BackgroundDifferential RNA-sequencing (dRNA-seq) is indispensable for determination of primary transcriptomes. However, using dRNA-seq data to map transcriptional start sites (TSSs) and promoters genome-wide is a bioinformatics challenge. We performed dRNA-seq of Bradyrhizobium japonicum USDA 110, the nitrogen-fixing symbiont of soybean, and developed algorithms to map TSSs and promoters.ResultsA specialized machine learning procedure for TSS recognition allowed us to map 15,923 TSSs: 14,360 in free-living bacteria, 4329 in symbiosis with soybean and 2766 in both conditions. Further, we provide proteomic evidence for 4090 proteins, among them 107 proteins corresponding to new genes and 178 proteins with N-termini different from the existing annotation (72 and 109 of them with TSS support, respectively). Guided by proteomics evidence, previously identified TSSs and TSSs experimentally validated here, we assign a score threshold to flag 14 % of the mapped TSSs as a class of lower confidence. However, this class of lower confidence contains valid TSSs of low-abundant transcripts. Moreover, we developed a de novo algorithm to identify promoter motifs upstream of mapped TSSs, which is publicly available, and found motifs mainly used in symbiosis (similar to RpoN-dependent promoters) or under both conditions (similar to RpoD-dependent promoters). Mapped TSSs and putative promoters, proteomic evidence and updated gene annotation were combined into an annotation file.ConclusionsThe genome-wide TSS and promoter maps along with the extended genome annotation of B. japonicum represent a valuable resource for future systems biology studies and for detailed analyses of individual non-coding transcripts and ORFs. Our data will also provide new insights into bacterial gene regulation during the agriculturally important symbiosis between rhizobia and legumes.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-016-2602-9) contains supplementary material, which is available to authorized users.

The use of an assay with rapid results reduced the use of antistaphylococcal therapy among patients who did not have S. aureus bacteremia; it also decreased the use of MRS drug therapy and led to earlier appropriate therapy among patients with MSSA bacteremia.

Urinary tract infections (UTI) are one of the most common indications for antibiotic prescriptions in the outpatient setting. Given rising rates of antibiotic resistance among uropathogens, antibiotic stewardship is critically needed to improve outpatient antibiotic use, including in outpatient clinics (primary care and specialty clinics) and emergency departments.

The use of an assay with rapid results reduced the use of antistaphylococcal therapy among patients who did not have S. aureus bacteremia; it also decreased the use of MRS drug therapy and led to earlier appropriate therapy among patients with MSSA bacteremia.

Letters to the Editor Identification of Methicillin-Resistant or Methicillin-Susceptible Staphylococcus aureus in Blood Cultures and Wound Swabs by GeneXpert ᰔ Until a decade ago, clinicians could use epidemiological clues to select empirical therapy for methicillin-susceptible Staphylococcus aureus (MSSA) or methicillin-resistant S. aureus (MRSA) (5). The emergence of MRSA as a community pathogen and the documentation of the inferiority of nonbeta-lactam antibiotics in treating MSSA bacteremia greatly complicate initial antibiotic choice (2-4, 7, 8, 9). Early identification and determination of antibiotic susceptibility might help focus initial antibiotic therapy. We compared a multiplex PCR that identifies MSSA and MRSA to standard microbiologic techniques for evaluating the results of blood cultures (BCs) and wound swab (WS) cultures. Blood was cultured in BacT/Alert, and drug susceptibility was determined with a Vitek 2 system (both from BioMerieux, Durham, NC). For BCs judged to contain gram-positive cocci in clusters (GPCCl), 1-ml aliquots were centrifuged (2 min at 3,000 rpm) to remove charcoal, and the supernatant was studied in a GeneXpert system (Cepheid, Sunnyvale, CA). WS samples were streaked to standard media (blood, chocolate, McConkey, and colistin-nalidixic acid) and then studied in the GeneXpert system within 48 h of collection. GeneXpert realtime PCR detects proprietary sequences of the S. aureus protein A gene, the staphylococcal cassette chromosome, and the methicillin resistance element (1). Of 223 blood samples, 68 yielded S. aureus by culture, 47 with MRSA and 21 with MSSA. PCR correctly identified 67/68 (98.5%) S. aureus isolates (Tables 1 and 2), including 46/47 (97.9%) MRSA and 21/21 (100%) MSSA isolates. No BC (155/155; 100%) that contained GPCCl without S. aureus contained S. aureus by PCR. Of 321 WS samples, 106 yielded MRSA and 51 MSSA by culture. PCR identified 104/106 (98.1%) MRSA isolates correctly but misidentified 2 MRSA isolates as MSSA (Table 1). Of 51 MSSA isolates, 47 (92.2%) were identified correctly, 3 incorrectly as MRSA, and 1 incorrectly as no S. aureus by PCR.

