Multiplex PCR assay for identification of three major pathogenic Vibrio spp., Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus

A multiplex PCR assay was developed based on atpA-sequence diversification for molecular identification of 3 major pathogenic Vibrio species: Vibrio cholerae, Vibrio parahaemolyticus, and Vibrio vulnificus. It specifically identified them from among 133 strains of various Vibrio species and other genera, and was applicable for testing seawater, suggesting its usefulness.     

Mediastinal ultrasonography for the assessment of mediastinal lymph node metastases in lung cancer patients

Using an ultrasonic probe inserted into the mediastinum during cervical mediastinoscopy, mediastinal ultrasonography (USM) was performed on 63 patients with lung cancer. The patients with a small peripheral mass of less than 2 cm in diameter, according to the chest X-ray results, and with mediastinal lymph nodes smaller than 1 cm in their short axes as determined by computed tomography (CT), were excluded from this study. An analysis of the areas under the receiver operating characteristic curves derived from CT and USM showed that USM was superior (P=0.043) to CT in terms of the diagnosis for mediastinal lymph node metastases, when the short axis dimension of mediastinal lymph nodes was employed for the diagnosis of metastases. The reason for this is that 97% of the mediastinal lymph nodes imaged by USM were located vertically along the body axis of the patient, and hence USM imaged the true short axis of the node in many cases. Our results indicate that USM is useful for performing a safe biopsy of lymph nodes during mediastinoscopy as well as for obtaining a clear imaging of the subcarinal nodes, which are inaccessible by normal cervical mediastinoscopy.     

The impact of ferritin fluctuations on stable hemoglobin levels in hemodialysis patients

Background

Hemoglobin (Hb) cycling in patients with renal anemia might be associated with a higher mortality rate. We investigated the association of factors relating serum ferritin and dose of erythropoiesis-stimulating agents (ESAs) with Hb levels.

Methods

We measured Hb and ferritin levels every month in 266 hemodialysis (HD) patients for 12?months.

Results

The standard deviation (SD) and residual SD (RSD) (liner regression of Hb or ferritin SD values) values of Hb were significantly correlated with ferritin SD or RSD values, respectively. The percentage achievement of target Hb in the target-ferritin group was significantly higher than in the high-amplitude fluctuation ferritin group. Ferritin SD and RSD values in patients with oral or no iron supplementation were significantly lower than those who received intravenous iron.

Conclusion

Iron storage varies over a relatively wide range in HD patients, and this variation is closely associated with Hb cycling. The stability of iron storage and ESA dosage is important for maintaining stable Hb levels.     

Elimination of Viruses (Human Immunodeficiency, Hepatitis B, Vesicular Stomatitis and Sindbis Viruses) from an Intravenous Immunoglobulin Preparation

More than 10(4) plaque-forming units (pfu)/ml of HIV are inactivated during the alcohol fractionation step from plasma to fraction (Fr)-II+III, greater than 10(4) pfu/ml is inactivated from Fr-II+III to Fr-II and greater than 10(4) pfu/ml is inactivated during the polyethylene glycol (PEG) fractionation process from Fr-II+III to intravenous IgG (IVIG). The total inactivation rate from plasma to IVIG via Fr-II+III or Fr-II was calculated to be greater than 10(8) or 10(12), respectively. The PEG fractionation method produces an intact and unmodified IVIG. In addition, the PEG fractionation method at a low ionic strength was found to be effective for the elimination of greater than 10(5) units of other viruses, including hepatitis B, vesicular stomatitis and Sindbis viruses.     

