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Adenosine kinase (ADK) is a purine salvage enzyme and a typical housekeeping enzyme in eukaryotes which catalyzes the phosphorylation of adenosine to form AMP. Since prokaryotes synthesize purines de novo and no endogenous ADK activity is detectable in Escherichia coli, ADK has long been considered to be rare in bacteria. To date, only two prokaryotes, both of which are gram-positive bacteria, have been reported to contain ADK. Here we report that the gram-negative bacterium Xanthomonas campestris pathovar campestris, the causal agent of black rot of crucifers, possesses a gene (designated [adk.sub.Xcc]) encoding an ADK (named [ADK.sub.Xcc]), and we demonstrate genetically that the [ADK.sub.Xcc] is involved in extracellular polysaccharide (EPS) production, cell motility, and pathogenicity of X. campestris pv. campestris, [adk.sub.Xcc] was overexpressed as a [His.sub.6]-tagged protein in E. coli, and the purified [His.sub.6]-tagged protein exhibited ADK activity. Mutation of [adk.sub.Xcc] did not affect bacterial growth in rich and minimal media but led to an accumulation of intracellular adenosine and diminutions of intracellular ADK activity and ATP level, as well as EPS. The [adk.sub.Xcc] mutant displayed significant reductions in bacterial growth and virulence in the host plant.

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) plays an important role in glucose catabolism, converting glyceraldehyde 3-phosphates to 1,3-bisphosphoglycerates. Open reading frame (ORF) XC_0972 in the genome of Xanthomonas campestris pv. campestris (Xcc) strain 8004 is the only ORF in this strain annotated to encode a GAPDH. In this work, we have demonstrated genetically that this ORF encodes a unique GAPDH in Xcc strain 8004, which seems to be constitutively expressed. A GAPDH-deficient mutant could still grow in medium with glucose or other sugars as the sole carbon source, and no phosphofructokinase activity was detectable in strain 8004. These facts suggest that Xcc may employ the Entner--Doudoroff pathway, but not glycolysis, to utilize glucose. The mutant could not utilize pyruvate as sole carbon source, whereas the wild-type could, implying that the GAPDH of Xcc is involved in gluconeogenesis. Furthermore, inactivation of the Xcc GAPDH resulted in impairment of bacterial growth and virulence in the host plant, and reduction of intracellular ATP and extracellular polysaccharide (EPS). This reveals that GAPDH is required for EPS production and full pathogenicity of Xcc.

Xanthomonas campestris pathovar campestris causes black rot, a vascular disease on cruciferous plants, including Arabidopsis thaliana. The gene XC1553 from X. campestris pv. campestris strain 8004 encodes a protein containing leucine-rich repeats (LRRs) and appears to be restricted to strains of X. campestris pv. campestris. LRRs are found in a number of type III-secreted effectors in plant and animal pathogens. These prompted us to investigate the role of the XC1553 gene in the interaction between X. campestris pv. campestris and A. thaliana. Translocation assays using the hypersensitive-reaction-inducing domain of X. campestris pv. campestris AvrBs1 as a reporter revealed that XC1553 is a type III effector. Infiltration of Arabidopsis leaf mesophyll with bacterial suspensions showed no differences between the wild-type strain and an XC1553 gene mutant: both strains induced disease symptoms on Kashmir and Col-0 ecotypes. However, a clear difference was observed when bacteria were introduced into the vascular system by piercing the central vein of leaves. In this case, the wild-type strain 8004 caused disease on the Kashmir ecotype, but not on ecotype Col-0; the XC1553 gene mutant became virulent on the Col-0 ecotype and still induced disease on the Kashmir ecotype. Altogether, these data show that the XC1553 gene, which was renamed [avrAC.sub.Xcc8004], functions as an avirulence gene whose product seems to be recognized in vascular tissues.

