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Perkins Koefoed posted an update 6 months, 1 week ago
udy, we determined that mutations in the C. difficile SpoIVA ATPase motifs result in relatively minor defects in spore formation, in contrast with Bacillus subtilis Nevertheless, our data suggest that SipL preferentially recognizes the ATP-bound form of SpoIVA and identify a specific residue in the SipL C-terminal LysM domain that is critical for recognizing the ATP-bound form of SpoIVA. These findings advance our understanding of how SpoIVA-SipL interactions regulate C. difficile spore assembly.Fluctuations in osmolarity are one of the most prevalent stresses to which bacteria must adapt, both hypo- and hyperosmotic conditions. Most bacteria cope with high osmolarity by accumulating compatible solutes (osmolytes) in the cytoplasm to maintain the turgor pressure of the cell. Vibrio parahaemolyticus, a halophile, utilizes at least six compatible solute transporters for the uptake of osmolytes two ABC family ProU transporters and four betaine-carnitine-choline transporter (BCCT) family transporters. The full range of compatible solutes transported by this species has yet to be determined. Using an osmolyte phenotypic microarray plate for growth analyses, we expanded the known osmolytes used by V. parahaemolyticus to include N,N-dimethylglycine (DMG), among others. Growth pattern analysis of four triple-bccT mutants, possessing only one functional BCCT, indicated that BccT1 (VP1456), BccT2 (VP1723), and BccT3 (VP1905) transported DMG. BccT1 was unusual in that it could take up both compounds with methylytes that were utilized by V. parahaemolyticus We demonstrated that the compound DMG, which is present in the marine environment, was a highly effective osmolyte for Vibrio species. We determined that DMG is transported via BCCT family carriers, which have not been shown previously to take up this compound. BccT1 was a carrier for GB, DMG, and ectoine, and we identified the amino acid residues essential for the coordination of these compounds. The data suggest that for BccT1, GB is more easily accommodated than ectoine in the transporter binding pocket.Naturally competent organisms are capable of DNA uptake directly from the environment through the process of transformation. Despite the importance of transformation to microbial evolution, DNA uptake remains poorly characterized outside of the bacterial domain. Here, we identify the pilus as a necessary component of the transformation machinery in archaea. We describe two naturally competent organisms, Methanococcus maripaludis and Methanoculleus thermophilus In M. maripaludis, replicative vectors were transferred with an average efficiency of 2.4 × 103 transformants μg-1 DNA. In M. thermophilus, integrative vectors were transferred with an average efficiency of 2.7 × 103 transformants μg-1 DNA. Additionally, natural transformation of M. Epigenetic inhibitor ic50 thermophilus could be used to introduce chromosomal mutations. To our knowledge, this is the first demonstration of a method to introduce targeted mutations in a member of the order Methanomicrobiales For both organisms, mutants lacking structural components of the type IV-lests that pili are important for transformation across the tree of life and further expands our understanding of gene flow in archaea.Bordetella pertussis uses a type III secretion system (T3SS) to inject virulence proteins into host cells. Although the B. pertussis T3SS was presumed to be involved in host colonization, efficient secretion of type III secreted proteins from B. pertussis has not been observed. To investigate the roles of type III secreted proteins during infection, we attempted to optimize culture conditions for the production and secretion of a type III secreted protein, BteA, in B. pertussis We observed that B. pertussis efficiently secretes BteA in ascorbic acid-depleted (AsA-) medium. When L2 cells, a rat lung epithelial cell line, were infected with B. pertussis cultured in the AsA- medium, BteA-dependent cytotoxicity was observed. We also performed an immunofluorescence assay of L2 cells infected with B. pertussis Clear fluorescence signals of Bsp22, a needle structure of T3SS, were detected on the bacterial surface of B. pertussis cultured in the AsA- medium. Since ascorbic acid is known as a reducing agent, we culturx signaling is involved in the regulation of B. pertussis T3SS.Plasma metagenomic next-generation sequencing (mNGS) is a new diagnostic method used to potentially identify multiple pathogens with a single DNA-based diagnostic test. The test is expensive, and little is understood about where it fits into the diagnostic schema. We describe our experience at Texas Children’s Hospital with the mNGS assay by Karius from Redwood City, CA, to determine whether mNGS offers additional diagnostic value when performed within 1 week before or after conventional testing (CT) (i.e., concurrently). We performed a retrospective review of all patients who had mNGS testing from April to June of 2019. Results for mNGS testing, collection time, time of result entry into the electronic medical record, and turnaround time were compared to those for CT performed concurrently. Discordant results were further reviewed for changes in antimicrobials due to the additional organism(s) identified by mNGS. Sixty patients had mNGS testing; the majority were immunosuppressed (62%). There was 61% positive agreement and 58% negative agreement between mNGS and CT. The mean time of result entry into the electronic medical record for CT was 3.5 days earlier than the mean result time for mNGS. When an additional organism(s) was identified by mNGS, antimicrobials were changed 26% of the time. On average, CT provided the same result as mNGS, but sooner than mNGS. When additional organisms were identified by mNGS, there was no change in management in the majority of cases. Overall, mNGS added little diagnostic value when ordered concurrently with CT.The O-serogrouping of pathogenic Escherichia coli is a standard method for subtyping strains for epidemiological studies and controls. O-serogroup diversification shows a strong association with the genetic diversity in some O-antigen biosynthesis gene clusters. Through genomic studies, in addition to the types of O-antigen biosynthesis gene clusters (Og-types) from conventional O-serogroup strains, a number of novel Og-types have been found in E. coli isolates. To assist outbreak investigations and surveillance of pathogenic E. coli at inspection institutes, in previous studies, we developed PCR methods that could determine almost all conventional O-serogroups and some novel Og-types. However, there are still many Og-types that may not be determined by simple genetic methods such as PCR. Thus, in the present study, we aimed to develop an additional Og-typing PCR system. Based on the novel Og-types, including OgN32, OgN33, and OgN34, presented in this study, we designed an additional 24 PCR primer pairs targeting 14 novel and 2 diversified E.
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