Towards comprehensive understanding of bacterial genetic diversity: large-scale amplifications in Bordetella pertussis and Mycobacterium tuberculosis.

Bacterial genetic diversity is often described solely using base-pair changes despite a wide variety of other mutation types likely being major contributors. Tandem duplication/amplifications are thought to be widespread among bacteria but due to their often-intractable size and instability, comprehensive studies of these mutations are rare. We define a methodology to investigate amplifications in bacterial genomes based on read depth of genome sequence data as a proxy for copy number. We demonstrate the approach with Bordetella pertussis, whose insertion sequence element-rich genome provides extensive scope for amplifications to occur. Analysis of data for 2430 B. pertussis isolates identified 272 putative amplifications, of which 94 % were located at 11 hotspot loci. We demonstrate limited phylogenetic connection for the occurrence of amplifications, suggesting unstable and sporadic characteristics. Genome instability was further described in vitro using long-read sequencing via the Nanopore platform, which revealed that clonally derived laboratory cultures produced heterogenous populations rapidly. We extended this research to analyse a population of 1000 isolates of another important pathogen, Mycobacterium tuberculosis. We found 590 amplifications in M. tuberculosis, and like B. pertussis, these occurred primarily at hotspots. Genes amplified in B. pertussis include those involved in motility and respiration, whilst in M. tuberuclosis, functions included intracellular growth and regulation of virulence. Using publicly available short-read data we predicted previously unrecognized, large amplifications in B. pertussis and M. tuberculosis. This reveals the unrecognized and dynamic genetic diversity of B. pertussis and M. tuberculosis, highlighting the need for a more holistic understanding of bacterial genetics.

Impact of antibiotic allergy labels on patient outcomes in a tertiary paediatric hospital.

AIMS: Antibiotic allergies are reported in 5-15% of children. This study aimed to evaluate the impact of common ß-lactam antibiotic allergy labels (AALs) on hospital treatment, focusing on length of stay and appropriateness of antibiotic prescribing. METHODS: This was a retrospective cohort study over 21 months at the Royal Children's Hospital Melbourne, Australia. A subset of children with the most common ß-lactam allergies, and who required admission for intravenous antibiotics over a 12-month period, was analysed for appropriateness of prescribing. Non-allergic patients were matched to evaluate associations between AALs and hospital treatment. RESULTS: There were 98 912 children admitted over the study period, of whom 938 (1%) had at least one AAL on first admission. Of all encounters, 5145 (2.5%) were for children with AALs. The most common AALs were to amoxicillin and amoxicillin-clavulanic acid combinations (40.8%), cefalexin (14.4%) and trimethoprim-sulfamethoxazole (9.7%). For the subset, there were 66 admissions for children who required intravenous antibiotics. Documentation was adequate for 27% of AALs. Inappropriate prescribing occurred in almost half (47%). Hospital stay was longer for children with AALs (median 4.7 days; IQR 2.3-9.2) compared to non-allergic controls (median 3.9 days; IQR 1.9-6.8; P = .02). Children with AALs were more likely to receive restricted antibiotics (aOR 3.03; 95% CI, 1.45-6.30; P = .003). CONCLUSION: This is the first study to demonstrate high rates of inappropriate prescribing in children with AALs. Children with AALs were significantly more likely to receive restricted antibiotics and had a longer length of stay compared with non-allergic controls.

Author Correction: A robust benchmark for detection of germline large deletions and insertions.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

A robust benchmark for detection of germline large deletions and insertions.

New technologies and analysis methods are enabling genomic structural variants (SVs) to be detected with ever-increasing accuracy, resolution and comprehensiveness. To help translate these methods to routine research and clinical practice, we developed a sequence-resolved benchmark set for identification of both false-negative and false-positive germline large insertions and deletions. To create this benchmark for a broadly consented son in a Personal Genome Project trio with broadly available cells and DNA, the Genome in a Bottle Consortium integrated 19 sequence-resolved variant calling methods from diverse technologies. The final benchmark set contains 12,745 isolated, sequence-resolved insertion (7,281) and deletion (5,464) calls ≥50 base pairs (bp). The Tier 1 benchmark regions, for which any extra calls are putative false positives, cover 2.51 Gbp and 5,262 insertions and 4,095 deletions supported by ≥1 diploid assembly. We demonstrate that the benchmark set reliably identifies false negatives and false positives in high-quality SV callsets from short-, linked- and long-read sequencing and optical mapping.

Risk factors for avascular necrosis after closed reduction for developmental dysplasia of the hip.

PURPOSE: The purpose of this study was to identify and evaluate risk factors of avascular necrosis (AVN) after closed treatment for developmental dysplasia of the hip (DDH). METHODS: A retrospective review of children diagnosed with DDH at a tertiary-care children's hospital between 1986 and 2009 was performed. The presence of AVN was assessed according to Salter's classification system. RESULTS: Eighty-two affected hips in 70 children with an average age of 10 months at closed reduction (range 1-31 months) and 5 years (range 2-19 years) of follow-up met the inclusion criteria. Twenty-nine (of 82, 35 %) affected hips developed AVN. The use of pre-reduction traction (p = 0.019) increased the risk of AVN, while preoperative Pavlik harness or brace trial (p = 0.28), presence of ossific nucleus at the time of closed reduction (p = 0.16), and adductor tenotomy (p = 0.37) were not significant factors. Laterality (right vs. left) was also not a significant risk factor (p = 0.75), but patients who underwent closed reduction for bilateral DDH were less likely to develop AVN (p = 0.027). Overall, the degree of abduction did not affect the rate of AVN (p = 0.87). However, in patients treated with closed reduction younger than 6 months of age, the rate of AVN was increased with abduction ≥50° (9/15, 60 %) compared to abduction <50° (0/8, 0 %) (p = 0.007). Patients who developed AVN were more likely to require subsequent surgery (p = 0.034) and more likely to report a fair/poor clinical outcome (p = 0.049). CONCLUSIONS: The risk of AVN (35 %) following closed reduction and spica casting for DDH is high. The degree of abduction in spica casts appears to be a risk factor in patients ≤6 months old. The authors recommend that abduction in spica casts should be limited to <50° in children younger than 6 months of age. LEVEL OF EVIDENCE: IV.