The genome of the Xanthomonas campestris pathovar campestris (Xcc) strain 8004 encodes three uncharacterized proteins, XC1166, XC1223 and XC1976, annotated as glucose kinase (Glk) by bioinformatic studies. Here we have investigated the biochemical characteristics and physiological roles of these proteins with particular reference to the synthesis of extracellular polysaccharide (EPS). XC1166, XC1223 and XC1976 were overexpressed as fusion proteins with a [His.sub.6] affinity tag and purified by nickel affinity chromatography. The standard GIk activity assay revealed that all three proteins possessed apparent Glk activity, with XC1976-[His.sub.6] being the most active; the specific activity values were 1.16 x [10.sup.6] U [mg.sup.-1] for XC1166-[His.sup.6], 4.36x[10.sup.7] U [mg.sup.-1] for XC1223-[His.sub.6] and 2.63 x [10.sup.8] U [mg.sup.-1] for XC1976-[His.sub.6]. TLC analysis showed, however, that only XC1976-[His.sub.6] could phosphorylate glucose. Insertional mutants of XC1166, XC1223 and XC1976 were generated using the suicide plasmid pK18mob. Although mutant strains with insertions in XC1166 or XC1223 had GIk activity similar to that of the wild-type strain, the XC1976 mutant had only about 6% of the wild-type activity. Mutation in XC1976 had complex effects on EPS production. In media containing arabinose, glucose, galactose, sucrose or maltose, the XC1976 mutant produced about 40-75% of the wild-type level of EPS, whereas in medium containing fructose, the mutant showed a 30% increase in EPS production compared to the wild-type strain. The X01976 mutant also showed attenuated virulence on the host plant Chinese radish (Raphanus sativus). The results indicate that XC1976 has the most significant role for the parameters tested.

The MarR family of transcriptional regulators of bacteria are involved in the regulation of many cellular processes, including pathogenesis. In this work, we have demonstrated genetically that hpaR (hpa, hrp associated), which encodes a putative MarR family regulator, is involved in the hypersensitive response (HR), pathogenicity, and extracellular protease production of the phytopathogenic bacterium Xanthomonas campestris pathovar campestris. A mutation in hpaR resulted in complete loss of virulence in the host plant cabbage, a delayed and weakened HR in the nonhost plant pepper ECW-10R, and an increase in extracellular protease production. Detection of the [beta]-glucuronidase activity of a plasmid-driven hpaR promoter-gusA reporter revealed that the expression of hpaR is positively controlled by HrpG and HrpX and is suppressed in rich medium while being strongly induced in minimal and hrp-inducing media and inside the host. These findings indicate that hpaR belongs to the hrpG and hrpX regulon and that HrpX regulates the extracellular protease production via hpaR in X. campestris pv. campestris.

Xanthomonas campestris pathovar campestris (Xcc) is the causal agent of black rot disease in cruciferous plants. The extracellular polysaccharide (EPS) produced by Xcc is an important pathogenicity factor and also has a range of industrial uses. In preliminary work a number of transposon-mediated insertion mutants in Xcc with defects in EPS production were identified. Here, one of these mutated loci was investigated in detail. Six ORFs within the locus (ORFs XC3811-3816) were disrupted by plasmid integration. Mutation of XC3813, XC3814 or XC3815 resulted in significantly reduced EPS production and significantly reduced virulence on the host plant Chinese radish (Raphanus sativus). The EPS production and virulence of XC3813, XC3814 and XC3815 mutants could be restored by intact XC3813, XC3814 and XC3815 genes, respectively, when provided in trans. Although bioinformatic analysis suggested a role for XC3814 and XC3815 in lipopolysaccharide biosynthesis, the lipopolysaccharides produced by the mutants were indistinguishable from those of the wild-type, as judged by electrophoretic mobility in SDS-polyacrylamide gels. These results reveal that XC3813, XC3814 and XC3815 comprise a novel gene cluster involved in EPS production and virulence of Xcc.

