Genomic and phylogenetic characterization of Elizabethkingia anophelis strains: The first two cases of life-threatening infection in Japan.

OBJECTIVE: Elizabethkingia anophelis causes meningitis, bloodstream infections, and respiratory infections in immunocompromised individuals. We examined two E. anophelis strains isolated from the first life-threatening cases caused by this species in Japan to determine the phylogenetic origin and genomic features of them. METHODS: We performed whole genome-based analysis to clarify the genetic relationship for the two strains (EK0004 and EK0079) and Elizabethkingia sp. strains isolated from worldwide and to characterize the genomic features such as the prevalence of virulence- and antimicrobial resistance (AMR)-related genes. PATIENTS: A 29-year-old man with hepatosplenic T-cell lymphoma and a 52-year-old man with systemic lupus erythematosus developed fatal bacteremia and meningitis due to E. anophelis, respectively. RESULTS: Two strains, EK0004 and EK0079, were genetically different but most closely related to the strains isolated from the largest outbreak in Wisconsin, USA from 2015 to 2016, and the strain isolated from cerebrospinal fluid of a patient in Florida, USA in 1982, respectively. The two strains contained AMR-related genes such as those encoding for an extended-spectrum ß-lactamase and multiple metallo-ß-lactamases and several virulence-related genes such as capsular polysaccharide synthesis gene clusters. CONCLUSIONS: Although further functional analyses are required to understand the virulence of these clones, these finding suggests that enough caution of E. anophelis infection in immunocompromised patients is required since the number of infections by this species is increasing outside Japan.

Involvement of IGF-1R-PI3K-AKT-mTOR pathway in increased number of GnRH3 neurons during androgen-induced sex reversal of the brain in female tilapia.

The neuroplastic mechanism of sex reversal in the fish brain remains unclear due to the difficulty in identifying the key neurons involved. Mozambique tilapia show different reproductive behaviours between sexes; males build circular breeding nests while females hold and brood fertilized eggs in their mouth. In tilapia, gonadotropin-releasing hormone 3 (GnRH3) neurons, located in the terminal nerve, regulate male reproductive behaviour. Mature males have more GnRH3 neurons than mature females, and these neurons have been indicated to play a key role in the androgen-induced female-to-male sex reversal of the brain. We aimed to elucidate the signalling pathway involved in the androgen-induced increase in GnRH3 neurons in mature female tilapia. Applying inhibitors to organotypic cultures of brain slices, we showed that the insulin-like growth factor (IGF)-1 receptor (IGF-1R)/PI3K/AKT/mTOR pathway contributed to the androgen-induced increase in GnRH3 neurons. The involvement of IGF-1 and IGF-1R in 11-ketotestosterone (11-KT)-induced development of GnRH3 neurons was supported by an increase in Igf-1 mRNA shortly after 11-KT treatment, the increase of GnRH3 neurons after IGF-1 treatment and the expression of IGF-1R in GnRH3 neurons. Our findings highlight the involvement of IGF-1 and its downstream signalling pathway in the sex reversal of the tilapia brain.

Structural basis for sequence-dependent recognition of colicin E5 tRNase by mimicking the mRNA-tRNA interaction.

Colicin E5--a tRNase toxin--specifically cleaves QUN (Q: queuosine) anticodons of the Escherichia coli tRNAs for Tyr, His, Asn and Asp. Here, we report the crystal structure of the C-terminal ribonuclease domain (CRD) of E5 complexed with a substrate analog, namely, dGpdUp, at a resolution of 1.9 A. Thisstructure is the first to reveal the substrate recognition mechanism of sequence-specific ribonucleases. E5-CRD realized the strict recognition for both the guanine and uracil bases of dGpdUp forming Watson-Crick-type hydrogen bonds and ring stacking interactions, thus mimicking the codons of mRNAs to bind to tRNA anticodons. The docking model of E5-CRD with tRNA also suggests its substrate preference for tRNA over ssRNA. In addition, the structure of E5-CRD/dGpdUp along with the mutational analysis suggests that Arg33 may play an important role in the catalytic activity, and Lys25/Lys60 may also be involved without His in E5-CRD. Finally, the comparison of the structures of E5-CRD/dGpdUp and E5-CRD/ImmE5 (an inhibitor protein) complexes suggests that the binding mode of E5-CRD and ImmE5 mimics that of mRNA and tRNA; this may represent the evolutionary pathway of these proteins from the RNA-RNA interaction through the RNA-protein interaction of tRNA/E5-CRD.

Sequence-specific recognition of colicin E5, a tRNA-targeting ribonuclease.

Colicin E5 is a novel Escherichia coli ribonuclease that specifically cleaves the anticodons of tRNA(Tyr), tRNA(His), tRNA(Asn) and tRNA(Asp). Since this activity is confined to its 115 amino acid long C-terminal domain (CRD), the recognition mechanism of E5-CRD is of great interest. The four tRNA substrates share the unique sequence UQU within their anticodon loops, and are cleaved between Q (modified base of G) and 3' U. Synthetic minihelix RNAs corresponding to the substrate tRNAs were completely susceptible to E5-CRD and were cleaved in the same manner as the authentic tRNAs. The specificity determinant for E5-CRD was YGUN at -1 to +3 of the 'anticodon'. The YGU is absolutely required and the extent of susceptibility of minihelices depends on N (third letter of the anticodon) in the order A > C > G > U accounting for the order of susceptibility tRNA(Tyr) > tRNA(Asp) > tRNA(His), tRNA(Asn). Contrastingly, we showed that GpUp is the minimal substrate strictly retaining specificity to E5-CRD. The effect of contiguous nucleotides is inconsistent between the loop and linear RNAs, suggesting that nucleotide extension on each side of GpUp introduces a structural constraint, which is reduced by a specific loop structure formation that includes a 5' pyrimidine and 3' A.