Group B Streptococcus Infections Caused by Improper Sourcing and Handling of Fish for Raw Consumption, Singapore, 2015-2016.

We assessed microbial safety and quality of raw fish sold in Singapore during 2015-2016 to complement epidemiologic findings for an outbreak of infection with group B Streptococcus serotype III sequence type (ST) 283 associated with raw fish consumption. Fish-associated group B Streptococcus ST283 strains included strains nearly identical (0-2 single-nucleotide polymorphisms) with the human outbreak strain, as well as strains in another distinct ST283 clade (57-71 single-nucleotide polymorphisms). Our investigations highlight the risk for contamination of freshwater fish (which are handled and distributed separately from saltwater fish sold as sashimi) and the need for improved hygienic handling of all fish for raw consumption. These results have led to updated policy and guidelines regarding the sale of ready-to-eat raw fish dishes in Singapore.

Hypophysitis presented as inflammatory pseudotumor in immunoglobulin G4-related systemic disease.

Immunoglobulin (Ig) G4-related systemic disease is a recently characterized entity. The best-known manifestation is pancreatitis. Other systemic involvements are also described. Three cases of this disease with hypophyseal involvement have been reported in the literature, all diagnosed clinically. We herein present the first case of IgG4-related hypophysitis diagnosed histopathologically. The patient is a 77-year-old Chinese man with a pituitary tumor. Histologic examination of the resected tumor showed hypophysitis with features of inflammatory pseudotumor. Clinical review showed history of pancreatitis and cholecystitis 4 years ago. The pancreatic biopsy and gall bladder specimens obtained previously had the same pathologic features of inflammatory pseudotumor. Immunohistochemistry highlighted abundant IgG4-positive plasma cells in all 3 specimens. Serum IgG4 level was also elevated. A diagnosis of IgG4-related systemic disease was confirmed. This is the first case of intracranial inflammatory pseudotumor shown to be associated with IgG4-related systemic disease.

Evolutionary conservation and mutational spectrum of BMPR2 gene.

A variety of mutations in the bone morphogenetic protein receptor type 2 (BMPR2) have been identified in patients with pulmonary arterial hypertension. In this study, using our BMPR2 mutation database and BMPR-II protein sequences from eight distantly related species, we defined the relationship among evolutionary conservation, mutation frequency and mutation distribution. As a whole, BMPR2 is evolving slower than the average for mammalian protein-encoding genes. As expected, the kinase domain is evolving more slowly than the extracellular ligand-binding and C-terminal domains. A detailed map of evolutionary conservation shows that there are repeating peaks and valleys within the C-terminal domain, representing higher and lower evolutionary conservation. We observed a strong correlation between evolutionary conservation and the distribution of mutations along the gene. All except two, of the nineteen missense mutations occur in absolutely conserved amino acids among the vertebrate homologs. In addition, we identified six mutational hotspots (P<0.05) by comparing the observed distribution of mutations to the pattern expected from a random multinomial distribution. Furthermore, analysis of the sequence environment surrounding the mutations revealed a specific pattern of mutagenesis. Over 22% of all single base-paired substitutions and 30% of all deletions and insertions are situated within tandem or non-tandem direct repeats of at least 5-bp and may be explained by slipped-mispairing model of mutagenesis. Also, over 59% of single base-paired substitutions versus 20% of deletions and insertions are located in perfect palindromic sequences that could produce "hairpin-loop" secondary structures with relatively high thermodynamic stability under physiological conditions. In addition, 3.7% of single base-paired substitutions versus 30% of deletions and insertions are located either within or in close proximity to the Krawczak and Cooper consensus sequence (TG A/G A/G G/T A/C). Further study of the mechanism of mutagenesis in BMPR2 may help identify other potentially mutable sites and differentiate between deleterious mutations and harmless polymorphic variants.

Bone morphogenetic protein receptor type II C-terminus interacts with c-Src: implication for a role in pulmonary arterial hypertension.