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ObjectivesThe objective of our study was to evaluate the impact of a multifaceted stewardship intervention on adherence to the evidence-based practice guidelines on treatment of uncomplicated cystitis in primary care. We hypothesised that our intervention would increase guideline adherence in terms of antibiotic choice and duration of treatment.DesignA preintervention and postintervention comparison with a contemporaneous control group was performed. During the first two study periods, we obtained baseline data and performed interviews exploring provider prescribing decisions for cystitis at both clinics. During the third period in the intervention clinic only, the intervention included a didactic lecture, a decision algorithm and audit and feedback. We used a difference-in-differences analysis to determine the effects of our intervention on the outcome and guideline adherence to antibiotic choice and duration.SettingTwo family medicine clinics (one intervention and one control) were included.ParticipantsAll female patients with uncomplicated cystitis attending the study clinics between 2016 and 2019.ResultsOur sample included 932 visits representing 812 unique patients with uncomplicated cystitis. The proportion of guideline-adherent antibiotic regimens increased during the intervention period (from 33.2% (95% CI 26.9 to 39.9) to 66.9% (95% CI 58.4 to 74.6) in the intervention site and from 5.3% (95% CI 2.3 to 10.1) to 17.0% (95% CI 9.9 to 26.6) in the control site). The increase in guideline adherence was greater in the intervention site compared with the control site with a difference-in-differences of 22 percentage points, p=0.001.ConclusionA multifaceted intervention increased guideline adherence for antibiotic choice and duration in greater magnitude than similar trends at the control site. Future research is needed to facilitate scale-up and sustainability of case-based audit and feedback interventions in primary care.

Background In hospitalized people with HIV (PWH) there is an increased risk of mortality from COVID-19 among hospitalized PWH as compared to HIV-negative individuals. Evidence suggests that tocilizumab—a humanized monoclonal interleukin (IL)-6 receptor inhibitor (IL-6ri) antibody—has a modest mortality benefit when combined with corticosteroids in select hospitalized COVID-19 patients who are severely ill. Data on clinical outcomes after tocilizumab use in PWH with severe COVID-19 are lacking. Case presentation We present a multinational case series of 18 PWH with COVID-19 who were treated with IL-6ri’s during the period from April to June 2020. Four patients received tocilizumab, six sarilumab, and eight received an undocumented IL-6ri. Of the 18 patients in the series, 4 (22%) had CD4 counts < 200 cells/mm3; 14 (82%) had a suppressed HIV viral load. Eight patients (44%), all admitted to ICU, were treated for secondary infection; 5 had a confirmed organism. Of the four patients with CD4 counts < 200 cells/mm3, three were treated for secondary infection, with 2 confirmed organisms. Overall outcomes were poor—12 patients (67%) were admitted to the ICU, 11 (61%) required mechanical ventilation, and 7 (39%) died. Conclusions In this case series of hospitalized PWH with COVID-19 and given IL-6ri prior to the common use of corticosteroids, there are reports of secondary or co-infection in severely ill patients. Comprehensive studies in PWH, particularly with CD4 counts < 200 cells, are warranted to assess infectious and other outcomes after IL-6ri use, particularly in the context of co-administered corticosteroids.