Mili Estee Torok

Rapid Whole-Genome Sequencing for Investigation of a Suspected Tuberculosis Outbreak

ABSTRACT Two Southeast Asian students attending the same school in the United Kingdom presented with pulmonary tuberculosis. An epidemiological investigation failed to link the two cases, and drug resistance profiles of the Mycobacterium tuberculosis isolates were discrepant. Whole-genome sequencing of the isolates found them to be genetically identical, suggesting a missed transmission event.

Prevalence and characterization of human mecC methicillin-resistant Staphylococcus aureus isolates in England

Objectives There are limited data available on the epidemiology and prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in the human population that encode the recently described mecA homologue, mecC. To address this knowledge gap we undertook a prospective prevalence study in England to determine the prevalence of mecC among MRSA isolates. Patients and methods Three hundred and thirty-five sequential MRSA isolates from individual patients were collected from each of six clinical microbiology laboratories in England during 2011–12. These were tested by PCR or genome sequencing to differentiate those encoding mecA and mecC. mecC-positive isolates were further characterized by multilocus sequence typing, spa typing, antimicrobial susceptibility profile and detection of PBP2a using commercially available kits. Results Nine out of the 2010 MRSA isolates tested were mecC positive, indicating a prevalence among MRSA in England of 0.45% (95% CI 0.24%–0.85%). The remainder were mecA positive. Eight out of these nine mecC MRSA isolates belonged to clonal complex 130, the other being sequence type 425. Resistance to non-β-lactam antibiotics was rare among these mecC MRSA isolates and all were phenotypically identified as MRSA using oxacillin and cefoxitin according to BSAC disc diffusion methodology. However, all nine mecC isolates gave a negative result using three different commercial PBP2a detection assays. Conclusions mecC MRSA are currently rare among MRSA isolated from humans in England and this study provides an important baseline prevalence rate to monitor future changes, which may be important given the increasing prevalence of mecC MRSA reported in Denmark.

Rapid whole-genome sequencing of bacterial pathogens in the clinical microbiology laboratory—pipe dream or reality?

The ability to perform rapid, high-throughput whole-genome sequencing using bench-top platforms represents a step-change in capabilities for diagnostic and public health microbiology laboratories. As the cost of sequencing continues to decline, the challenge will be to define when and where to apply this technology. This article reviews its potential applications in the clinical microbiology laboratory and discusses the current barriers to implementation.

Use of Vitek 2 Antimicrobial Susceptibility Profile To Identify mecC in Methicillin-Resistant Staphylococcus aureus

ABSTRACT The emergence of mecC methicillin-resistant Staphylococcus aureus (MRSA) poses a diagnostic challenge for clinical microbiology laboratories. Using the Vitek 2 system, we tested a panel of 896 Staphylococcus aureus isolates and found that an oxacillin-sensitive/cefoxitin-resistant profile had a sensitivity of 88.7% and a specificity of 99.5% for the identification of mecC MRSA isolates. The presence of the mecC gene, determined by bacterial whole-genome sequencing, was used as the gold standard. This profile could provide a zero-cost screening method for identification of mecC-positive MRSA strains.

Read and assembly metrics inconsequential for clinical utility of whole-genome sequencing in mapping outbreaks

Read and assembly metrics inconsequential for clinical utility of whole-genome sequencing in mapping outbreaks

Genome-based characterization of hospital-adapted Enterococcus faecalis lineages

Vancomycin-resistant Enterococcus faecalis (VREfs) is an important nosocomial pathogen1,2. We undertook whole genome sequencing of E. faecalis associated with bloodstream infection in the UK and Ireland over more than a decade to determine the population structure and genetic associations with hospital adaptation. Three lineages predominated in the population, two of which (L1 and L2) were nationally distributed, and one (L3) geographically restricted. Genome comparison with a global collection identified that L1 and L3 were also present in the USA, but were genetically distinct. Over 90% of VREfs belonged to L1–L3, with resistance acquired and lost multiple times in L1 and L2, but only once followed by clonal expansion in L3. Putative virulence and antibiotic resistance genes were over-represented in L1, L2 and L3 isolates combined, versus the remainder. Each of the three main lineages contained a mixture of vancomycin-resistant and -susceptible E. faecalis (VSEfs), which has important implications for infection control and antibiotic stewardship.

Complex Routes of Nosocomial Vancomycin-Resistant Enterococcus faecium Transmission Revealed by Genome Sequencing

Summary We generated genome sequence data from 293 Enterococcus faecium isolates from patients with bacteremia spanning 7 years in one hospital. Cases were connected to numerous transmission routes, the complexity of which placed them beyond detection by conventional infection control methods.