Mutations of bone morphogenetic protein receptor type II (BMPR-II) have been associated with familial and idiopathic pulmonary arterial hypertension (PAH). BMPR-II is a member of the transforming growth factor-beta receptor superfamily. It consists of extracellular, transmembrane, and kinase domains, and a unique C-terminus with mostly unknown function. However, a number of PAH-causing mutations are predicted to truncate the C-terminus, suggesting that this domain plays an important role in the homeostasis of pulmonary vessels. In this study, we sought to elucidate the functional role of this C-terminus by seeking its interacting partners. Using yeast two-hybrid screening, we identified c-Src tyrosine kinase as a binding partner of this C-terminus. In vitro co-immunoprecipitation confirmed their interaction. Mutations truncating the C-terminus disrupted their interaction, while missense mutation within kinase domain reduced their interaction. In addition, BMPR-II and c-Src tyrosine kinase colocalized within intracellular aggregates when overexpressed in HEK293 cells. Moreover, mutations truncating the C-terminus disrupted their colocalization, whereas missense mutation within kinase domain had no effect on their colocalization. Furthermore, BMP ligand stimulation decreased c-Src-activating phosphorylation at Tyrosine 418 in pulmonary smooth muscle cells in both time- and concentration-dependent manners. Mutations that truncated the C-terminus abolished this response. Taken together, these results suggest a model in which proliferative effect of c-Src by vasoactive molecules is balanced by opposing effect of BMP signaling in basal state, and the loss of this balance due to BMPR2 mutations leads to increased c-Src activity and subsequently cell growth.

Decreased methylation and transcription repressor Sp3 up-regulated human monoamine oxidase (MAO) B expression during Caco-2 differentiation.

Monoamine oxidase (MAO) A and B catalyze the oxidative deamination of neuroactive and dietary monoamines such as serotonin, tyramine, and phenylethylamine. Here we show that MAO B, but not MAO A, gene expression was induced during Caco-2 cell differentiation; thus this cell line was used as a model system to study the gene regulation unique for MAO B. Luciferase and gel shift assays showed that transcription factors Sp1 and Sp3 binding to -246 and -99 bp were responsible for the observed gene activation. Overexpression of Sp3 inhibited the induction of MAO B gene by Sp1, and the expression of Sp3 was decreased during Caco-2 cell differentiation. Computer analysis revealed a putative CpG island containing 22 potential CpG methylation sites between -261 and -58 bp. In vitro methylation of MAO B promoter with 5-aza-2'-deoxycytidine, a DNA methyltransferase inhibitor, up-regulated MAO B gene expression in both HeLa and Caco-2 cells. Sodium bisulfite sequencing showed a gradually reduced methylation of the CpG sites during Caco-2 cell differentiation. These results suggested that MAO B gene expression is selectively induced by a decreased Sp3/Sp1 ratio and reduced DNA methylation. This new information may provide insights on the tissue-specific expression of these two isoenzymes.

Activation of human monoamine oxidase B gene expression by a protein kinase C MAPK signal transduction pathway involves c-Jun and Egr-1.

Monoamine oxidases (MAO) A and B deaminate a number of biogenic amines. Aberrant expression of MAO is implicated in several psychiatric and neurogenerative disorders. In this study, we have shown that phorbol 12-myristate 13-acetate (PMA) increases human MAO B, but not MAO A, gene expression. The sequence between -246 and -225 bp consists of overlapping binding sites (Sp1/Egr-1/Sp1) that are recognized by Sp1, Sp3, and PMA-inducible Egr-1 is essential for PMA activation. PMA transiently increases egr-1 and c-jun gene expression. Mutation studies show that Egr-1 and c-Jun transactivate the MAO B promoter and increase endogenous MAO B transcripts via the Sp1/Egr-1/Sp1 overlapping binding sites. Sp3 inhibits Sp1 and Egr-1 activation of MAO B gene expression. c-fos gene expression was increased by PMA but not involved in MAO B gene transcription. Furthermore, protein kinase C inhibitor blocks the PMA-dependent activation of MAO B. Co-transfection of the MAO B promoter with dominant negative forms of Ras, Raf-1, MEKK1, MEK1, MEK3, MEK7, ERK2, JNK1, and p38/RK inhibit the PMA-dependent activation of the MAO B promoter. These results indicate that MAO B expression is selectively induced by the activation of protein kinase C and MAPK signaling pathway and that c-Jun and Egr-1 appear to be the ultimate targets of this regulation.