Long-term persistence of Ebola virus (EBOV) in immunologically privileged sites has been implicated in recent outbreaks of Ebola virus disease (EVD) in Guinea and the Democratic Republic of Congo. ...This study was designed to understand how the acute course of EVD, convalescence, and host immune and genetic factors may play a role in prolonged viral persistence in semen.
A cohort of 131 male EVD survivors in Liberia were enrolled in a case-case study. "Early clearers" were defined as those with 2 consecutive negative EBOV semen test results by real-time reverse-transcription polymerase chain reaction (rRT-PCR) ≥2 weeks apart within 1 year after discharge from the Ebola treatment unit or acute EVD. "Late clearers" had detectable EBOV RNA by rRT-PCR >1 year after discharge from the Ebola treatment unit or acute EVD. Retrospective histories of their EVD clinical course were collected by questionnaire, followed by complete physical examinations and blood work.
Compared with early clearers, late clearers were older (median, 42.5 years; P < .001) and experienced fewer severe clinical symptoms (median 2, P = .006). Late clearers had more lens opacifications (odds ratio, 3.9 95% confidence interval, 1.1-13.3; P = .03), after accounting for age, higher total serum immunoglobulin G3 (IgG3) titers (P = .005), and increased expression of the HLA-C*03:04 allele (0.14 .02-.70; P = .007).
Older age, decreased illness severity, elevated total serum IgG3 and HLA-C*03:04 allele expression may be risk factors for the persistence of EBOV in the semen of EVD survivors. EBOV persistence in semen may also be associated with its persistence in other immunologically protected sites, such as the eye.
Here we present a novel and robust method for the identification of protein S-nitrosylation sites in complex protein mixtures. The approach utilizes the cysteinyl affinity resin to selectively enrich ...S-nitrosylated peptides reduced by ascorbate followed by nanoscale liquid chromatography tandem mass spectrometry. Two alkylation agents with different added masses were employed to differentiate the S-nitrosylation sites from the non-S-nitrosylation sites. We applied this approach to MDA-MB-231 cells treated with Angeli’s salt, a nitric oxide donor that has been shown to inhibit breast tumor growth and angiogenesis. A total of 162 S-nitrosylation sites were identified and an S-nitrosylation motif was revealed in our study. The 162 sites are significantly more than the number reported by previous methods, demonstrating the efficiency of our approach. Our approach will further facilitate the functional study of protein S-nitrosylation in cellular processes and may reveal new therapeutic targets.
Menaquinone is used for transporting electrons and is essential for the aerobic and anaerobic respiratory systems of all pathogens and prokaryotes. Many Gram-positive bacteria use only menaquinone in ...the electron transport system. Thus, menaquinone biosynthesis is a potential target for the development of inhibitors against bacteria including drug-resistant pathogens.
After modeling, synthesis and in vitro testing, we determined that 7-methoxy-2-naphthol-based inhibitors targeted the MenA enzyme of the menaquinone biosynthesis pathway. The developmental compounds 1 and 2 were active against Mycobacterium tuberculosis and methicillin-resistant Staphylococcus aureus with a minimal inhibitory concentration of 3-5 μg/ml.
Nontraditional bicyclic inhibitors, compounds 1 and 2 could serve as lead compounds for the development of an antimicrobial agent, with activities against M. tuberculosis and methicillin-resistant S. aureus.
The taxonomic status of the bacterium Wolbachia persica is described, and based on the evidence presented, transfer of this species to the genus Francisella as Francisella persica comb. nov. is ...proposed. This reclassification is supported by data generated from genomic comparisons of W. persica ATCC VR-331T ( = FSC845T = DSM 101678T) to other near neighbours, including Francisella tularensis subsp. novicida. The full-length 16S rRNA gene sequence of strain ATCC VR-331T had 98.5 % nucleotide identity to the cognate gene in F. tularensis, with the highest similarity to subspecies novicida. Phylogenetic trees of full-length 16S rRNA gene, gyrA and recA sequences from species of the genera Wolbachia (class Alphaproteobacteria) and Francisella (class Gammaproteobacteria) indicated that W. persica ATCC VR-331T was most closely related to members of the genus Francisella and not Wolbachia. Local collinear blocks within the chromosome of strain ATCC VR-331T had considerable similarity with F. tularensis subsp. novicida, but not with any Wolbachia strain. The genomes of strain ATCC VR-331T and F. tularensis subsp. novicida Utah 112T ( = ATCC 15482T) contained an average nucleotide identity mean of 88.72 % and median of 89.18 %. Importantly, the genome of strain ATCC VR-331T contained one Francisella Pathogenicity Island, similar to F. tularensis subsp. novicida, as well as the Francisella-specific gene fopA1 and F. tularensis-specific genes fopA2 and lpnA (also referred to as tul4). In contrast to the obligate intracellular genus Wolbachia, strain ATCC VR-331T and facultative intracellular Francisella can replicate in specialized cell-free media. Collectively, these results demonstrate that Wolbachia persica should be reclassified in the genus Francisella as Francisella persica comb. nov. The type strain of Francisella persica comb. nov. is ATCC VR-331T ( = FSC845T = DSM 101678T). An emended description of the family Francisellaceae is also provided.