Subtilase Cytotoxin Enhances Escherichia coli Survival in Macrophages by Suppression of Nitric Oxide Production through the Inhibition of NF-κB Activation

Subtilase cytotoxin (SubAB), which is produced by certain strains of Shiga-toxigenic Escherichia coli (STEC), cleaves an endoplasmic reticulum (ER) chaperone, BiP/Grp78, leading to induction of ER stress and caspase-dependent apoptosis. SubAB alters the innate immune response. SubAB pretreatment of macrophages inhibited lipopolysaccharide (LPS)-induced production of both monocyte chemoattractant protein 1 (MCP-1) and tumor necrosis factor α (TNF-α). We investigated here the mechanism by which SubAB inhibits nitric oxide (NO) production by mouse macrophages. SubAB suppressed LPS-induced NO production through inhibition of inducible NO synthase (iNOS) mRNA and protein expression. Further, SubAB inhibited LPS-induced IκB-α phosphorylation and nuclear localization of the nuclear factor-κB (NF-κB) p65/p50 heterodimer. Reporter gene and chromatin immunoprecipitation (ChIP) assays revealed that SubAB reduced LPS-induced NF-κB p65/p50 heterodimer binding to an NF-κB binding site on the iNOS promoter. In contrast to the native toxin, a catalytically inactivated SubAB mutant slightly enhanced LPS-induced iNOS expression and binding of NF-κB subunits to the iNOS promoter. The SubAB effect on LPS-induced iNOS expression was significantly reduced in macrophages from NF-κB1 (p50)-deficient mice, which lacked a DNA-binding subunit of the p65/p50 heterodimer, suggesting that p50 was involved in SubAB-mediated inhibition of iNOS expression. Treatment of macrophages with an NOS inhibitor or expression of SubAB by E. coli increased E. coli survival in macrophages, suggesting that NO generated by macrophages resulted in efficient killing of the bacteria and SubAB contributed to E. coli survival in macrophages. Thus, we hypothesize that SubAB might represent a novel bacterial strategy to circumvent host defense during STEC infection.     

Identification of a distinct lineage of aviadenovirus from crane feces

Viruses are believed to be ubiquitous; however, the diversity of viruses is largely unknown because of the bias of previous research toward pathogenic viruses. Deep sequencing is a promising and unbiased approach to detect viruses from animal-derived materials. Although cranes are known to be infected by several viruses such as influenza A viruses, previous studies targeted limited species of viruses, and thus viruses that infect cranes have not been extensively studied. In this study, we collected crane fecal samples in the Izumi plain in Japan, which is an overwintering site for cranes, and performed metagenomic shotgun sequencing analyses. We detected aviadenovirus-like sequences in the fecal samples and tentatively named the discovered virus crane-associated adenovirus 1 (CrAdV-1). We determined that our sequence accounted for approximately three-fourths of the estimated CrAdV-1 genome size (33,245 bp). The GC content of CrAdV-1 genome is 34.1%, which is considerably lower than that of other aviadenoviruses. Phylogenetic analyses revealed that CrAdV-1 clusters with members of the genus Aviadenovirus, but is distantly related to the previously identified aviadenoviruses. The protein sequence divergence between the DNA polymerase of CrAdV-1 and those of other aviadenoviruses is 45.2–46.8%. Based on these results and the species demarcation for the family Adenoviridae, we propose that CrAdV-1 be classified as a new species in the genus Aviadenovirus. Results of this study contribute to a deeper understanding of the diversity and evolution of viruses and provide additional information on viruses that infect cranes, which might lead to protection of the endangered species of cranes.

     

Effects of addition of arginine, cystine, and glycine to the bovine milk-simulated amino acid mixture on the level of plasma and liver cholesterol in rats

The effects of the addition of varying levels of arginine (Arg), cystine (Cys), and glycine (Gly) to the bovine milk-simulated amino acid mixture on the levels of plasma and liver cholesterol were investigated in rats. The diets containing a high amount of Cys lowered significantly the level of plasma cholesterol as the amount of Cys in diets increased. The serum high density lipoprotein (HDL)-cholesterol level and fecal excretion of acidic steroids were higher in rats fed the Cys diets than in those fed the Arg and Gly diets. No differences, however, was observed in the content of liver cholesterol. Liver triglyceride contents elevated significantly on feeding the Arg and Cys diets. Furthermore, liver phospholipid contents elevated significantly on feeding the Arg diet but lowered on feeding the Cys diet. Therefore, these results indicated that the feeding of a high amount of Cys lowers the plasma cholesterol levels as the result of the enhanced conversion of cholesterol to bile acids.