Disruption of ppsA, a key gene in gluconeogenesis, of Xanthomonas campestris pv. campestris resulted in the failure of the pathogen to grow in medium with pyruvate or C4-dicarboxylates as the sole carbon source and a significant reduction in virulence, indicating that X. campestris pv. campestris possesses only the malic enzyme-PpsA route in gluconeogenesis, which is required for virulence.

Xanthomonas oryzae pv. textitoryzae (Xoo) is a causal agent of rice bacterial leaf blight (BLB), the major rice disease, which is seriously constraining rice production in Asia. The interaction between Xoo and rice is in a dynamic process, essentially the co-evolution. Tracking the occurrence of plant diseases and identifying the epidemic pathogens in time are critical to assessing the epidemic disease status and understanding the pathogen evolution. In 2020, the occurrences of rice BLB were spotted in many places of Guangxi, the major rice growing region in China. Two of the 2020-epidemic Xoo strains, namely, GXO20-01 and GXO20-06, were isolated from low land and high mountain paddies in Guangxi, respectively, and were demonstrated to be race R8 of Chinese Xoo strains, but with significantly different virulence on certain susceptible varieties of rice. The HiFi PacBio sequencing revealed that GXO20-01 and GXO20-06 share the highly syntenic genome structures and the major genome contents, but only differ in <10 genes, including one gene encoding for transcription activator-like effector (TALE). A phylogenomic analysis grouped GXO20-01 and GXO20-06 into the PX-A lineage, stood close to PXO563 and PXO71 strains, but stood away from the other Chinese Xoo strains; for example, the JL25 and YC11. A comparative genomic analysis revealed that the major pathogenicity/virulence genes are conserved in two, newly isolated Xoo strains and the other Xoo strains in PX-A lineage, including the majority genes for the TALomes. The genomic differences between the Xoo strains were pinpointed to a few tal genes, which were variable in both their numbers and sequences, even between GXO20-01 and GXO20-06, the two 2020-epidemic Xoo strains. The study further revealed the instability and variability of tal genes in Xoo and highlighted the utility of HiFi long-read sequencing in TALE analysis and pathogen tracking. [ABSTRACT FROM AUTHOR]

Abstract Background One of the major tasks of the post-genomic era is "reading" genomic sequences in order to extract all the biological information contained in them. Although a wide variety of techniques is used to solve the gene finding problem and a number of prokaryotic gene-finding software are available, gene recognition in bacteria is far from being always straightforward. Results This study reported a thorough search for new CDS in the two published Xcc genomes. In the first, putative CDSs encoded in the two genomes were re-predicted using three gene finders, resulting in the identification of 2850 putative new CDSs. In the second, similarity searching was conducted and 278 CDSs were found to have homologs in other bacterial species. In the third, oligonucleotide microarray and RT-PCR analysis identified 147 CDSs with detectable mRNA transcripts. Finally, in-frame deletion and subsequent phenotype analysis of confirmed that Xcc_CDS002 encoding a novel SIR2-like domain protein is involved in virulence and Xcc_CDS1553 encoding a ArsR family transcription factor is involved in arsenate resistance. Conclusions Despite sophisticated approaches available for genome annotation, many cellular transcripts have remained unidentified so far in Xcc genomes. Through a combined strategy involving bioinformatic, postgenomic and genetic approaches, a reliable list of 306 new CDSs was identified and a more thorough understanding of some cellular processes was gained.

