Xanthomonas campestris pathovar campestris (Xcc) is the causative agent of crucifer black rot disease, which causes severe losses in agricultural yield world-wide. This bacterium is a model organism for studying plant-bacteria interactions. We sequenced the complete genome of Xcc 8004 (5,148,708 bp), which is highly conserved relative to that of Xcc ATCC 33913. Comparative genomics analysis indicated that, in addition to a significant genomic-scale rearrangement cross the replication axis between two IS1478 elements, loss and acquisition of blocks of genes, rather than point mutations, constitute the main genetic variation between the two Xcc strains. Screening of a high-density transposon insertional mutant library (16,512 clones) of Xcc 8004 against a host plant (Brassica oleraceae) identified 75 nonredundant, single-copy insertions in protein-coding sequences (CDSs) and intergenic regions. In addition to known virulence factors, full virulence was found to require several additional metabolic pathways and regulatory systems, such as fatty acid degradation, type IV secretion system, cell signaling, and amino acids and nucleotide metabolism. Among the identified pathogenicity-related genes, three of unknown function were found in Xcc 8004-specific chromosomal segments, revealing a direct correlation between genomic dynamics and Xcc virulence. The present combination of comparative and functional genomic analyses provides valuable information about the genetic basis of Xcc pathogenicity, which may offer novel insight toward the development of efficient methods for prevention of this important plant disease.

Adherence of enterohemorrhagic Escherichia coli (EHEC) to intestinal epithelium is essential for initiation of infections, including diarrhea, and expression of the genes of the locus of enterocyte effacement (LEE) is thought to be crucial for adherence. To identify genes involved in modulating the adherent capacity, bacteria collected from an EHEC O157:H7 strain (O157Sakai) mutagenized by mini-Tn5Km2 were screened for their ability to increase the number of microcolonies (MC) on Caco-2 cells and eight mutants with increased adherence were isolated. Analysis of the mini-Tn5Km2-flanked DNA sequences indicated that one possessed the insertion within an O157 antigen gene cluster, another possessed the insertion within the yhiF gene, and the remaining six mutants had their insertions in the yhiE gene. yhiE and yhiF products share amino acid homology (23% identity) to each other and with members of the LuxR family, which are known as transcriptional regulatory proteins. The mutant having the insertion within the O157 antigen gene cluster did not express the O157 side chain (as determined by agglutination test and immunoblotting with polyclonal O157-specific antiserum), unlike the other seven mutants. Importantly, the other mutants showed enhanced type III secretion. Levels of the related mRNAs of genes of the LEE, but not that of ler mRNA, were also increased compared with those in the wild type. Indeed, when we introduced an in-frame deletion into the yhiE or yhiF gene in O157Sakai, the capacity of the resultant mutants to adhere to Caco-2 cells was greatly increased. When one of the yhiE insertion mutants was orally inoculated into ICR mice, the number of bacteria shed into feces by day 14 was greater than that for the wild type. These results suggest that yhiE and yhiF are involved in the adherence of O157Sakai to epithelial cells as negative regulators for the expression of the genes required for the type III secretion system.Enterohemorrhagic Escherichia coli (EHEC) are members of a class of pathogenic E. coli that cause a range of illnesses, including nonbloody diarrhea, hemorrhagic colitis, and hemolytic uremic syndrome. Although the mechanisms involved in infection of intestines in humans are still to be elucidated, the pathogenesis of EHEC (represented by strain O157:H7) has been shown to have several characteristics, including the production of Shiga toxins, the ability to produce an attaching and effacing (A/E) intestinal lesion, and the ability to induce enterohemolytic activity (6,17,32,35).Among these characteristics, the ability to induce A/E intestinal lesions is shared by enteropathogenic E. coli (EPEC) (32). The attachment of both pathogens to intestinal epithelial cells, leading to the formation of A/E lesions, depends on the presence of the locus of enterocyte effacement (LEE) region, which contains a subset of genes encoding intimin (eae), its receptor (tir/espE), a type III secretion machine, and the type III secreted proteins, such as EspA, EspB, and EspD (9,15,22,25). In vitro assays have indi...

Huntington’s disease (HD) is an inherited neurodegenerative disorder caused by the abnormal expansion of CAG repeats in the huntingtin (HTT) gene, which leads to progressive loss of neurons starting in the striatum and cortex. One possible mechanism for this selective loss of neurons in the early stage of HD is altered neurotransmission at synapses. Despite the recent finding that presynaptic terminals play an important role in HD, neurotransmitter release at synapses in HD remains poorly understood. Here, we measured synaptic vesicle release in real time at single presynaptic terminals during electrical field stimulation. We found the increase in synaptic vesicle release at presynaptic terminals in primary cortical neurons in a knock-in mouse model of HD (zQ175). We also found the increase in Ca2+ influx at presynaptic terminals in HD neurons during the electrical stimulation. Consistent with increased Ca2+-dependent neurotransmission in HD neurons, the increase in vesicle release and Ca2+ influx was rescued with Ca2+ chelators or by blocking N-type voltage-gated Ca2+ channels, suggesting N-type voltage-gated Ca2+ channels play an important role in HD. Taken together, our results suggest that the increased synaptic vesicles release due to increased Ca2+ influx at presynaptic terminals in cortical neurons contributes to the selective neurodegeneration of these neurons in early HD and provide a possible therapeutic target.