxidative stress, OxyR is reduced and acts as a repressor of several genes, including oxyR itself (47, 52). In the presence of hydrogen peroxide, the conserved cysteines of OxyR (C199 and C208, in Escherichia coli) form a disulfide bond that changes its conformation and converts OxyR into a transcriptional activator (52, 53). In E. coli, OxyRregulated genes include catalase (kat), alkyl hydroperoxide reductase (ahp), glutaredoxin (grx), and glutathione reductase (gor) (53).The plant-pathogenic bacterium Ralstonia solanacearum causes bacterial wilt disease on many economically important crops, including tomato (19). Multiple quantitative virulence factors contribute to the disease (11); however, little is known about how R. solanacearum adapts to its host environment, which is a critical prerequisite for pathogen success. An in vivo expression technology (IVET) screen performed on R...

Recurrent honey bee losses make it critical to understand the impact of human interventions, such as antibiotics use in apiculture. Antibiotics are used to prevent or treat bacterial infections in colonies. However, little is known about their effects on honey bee development. We studied the effect of two commercial beekeeping antibiotics on the bee physiology and behavior throughout development. Our results show that antibiotic treatments have an effect on amount of lipids and rate of behavioral development. Lipid amount in treated bees was higher than those not treated. Also, the timing of antibiotic treatment had distinct effects for the age of onset of behaviors starting with cleaning, then nursing and lastly foraging. Bees treated during larva-pupa stages demonstrated an accelerated behavioral development and loss of lipids, while bees treated from larva to adulthood had a delay in behavioral development and loss of lipids. The effects were shared across the two antibiotics tested, TerramycinR (oxytetracycline) and TylanR (tylosin tartrate). These results on effects of antibiotic treatments suggest a role of microbiota in the interaction between the fat body and brain that is important for honey bee behavioral development.