Abstract Background In bacteria, small non-coding RNAs (sRNAs) have been recognized as important regulators of various cellular processes. Approximately 200 bacterial sRNAs in total have been reported. However, very few sRNAs have been identified from phytopathogenic bacteria. Results Xanthomons campestris pathovar campestris (Xcc) is the causal agent of black rot disease of cruciferous crops. In this study, a cDNA library was constructed from the low-molecular weight RNA isolated from the Xcc strain 8004 grown to exponential phase in the minimal medium XVM2. Seven sRNA candidates were obtained by sequencing screen of 2,500 clones from the library and four of them were confirmed to be sRNAs by Northern hybridization, which were named sRNA-Xcc1, sRNA-Xcc2, sRNA-Xcc3, and sRNA-Xcc4. The transcription start and stop sites of these sRNAs were further determined. BLAST analysis revealed that the four sRNAs are novel. Bioinformatics prediction showed that a large number of genes with various known or unknown functions in Xcc 8004 are potential targets of sRNA-Xcc1, sRNA-Xcc3 and sRNA-Xcc4. In contrast, only a few genes were predicted to be potential targets of sRNA-Xcc2. Conclusion We have identified four novel sRNAs from Xcc by a large-scale screen. Bioinformatics analysis suggests that they may perform various functions. This work provides the first step toward understanding the role of sRNAs in the molecular mechanisms of Xanthomonas campestris pathogenesis.

In many tunnels in China, the groundwater is acidic and carbonated, causing in challenges in the prevention and control of leakages of the existing tunnels and tunnels under construction. Research on tunnel leakage and the physical and chemical effects of water due to water-rock interactions is a trending topic. In addition, there is a big difference between the waterproof and drainage of multiarch tunnels and separate tunnels. In this study, the mechanism of the interaction between the groundwater and the surrounding rock of a multiarch tunnel were analyzed. The relationship between the leakage of a multiarch tunnel and the interaction between the surrounding rocks and groundwater was determined by analyzing the interaction between the chemical components in the groundwater, rocks, and the concrete lining. A mathematical model was established based on the physical and chemical reactions in the rock surrounding the tunnel, and the RNCDX.FOR program was compiled using the simplex Monte Carlo method. The total free energy in the entire system varied significantly, and the value of objective function (J) reflected the trend of the system's spontaneous reaction. As J decreased, the reaction power of the system increased. The more significant the erosion of the rocks was by the groundwater, the more channels were created by the groundwater in the rock, and the more likely the water leakage was. [ABSTRACT FROM AUTHOR]

Xanthomonas is a well-studied genus of bacterial plant pathogens whose members cause a variety of diseases in economically important crops worldwide. Genomic and functional studies of these phytopathogens have provided significant understanding of microbial-host interactions, bacterial virulence and host adaptation mechanisms including microbial ecology and epidemiology. In addition, several strains of Xanthomonas are important as producers of the extracellular polysaccharide, xanthan, used in the food and pharmaceutical industries. This polymer has also been implicated in several phases of the bacterial disease cycle. In this review, we summarise the current knowledge on the infection strategies and regulatory networks controlling virulence and adaptation mechanisms from Xanthomonas species and discuss the novel opportunities that this body of work has provided for disease control and plant health. [ABSTRACT FROM AUTHOR]

The Gram-negative bacterium Xanthomonas oryzae pv. oryzicola (Xoc) is the causal agent of rice bacterial leaf streak (BLS), one of the most destructive diseases of rice (Oryza sativa L.) that is the important staple crop. Xoc can invade host leaves via stomata and wounds and its type three secretion system (T3SS) is pivotal to its pathogenic lifestyle. In this study, using a novel dual RNA-seq approach, we examined transcriptomes of rice and Xoc in samples inoculated with wild type Xoc GX01 and its T3SS defective strain (T3SD), to investigate the global transcriptional changes in both organisms. Compared with T3SD strain, rice inoculated with wild type Xoc GX01 resulted in significant expression changes of a series of plant defence related genes, including ones altered in plant signalling pathway, and downregulated in phenylalanine metabolism, flavonoid and momilactone biosynthesis, suggesting repression of plant defence response and reduction in both callose deposition and phytoalexin accumulation. Also, some known transcription activator-like effector (TALE) targets were induced by Xoc GX01, e.g. OsSultr3;6 which contributes to rice susceptibility. Some cell elongation related genes, including several expansin genes, were induced by GX01 too, suggesting that Xoc may exploit this pathway to weaken cell wall strength, beneficial for bacterial infection. On the other hand, compared with wild type, the T3SD strain transcriptome in planta was characterized by downregulation of ATP, protein and polysaccharide synthesis, and upregulation of antioxidation and detoxification related genes, revealing that T3SD strain faced serious starvation and oxidation stresses in planta without a functional T3SS. In addition, comparative global transcript profiles of Xoc in planta and in medium revealed an upregulation of virulence factor synthesis and secretion in planta in favour of bacterial infection. Collectively, this study provides a comprehensive representation of cross talk between the host and bacterial pathogen, revealing insights into the Xoc-rice pathogenic dynamic and reveals novel strategies exploited by this important pathogen to cause disease. [ABSTRACT FROM AUTHOR]

