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Since STEC infection does not always lead to severe symptoms, it is possible that many more cases were associated with this cluster and largely went unrecognized.
Since STEC infection does not always lead to severe symptoms, it is possible that many more cases were associated with this cluster and largely went unrecognized.Control of biological populations remains a critical goal to address the challenges facing ecosystems and agriculture and those posed by human disease, including pests, parasites, pathogens and invasive species. A particular architecture of the CRISPR/Cas biotechnology - a gene drive - has the potential to modify or eliminate populations on a massive scale. Super-Mendelian inheritance has now been demonstrated in both fungi and metazoans, including disease vectors such as mosquitoes. Studies in yeast and fly model systems have developed a number of molecular safeguards to increase biosafety and control over drive systems in vivo, including titration of nuclease activity, anti-CRISPR-dependent inhibition and use of non-native DNA target sites. We have developed a CRISPR/Cas9 gene drive in Saccharomyces cerevisiae that allows for the safe and rapid examination of alternative drive designs and control mechanisms. In this study, we tested whether non-homologous end-joining (NHEJ) had occurred within diploid cells displaying a loss of the target allele following drive activation and did not detect any instances of NHEJ within multiple sampled populations. We also demonstrated successful multiplexing using two additional non-native target sequences. Furthermore, we extended our analysis of 'resistant' clones that still harboured both the drive and target selection markers following expression of Streptococcus pyogenes Cas9; de novo mutation or NHEJ-based repair could not explain the majority of these heterozygous clones. Finally, we developed a second-generation gene drive in yeast with a guide RNA cassette integrated within the drive locus with a near 100 % success rate; resistant clones in this system could also be reactivated during a second round of Cas9 induction.Group A streptococcus (GAS) is a rare cause of bacterial meningitis in children and is associated with a high cerebral complication rate. In this case report, we present a 9-year-old girl with GAS meningitis complicated with cerebritis. Clear guidelines about choice of treatment and indications of follow-up by imaging tests are lacking, making GAS meningitis unpredictable and difficult to treat. Eventually, we found 25 paediatric cases of GAS meningitis presented in the literature and reviewed their treatment choices, outcomes and follow-up by imaging tests. Penicillin and ceftriaxone are most preferred for the treatment of GAS meningitis and adding rifampicin to the antibiotic treatment could be of potential benefit. When considering the duration of antibiotic treatment and follow-up by imaging tests, no clear recommendations were found. We found that GAS meningitis is associated with higher mortality and cerebral complication rates compared to other, more common, bacterial causes of meningitis in children. This should alert the clinician to consider imaging tests routinely, even if the patient improves clinically. We advise clinicians to routinely evaluate for possible cerebral complications through magnetic resonance imaging (MRI) scans. When cerebral complications are found, antibiotic treatment should be prolonged and adding rifampicin to the antibiotic regime may be considered.Middle East respiratory syndrome coronavirus (MERS-CoV) is a novel zoonotic coronavirus that was identified in 2012. MERS-CoV infection in humans can result in an acute, severe respiratory disease and in some cases multi-organ failure; the global mortality rate is approximately 35 %. The MERS-CoV spike (S) protein is a major target for neutralizing antibodies in infected patients. The MERS-CoV microneutralization test (MNt) is the gold standard method for demonstrating prior infection. However, this method requires the use of live MERS-CoV in biosafety level 3 (BSL-3) containment. The present work describes the generation and validation of S protein-bearing vesicular stomatitis virus (VSV) pseudotype particles (VSV-MERS-CoV-S) in which the VSV glycoprotein G gene has been replaced by the luciferase reporter gene, followed by the establishment of a pseudoparticle-based neutralization test to detect MERS-CoV neutralizing antibodies under BSL-2 conditions. Using a panel of human sera from confirmed MERS-CoV patients, the VSV-MERS-CoV particle neutralization assay produced results that were highly comparable to those of the microneutralization test using live MERS-CoV. The results suggest that the VSV-MERS-CoV-S pseudotype neutralization assay offers a highly specific, sensitive and safer alternative method to detect MERS-CoV neutralizing antibodies in human sera.
Standard culture methods may fail to detect the causative agents of bacterial infection for various reasons including specimen collection after antibiotic administration, or when standard techniques or environmental conditions are not appropriate for growth of the microorganisms. Conventional 16S rRNA gene sequencing is sometimes a useful alternative technique for identification of bacteria, but is confounded by polymicrobial infection. We present a case of a patient who developed a serious neurological infection for which causative oral flora organisms were observed by microscopy, failed to culture but were identified by next-generation DNA sequencing.
A male in his forties developed sinus pain and congestion, followed by facial and eye pain, and several weeks later acute-onset confusion and neck stiffness. Cerebrospinal fluid examination revealed pleocytosis and several bacterial morphologies, which were subsequently identified by next-generation sequencing as oral flora constituents
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Oral flora can cause meningoencephalitis and brain abscess formation if translocation occurs by injury or surgical procedures. Next-generation sequencing is often not available at healthcare facilities, or when available may not have been validated for a wide spectrum of specimen sources, but is available at reference laboratories and should be considered when routine methods fail to provide a diagnosis for serious infections.