The kinetic chromogenic endotoxin assay measures the release of p-nitroaniline from the chromogenic peptide substrate Ac-IEAR-pNA. As part of our project to purify large quantities of human ...butyrylcholinesterase (HuBChE), we evaluated pure HuBChE for endotoxin levels. We found that HuBChE contributed up to 90% of the yellow p-nitroaniline product in a standard endotoxin assay through the catalytic hydrolysis of Ac-IEAR-pNA with a rate constant of 0.016 min−1 and a Km of 2.9 mM in potassium phosphate buffer pH 7.0 at 24 °C. Thus, endotoxin concentrations for native BChE are artificially high in the kinetic chromogenic assay. Destruction of HuBChE catalytic activity by boiling yields endotoxin concentrations that more accurately reflect the endotoxin concentration in purified HuBChE preparations.
•Human BChE cleaves the chromogenic substrate used in endotoxin assays.•Endotoxin levels are artificially high for native BChE.•Boiled HuBChE has up to 90% lower endotoxin levels.
Of the estimated 1.4 million salmonella infections that occur each year in the United States, most are in children and the elderly and approximately 600 are fatal.
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Septicemia complicates at least 7 ...percent of the 40,000 culture-confirmed infections that are reported each year.
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Although antibiotics are not essential for the treatment of most cases of salmonellosis, they can be lifesaving in persons with invasive disease. Expanded-spectrum cephalosporins (e.g., ceftriaxone) are commonly used to treat salmonella infections in children because of their pharmacodynamic properties and the very low prevalence of resistance to these agents. Since 1991, salmonella species that are . . .
The prevalence of resistant organisms has increased throughout the last decade. In order to determine best treatment strategies, studies on reimplantation must address this population. The purpose of ...this study is to compare the results of delayed reconstruction for total joint infections caused by methicillin-sensitive and methicillin-resistant organisms. This investigation included 46 patients (21 men, 25 women) with an average age 67 years (range, 36-86 years) who had a two-staged reimplantation after periprosthetic total joint infection. Thirty-seven patients cultured methicillin-sensitive organisms and nine cultured methicillin-resistant organisms. Treatment included resection, aggressive debridement, 6 weeks of parenteral antibiotics, and subsequent debridement and reimplantation. Treatment was considered successful if the patient was disease free at last followup (average 5 years, range 2-10 years). The efficacy of two-stage reconstruction was similar for infections with sensitive and resistant bacteria. Only 3 of 46 patients failed delayed reconstruction requiring further treatment. Failure rates with regards to infections with sensitive bacteria were 5.4% (2 of 37 infections) and resistant bacteria 11.1% (1 of 9 infections), which was not significant. The current study shows clinical success of two-stage reimplantation for patients with methicillin-resistant bacteria complicating total joint arthroplasty.
: We have described recently an acetylcholinesterase (AChE) knockout mouse. While comparing the tissue distribution of AChE and butyrylcholinesterase (BChE), we found that extraction buffers ...containing Triton X‐100 strongly inhibited mouse BChE activity. In contrast, buffers with Tween 20 caused no inhibition of BChE. Conventional techniques grossly underestimated BChE activity by up to 15‐fold. In Tween 20 buffer, the intestine, serum, lung, liver, and heart had higher BChE than AChE activity. Only brain had higher AChE than BChE activity in AChE +/+ mice. These findings contradict the dogma, based mainly on observations in Triton X‐100 extracts, that BChE is a minor cholinesterase in animal tissues. AChE +/‐ mice had 50% of normal AChE activity and AChE ‐/‐ mice had none, but all mice had similar levels of BChE activity. BChE was inhibited by Triton X‐100 in all species tested, except rat and chicken. Inhibition was reversible and competitive with substrate binding. The active site of rat BChE was unique, having an arginine in place of leucine at position 286 (human BChE numbering) in the acyl‐binding pocket of the active site, thus explaining the lack of inhibition of rat BChE by Triton X‐100. The generally high levels of BChE activity in tissues, including the motor endplate, and the observation that mice live without AChE, suggest that BChE has an essential function in nullizygous mice and probably in wild‐type mice as well.
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