Two-component regulatory system (TCS), a major type of cellular signal transduction system, is widely used by bacteria to adapt to different conditions and to colonize certain ecological niches in response to environmental stimuli. TCSs are of distinct functional diversity, genetic diversity, and species specificity (pathovar specificity, even strain specificity) across bacterial groups. Although TCSs have been demonstrated to be crucial to the virulence of Xanthomonas , only a few researches have been reported about the studies of TCSs in Xanthomonas oryzae pathovar oryzicola (hereafter Xoc), the pathogen of rice bacterial streak disease. In the genome of Xoc strain GX01, it has been annotated 110 TCSs genes encoding 54 response regulators (RRs), 36 orthodox histidine kinase (HKs) and 20 hybrid histidine kinase (HyHKs). To evaluate the involvement of TCSs in the stress adaptation and virulence of Xoc , we mutated 50 annotated RR genes in Xoc GX01 by homologous vector integration mutagenesis and assessed their phenotypes in given conditions and tested their virulence on host rice. 17 RR genes were identified to be likely involved in virulence of Xoc , of which 10 RR genes are novel virulence genes in Xanthomonas , including three novel virulence genes for bacteria. Of the novel candidate virulence genes, some of which may be involved in the general stress adaptation, exopolysaccharide production, extracellular protease secretion and swarming motility of Xoc. Our results will facilitate further studies on revealing the biological functions of TCS genes in this phytopathogenic bacterium. [ABSTRACT FROM AUTHOR]

Background and objectives: Increase in the world population is usually followed by a growing demand for food and increasingly higher requirements for the quality of food. The existing crops may require immediate improvement and optimization in order to adjust to market demands and the deteriorating environment. Findings: Genome-editing technology, a relatively advanced technology method that appeared in recent years, can accurately edit (i.e., insert, delete, or replace) genes and directionally modify crops by changing the plant's heredity and performance. Conclusions: Currently, some of the modern genome-editing approaches have been applied rather widely in crop breeding: for example, zinc finger nuclease, transcription activator-like effector nuclease, and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein. [ABSTRACT FROM AUTHOR]

A correction to the article "Cyclic di-GMP Signalling in the Virulence and Environmental Adaptation of Xanthomonas Campestris," by Robert P. Ryan and colleagues is presented.

An effective multiplex real-time polymerase chain reaction (PCR) assay for the simultaneous detection of three major pathogens, Nosema bombycis Nägeli (Microsporidia: Nosematidae), Bombyx mori nucleopolyhedrovirus (Baculoviridae: genus Alphabaculovirus) (NPV), and Bombyx mori densovirus (Parvoviridae: genus Iteravirus) (DNV), in silkworms (Bombyx mori (Linnaeus); Lepidoptera: Bombycidae) was developed in this study. Polymerase chain reaction and real-time PCR tests and basic local alignment search tool searches revealed that the primers and probes used in this study had high specificities for their target species. The ability of each primer/probe set to detect pure pathogen DNA was determined using a plasmid dilution panel, in which under optimal conditions the multiplex real-time PCR assay showed high efficiency in the detection of three mixed target plasmids with a detection limit of 8.5×103 copies for N. bombycis and Bombyx mori NPV (BmNPV) and 8.5×104 copies for Bombyx mori DNV (BmDNV). When the ability to detect these three pathogens was examined in artificially inoculated silkworms, our method presented a number of advantages over traditional microscopy, including specificity, sensitivity, and high-throughput capabilities. Under the optimal volume ratio for the three primer/probe sets (3:2:2=N. bombycis:BmNPV:BmDNV), the multiplex real-time PCR assay showed early detection of BmNPV and BmDNV by day 1 post inoculation using DNA templates of the three pathogens in various combinations from individually infected silkworms; the early detection of N. bombycis was possible by day 3 post inoculation using the DNA isolated from the midgut of N. bombycis-infected silkworms. [ABSTRACT FROM AUTHOR]