Longitudinal genomic surveillance of MRSA in the UK reveals transmission patterns in hospitals and the community

Longitudinal genomic and epidemiological surveillance of methicillin-resistant Staphylococcus aureus in the UK reveals extensive transmission in hospitals and the community. On the trail of MRSA Genome sequencing of methicillin-resistant Staphylococcus aureus (MRSA) has been successfully applied to investigate suspected outbreaks. Coll et al. now extend its application to the genomic surveillance of MRSA in samples from 1465 people identified over a 12-month period by a diagnostic laboratory in the East of England. This analysis identified 173 putative outbreaks involving 598 patients and included hospital outbreaks, those spanning the hospital and community, and community outbreaks among people registered with the same medical practice or living in the same household or long-term care facility. This study illustrates that sequencing is a powerful tool that could be used to identify infectious disease outbreaks as they happen. Genome sequencing has provided snapshots of the transmission of methicillin-resistant Staphylococcus aureus (MRSA) during suspected outbreaks in isolated hospital wards. Scale-up to populations is now required to establish the full potential of this technology for surveillance. We prospectively identified all individuals over a 12-month period who had at least one MRSA-positive sample processed by a routine diagnostic microbiology laboratory in the East of England, which received samples from three hospitals and 75 general practitioner (GP) practices. We sequenced at least 1 MRSA isolate from 1465 individuals (2282 MRSA isolates) and recorded epidemiological data. An integrated epidemiological and phylogenetic analysis revealed 173 transmission clusters containing between 2 and 44 cases and involving 598 people (40.8%). Of these, 118 clusters (371 people) involved hospital contacts alone, 27 clusters (72 people) involved community contacts alone, and 28 clusters (157 people) had both types of contact. Community- and hospital-associated MRSA lineages were equally capable of transmission in the community, with instances of spread in households, long-term care facilities, and GP practices. Our study provides a comprehensive picture of MRSA transmission in a sampled population of 1465 people and suggests the need to review existing infection control policy and practice.

Strongyloides stercoralis hyperinfection in a patient treated for multiple myeloma

A 53-year-old Ghanaian man, resident in the UK for many years, was diagnosed with myeloma and commenced treatment with cyclophosphamide, thalidomide and dexamethasone. Towards the end of his first cycle he presented with fever, abdominal pain, constipation and anorexia. There was no recent travel history. Blood tests on admission to hospital showed anaemia (haemoglobin concentration 87 g/l), lymphopenia (white cell count 8·4 9 10/l, lymphocytes 0·7 9 10/l, neutrophils 6·5 9 10/l, eosinophils 0·3 9 10/l) and profound hyponatraemia (plasma sodium 115 mmol/l) as a consequence of the syndrome of inappropriate anti-diuretic hormone release (SIADH). C-reactive protein was mildly raised (24 mg/l). Following an episode of haematemesis, gastroscopy revealed gastritis and duodenitis. Computed tomography (CT) of his abdomen showed diffuse thickening of the entire small bowel and proximal colon (left). He deteriorated clinically with type-one respiratory failure requiring ventilation and intensive care unit (ICU) support. A CT chest scan was consistent with acute respiratory distress syndrome. The diagnosis of Strongyloides stercoralis hyperinfection was made following gastric biopsy, where the filariform larvae of the parasite were seen (right). Parasites were subsequently observed in bronchial washes. Treatment with oral ivermectin was started, but later converted to subcutaneous administration as persistent gastric bleeding abrogated enteric absorption. Slow improvement was made thereafter and the ivermectin dose was gradually tapered. The patient was discharged from hospital on day 42 and remains well. On review of his previous blood results, an unexplained mild eosinophilia (2·2 9 10/l) had been present prior to commencement of chemotherapy. Strongyloides stercoralis infection is endemic in the tropics and subtropics and can persist subclinically for many years. Strongyloides hyperinfection is a serious and well recognized complication of immunosuppression, which is characteristically not accompanied by eosinophilia. Chronic Strongyloides infection should be excluded in patients with eosinophilia originating from, or having travelled to, endemic areas prior to initiating immunosuppressive therapy or steroids. Parenteral ivermectin is unlicensed for human use but is an effective alternative if the oral route is unavailable.

Community outbreaks of group A Streptococcus revealed by genome sequencing

The frequent occurrence of disease outbreaks in humans caused by group A Streptococcus (GAS) is an on-going public health threat. Conventional bacterial typing methods lack the discriminatory power to confidently confirm or refute outbreaks in hospital and community settings. Microbial whole genome sequencing (WGS) provides a potential solution to this, but, there has been limited population-based surveillance with accompanying sequence data. We performed retrospective genomic surveillance of 93 clinical GAS isolates from individuals in a defined geographic region. Detailed clinical information was obtained for closely related clusters of isolates. Genomic sequence data was contextualised through comparison with international data. We identified 18 different emm genotypes within our bacterial population, and revealed both highly diverse and closely related isolates. This high level of diversity was maintained even in the context of international sequence data. We also identified two emm1 clusters, and one emm3 cluster, of closely-related isolates that differed only by 1 to 4 single nucleotide polymorphisms. Analysis of clinical information identified no healthcare associated contact between patients, indicating cryptic community transmission. Our findings suggest that genomic surveillance of GAS would increase detection of transmission and highlight opportunities for intervention.