Oral flora can cause meningoencephalitis and brain abscess formation if translocation occurs by injury or surgical procedures. Next-generation sequencing is often not available at healthcare facilities, or when available may not have been validated for a wide spectrum of specimen sources, but is available at reference laboratories and should be considered when routine methods fail to provide a diagnosis for serious infections.The present report describes a case of infective endocarditis complicated with aortic root abscess caused by Cardiobacterium hominis in a 56-year-old man. C. hominis is a microaerophilic, pleomorphic Gram-negative bacillus and member of the Haemophilus species, Aggregatibacter actinomycetemcomitans , C. hominis , Eikenella corrodens and Kingella kingae (HACEK) group, a group of bacteria known to be a rare cause of endocarditis. With prompt diagnosis and initiation of antimicrobial and surgical management, a successful outcome was achieved.The type II toxin-antitoxin (TA) modules, mazEF and relBE, in Streptococcus mutans have been implicated in stress response, antibiotic tolerance and persister cell formation. However, how S. mutans regulates these systems to prevent unwanted toxin activation and persister cell formation is unclear. In this study, we provide evidence that ClpP is required for the proteolytic regulation of these TA systems and persister cell formation in S. mutans following antibiotic challenge. A persister viability assay showed that S. mutans UA159 (WT) formed a larger quantity of persister cells than its isogenic mutant ΔclpP following antibiotic challenge. However, the lux reporter assay revealed that clpP deletion did not affect the transcriptional levels of mazEF and relBE, since no significant differences (P>0.05) in the reporter activities were detected between the wild-type and ΔclpP background. Instead, all antibiotics tested at a sub-minimum inhibitory concentration (sub-MIC) induced transcriptional levels of mazEF and relBE operons. We then examined the protein profiles of His-tagged MazE and RelB proteins in the UA159 and ΔclpP backgrounds by Western blotting analysis. The results showed that S. mutans strains grown under non-stress conditions expressed very low but detectable levels of MazE and RelB antitoxin proteins. Antibiotics at sub-MICs induced the levels of the MazE and RelB proteins, but the protein levels decreased rapidly in the wild-type background. In contrast, a stable level of MazE and RelB proteins could be detected in the ΔclpP mutant background, suggesting that both proteins accumulated in the ΔclpP mutant. We conclude that ClpP is required for the proteolytic regulation of cellular levels of the MazE and RelB antitoxins in S. mutans , which may play a critical role in modulating the TA activities and persister cell formation of this organism following antibiotic challenge.We present a case of Ruminococcus gnavus sepsis in a woman suffering from multiple myeloma and myelodysplastic syndrome. R. gnavus , a Gram-positive coccus and a gut commensal, has been described in nine cases of infection in the literature, with most infections having occurred in patients with either gastrointestinal symptoms or prosthesis infections. In this case, R gnavus was identified by mass spectrometry, and showed susceptibility to penicillin, meropenem, tetracycline, metronidazole and clindamycin. The patient was successfully treated initially with intravenous piperacillin/tazobactam and metronidazole, and then switched to oral penicillin and metronidazole. The cause of infection is hypothesized to have been a shift in the gut microbiota towards an excess growth of R. gnavus caused by immunosuppression, and bacterial translocation across a vulnerable mucosal barrier due to prednisolone treatment and severe thrombocytopenia.Strains of Rotarix, a live attenuated monovalent oral rotavirus vaccine, replicate in the intestine and are shed for about one month in immunocompetent recipients. The current study aimed to identify genetic changes of shed strains to reveal any significant mutations and their clinical impact on recipients. Stool samples of recipients of the first dose of Rotarix were sequentially collected for one month from the day of administration. Sequence analyses of the VP7 gene in eight recipients revealed five amino acid substitutions. Among them, two were observed in aa123, which is located in antigenic region 7-1a. Since there were no associated clinical symptoms, the genetic changes were unlikely to have caused reversion of pathogenicity of vaccine strain. Of interest, the virus in one case became closer to wild-type rotavirus via an amino acid change at aa123 occurring 14 days after administration, which might have resulted from multiple replications and long-term shedding of the vaccine strain.Automated, high-throughput technologies are becoming increasingly common in microbiome studies to decrease costs and increase efficiency. However, in microbiome studies, small differences in methodology - including storage conditions, wet lab methods, sequencing platforms and data analysis - can influence the reproducibility and comparability of data across studies. There has been limited testing of the effects of high-throughput methods, including microfluidic PCR technologies. In this paper, we compare two extraction methods (the QIAamp DNA Stool Mini Kit and the MoBio PowerSoil DNA Isolation kit), two taq polymerase enzymes (MyTaq HS Red Mix and Accustart II PCR ToughMix), two primer sets (V3-V4 and V4-V5) and two amplification methods (a common two-step PCR protocol and amplicon library preparation on the Fluidigm Access Array system that allows automated multiplexing of primers). Gut microbial community profiles were significantly affected by all variables. While there were no significant differences in alpha diversity measured between the two extraction methods, there was an effect of extraction method on community composition measured by unweighted UniFrac distances.
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