In opportunistic networks, temporary nodes choose neighbor nodes to forward messages while communicating. However, traditional forward mechanisms don't take the importance of nodes into consideration while forwarding. In this paper, we assume that each node has a status indicating its importance, and temporary nodes choose the most important neighbors to forward messages. While discovering important neighbors, we propose a binary tree random walk based algorithm. We analyze the iteration number and communication cost of the proposed algorithm, and they are much less than related works. The simulation experiments validate the efficiency and effectiveness of the proposed algorithm. [ABSTRACT FROM AUTHOR]

Location estimation of mobile target is an important research in wireless sensor networks. However, the phenomenon of non line of sight affects the accuracy of location estimating algorithms greatly. In order to improve the accuracy of location estimation of mobile targets, we proposed a game theory based location method algorithm in wireless sensor networks. In the proposed method, we first estimate the initial locations of mobile target with a support vector regression model, and then use game theory to smooth the already estimated locations. In the game model, game based filter and noise generator are two game rivals. The game filter aims to minimize the objective function of the game model, and the noise generator aims to maximize the same objective function. We propose a differential game theory based algorithm for solving the optimal resolution. The experiments show that, the proposed method is more powerful, and can be used to mobile target locating and tracking. [ABSTRACT FROM AUTHOR]

A number of species of bacteria from the genus B urkholderia have been shown to be causal agents of diseases of rice. These diseases, caused by B urkholderia glumae, B . gladioli and B . plantarii, are becoming increasingly common across the globe. This is particularly so for B . glumae, whose ability to grow at elevated temperatures suggests that it may become a prevalent problem in an era of global warming. Despite the increasing threat to rice, relatively little is known about the virulence mechanisms employed by these pathogens. Work over the last 5 years has provided an increasing insight into these factors and their control by environmental and other cues. In addition, the determination of a number of genome sequences has allowed bioinformatic predictions of further possible mechanisms, which can now be investigated experimentally. Here, we review recent advances in the understanding of virulence of B urkholderia to rice, to include discussion of the roles of toxins, type II secreted enzymes, type III secreted effectors and motility as well as their regulation by quorum sensing, two-component systems and cyclic di- GMP signalling. Finally, we consider a number of approaches for the control of bacterial virulence through the modulation of quorum sensing and toxin degradation. [ABSTRACT FROM AUTHOR]

In order to improve the Internet coverage ratio and provide connectivity guarantee, based on sensor opportunistic coverage mechanism and cooperative cloud service, we proposed the coverage connectivity guarantee protocol for mobile Internet. In this scheme, based on the opportunistic covering rules, the network coverage algorithm of high reliability and real-time security was achieved by using the opportunity of sensor nodes and the Internet mobile node. Then, the cloud service business support platform is created based on the Internet application service management capabilities and wireless sensor network communication service capabilities, which is the architecture of the cloud support layer. The cooperative cloud service aware model was proposed. Finally, we proposed the mobile Internet coverage connectivity guarantee protocol. The results of experiments demonstrate that the proposed algorithm has excellent performance, in terms of the security of the Internet and the stability, as well as coverage connectivity ability. [ABSTRACT FROM AUTHOR]

The bacterial phytopathogen X anthomonas campestris pv. campestris ( Xcc) relies on the hrp (hypersensitive response and pathogenicity) genes to cause disease and induce hypersensitive response ( HR). The hrp genes of bacterial phytopathogens are divided into two groups. Xcc hrp genes belong to group II. It has long been known that the group II hrp genes are activated by an AraC-type transcriptional regulator whose expression is controlled by a two-component system ( TCS) response regulator (named HrpG in Xcc). However, no cognate sensor kinase has yet been identified. Here, we present evidence showing that the Xcc open-reading frame XC _3670 encodes a TCS sensor kinase (named HpaS). Mutation of hpaS almost completely abolished the HR induction and virulence. Bacterial two-hybrid and protein pull-down assays revealed that HpaS physically interacted with HrpG. Phos-tag™ SDS-PAGE analysis showed that mutation in hpaS reduced markedly the phosphorylation of HrpG in vivo. These data suggest that HpaS and HrpG are most likely to form a TCS. We also showed that XC _3669 (named hpaR2), which is adjacent to hpaS and encodes a putative TCS response regulator, is required for full virulence but not HR induction. HpaR2 also physically interacted with HpaS, suggesting that HpaS may also form another TCS with HpaR2. [ABSTRACT FROM AUTHOR]

Mip (macrophage infectivity potentiator) and Mip-like proteins have been demonstrated to be involved in virulence of several animal pathogens, but as yet none of their native bacterial targets has been identified. Our previous work demonstrated that the Mip-like protein found in the plant pathogen X anthomonas campestris pv. campestris ( Xcc) (hereafter called MipXcc) is also involved in virulence. Inactivation of the mipXcc gene leads to a significant reduction in exopolysaccharide production and extracellular protease activity via an unknown mechanism. The Xcc genome encodes six extracellular proteases, all of which are secreted via the type II secretion system. The serine protease PrtA makes the largest contribution to Xcc's total extracellular proteolytic activity. In this study, Western blotting analysis demonstrated that MipXcc was located in the periplasm. Bacterial two-hybrid and far-Western analysis indicated that MipXcc interacted with PrtA directly. Purified MipXcc was found to be able to rescue the protease activity of periplasmic proteins extracted from the mipXcc mutant. These findings show that MipXcc plays a role in the maturation of PrtA, which is the novel native target for at least one Mip or Mip-like protein. [ABSTRACT FROM AUTHOR]

Abstract: The ColR–ColS two-component signal transduction system was originally characterized as a regulatory system involved in the capacity of root-colonizing biocontrol bacterium Pseudomonas fluorescens to colonize plant roots. There are three pairs of putative colR–colS two-component regulatory systems annotated in the phytopathogen Xanthomonas campestris pathovar campestris. Mutational studies revealed that one of them, named colRXC1049 and colSXC1050 , is a global regulatory system involved in various cellular processes, including virulence, hypersensitive response and stress tolerance. Growth rate determination showed that, although the colRXC1049 and colSXC1050 mutants are not auxotrophic, colRXC1049 and colSXC1050 are required for the pathogen to proliferate well in standard media and host plants. Assays of β-glucuronidase activities of plasmid-driven promoter-gusA reporters and/or semi-quantitative RT-PCR demonstrated that colRXC1049 and colSXC1050 positively regulate expression of hrpC and hrpE operons, and that expression of colRXC1049 and colSXC1050 is not controlled by key hrp regulators HrpG and HrpX. [Copyright &y& Elsevier]

In this work, a sensitive electrochemical method for bleomycin (BLM) determination was reported on the basis of BLM-mediated activation of Zn2+-dependent DNAzyme and the adsorption of signal probes by a metal-organic framework (MOF) modified electrode. Two hairpin DNAs were employed in this protocol, one (HP1) for BLM recognition and one (HP2) for amplified signal output. The presence of BLM and Fe2+ caused the formation of BLM-Fe (II) complex to cleave HP1, releasing DNAzyme fragments, which could further hybridize with substrate HP2 to form a partial double-stranded DNA duplex and enable the activation of Zn2+-dependent DNAzyme with the coexistence of Zn2+. The Zn2+-dependent DNAzyme catalyzed the cyclic cleavage of magnetic beads (MB)-immobilized HP2 to release massive DNA fragments with a Fc-labeled- terminal, which could be used for BLM quantification through electrochemical measurement after their adsorption on a MOF modified electrode. Attributing to the high catalytic efficiency of DNAzyme and excellent electrochemical performance of MOF modified electrode, our method revealed an impressive limit of detection as low as 4 pM BLM with a linear range of 5–2000 pM. Besides, the easy synthesis of MOF without further modification and the easy way of adsorption for signal achievement facilitated the operation process. In virtue of the high sensitivity, selectivity and the simple-to-implement features, this method is believed to hold a great promising application for BLM determination in biomedical and clinical study. • Target-mediated activation of Zn2+-dependent DNAzyme was utilized for amplified bleomycin determination. • The MOF-based electrochemical signal was creatively and easily achieved without complicated modification. • This method revealed a low detection limit, a wide linear range and the ability for complex sample analysis. [ABSTRACT FROM AUTHOR]

Xanthomonas TALE transcriptional activators act as virulence or avirulence factors by activating host disease susceptibility or resistance genes. Their specificity is determined by a tandem repeat domain. Some Xanthomonas pathogens contain 10-30 TALEs per strain. Although TALEs play critical roles in pathogenesis, their studies have so far been limited to a few examples, due to their highly repetitive gene structure and extreme similarity among different members, which constrict sequencing and assembling. To facilitate TALE studies, we developed an efficient and rapid pipeline for genome-wide cloning of tal genes as many as possible from a strain. Here, we report the pipeline and its use to identify all 18 tal genes from a newly isolated strain of the rice pathogen Xathomonas oryzae. Target prediction revealed a number of potential rice targets including several notable genes such as genes encoding SWEET, WRKY, Hen1, and BAK1 proteins, which provide candidates for further experimental functional analysis of the TALEs. [ABSTRACT FROM AUTHOR]

• A panel of PEDV structural and nonstructural proteins were expressed. • These PEDV antigens were tested for reactivity with sera from PEDV-infected pigs. • 851 porcine sera were analyzed by ELISA with antigens showing immune-reactivity. • Pattern of neutralizing antibody was more similar to that of IgA in weaning piglets. • ORF3C and E in addition to S1 can be used as diagnostic markers for serologic detection. Given the highly contagious and acute nature of porcine epidemic diarrhea (PED), especially in piglets, there is an urgent need for the development of rapid and sensitive diagnostic assays. The diagnostic potentials of specific porcine epidemic diarrhea virus (PEDV) accessory and nonstructural proteins, if any, have not yet been investigated. In order to determine and compare which of the viral proteins may be useful as diagnostic antigens, whole virus (WV) particles and a panel of structural and nonstructural PEDV proteins [spike subunit 1 (S1), the C-terminal part of ORF3 (ORF3C), envelope (E), nonstructural protein 1 (Nsp1), Nsp2, Ac (acidic domain of Nsp3), and ADRP (ADP-ribose-1-monophosphatase domain of Nsp3), expressed individually in bacterial and/or mammalian cells] were tested for reactivity with sera from PEDV-infected pigs by ELISA and/or western blot analysis. According to western blots, serum antibody interactions with the S1 protein were relatively more sensitive and specific than ORF3C, E and Ac. Furthermore, a total of 851 serum samples from diarrheal pigs of different ages were analyzed by ELISA, with most showing immune-reactivity towards the WV, S1, ORF3C, and E proteins. The earliest IgG antibody response was observed in the one-week-old piglets, with similar antibody ontogeny and patterns of seroconversion for S1, ORF3C, E, and WV antigens. In addition, the pattern of neutralizing antibody was more similar to that of IgA in weaning piglets after PEDV infection. Collectively, these data provide more reliable information on the host immune response to different viral proteins, which will be useful for development of novel serological assays and for design of vaccines that better stimulate protective immunity. [ABSTRACT FROM AUTHOR]