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The COVID-19 pandemic has led to the rapid development of a plethora of molecular diagnostic assays with real-time polymerase chain reaction (RT-PCR) at the forefront. In this review, we will discuss the history and utility of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) molecular diagnostics and the associated current and future regulatory process in Europe. We will assess the performance characteristics of a range of the most common SARS-CoV-2 molecular tests currently used in Europe with a focus on as rapid molecular platforms, stand-alone RT-PCR kits, the role of low-throughput and high-throughput end-to-end testing platforms, and the rapidly evolving field of SARS-CoV-2 variant of concern identification.

Here we describe a retrospective clinical evaluation of the QIAGEN

Objectives SARS-CoV-2 emerged in South Asia in 2019 and has resulted in a global pandemic. Public Health England (PHE) Manchester rapidly escalated testing for SARS-CoV-2 in the highest COVID-19 incidence location in England. The results of the PHE Manchester SARS-CoV-2 surveillance during the first wave are presented. Methods Retrospective data were collected for patients fitting the PHE SARS-CoV-2 case definition from 11th February to 31st August 2020. Respiratory tract, tissue, faecal, fluid and cerebrospinal (CSF) samples were tested for SARS-CoV-2 by a semi-quantitative real-time reverse-transcription PCR. Results Of the 204,083 tests for SARS-CoV-2, 18,011 were positive demonstrating a positivity of 8.90%. Highest positivity was in nasal swabs (20.99%) followed by broncheo-alveolar lavage samples (12.50%). None of the faecal, fluid or CSF samples received were positive for SARS-CoV-2. Conclusions There was a high incidence of SARS-CoV-2 patients in the North-West of England during the first UK wave of the Covid-19 pandemic. Highest positivity rate was in nasal specimens suggesting this is the optimum sample type within this dataset for detecting SARS-CoV-2. Further studies are warranted to assess the utility of testing faecal, fluid and CSF samples. Rapid escalation of testing via multiple platforms was required to ensure prompt diagnosis and isolate infected cases to reduce transmission of the virus.

Two apparently linked fatal cases of pneumococcal meningitis were investigated. Pneumolysin PCR performed on blood and cerebrospinal fluid was positive in a culture-negative case. A second case yielded Streptococcus pneumoniae from blood culture. Multilocus sequence typing performed on DNA extracted from case 1 (specimens) and case 2 (isolate) revealed identical sequence types (ST53) substantiating the link between them.

In a study of Campylobacter infection in northwestern England, 2003–2006, C. jejuni multilocus sequence type (ST)–45 was associated with early summer onset and was the most prevalent C. jejuni type in surface waters. ST-45 is likely more adapted to survival outside a host, making it a key driver of transmission between livestock, environmental, and human settings.

Multilocus sequence typing (MLST), an accurate and phylogenetically robust characterization method for population studies of Campylobacter , was applied to Campylobacter jejuni isolates ( n = 297) from the fecal samples of cattle from five dairy farms in Cheshire, United Kingdom, collected throughout 2003. The population dynamics of the C. jejuni strains, as identified by the occurrence of sequence types and clonal complexes, demonstrated variations within and between cattle populations over time. Three clonal lineages have emerged to predominate among the cattle isolates, namely, the ST-61 complex (24.2%), ST-21 complex (23.6%), and ST-42 complex (20.5%). This provided further evidence that the ST-61 clonal complex may present a cattle-adapted C. jejuni genotype. In addition, the ST-42 clonal complex may also represent an important cattle-associated genotype. Strong geographical associations for these genotypes were also found among the farms. This is the first longitudinal study and the largest study to date for C. jejuni involving cattle populations using MLST for accurate strain characterization. This study shows the important associations between cattle and C. jejuni clonal complexes ST-61, ST-21, and ST-42, and it suggests that cattle and/or dairy products are likely to be a source of the human Campylobacter gastroenteritis caused by such genotypes. The reported findings have significant implications for the design of effective intervention strategies for disease control and prevention.

Infections associated with Neisseria meningitidis are a major public health problem in England, Wales, and Northern Ireland. Currently, over 40% of cases are confirmed directly from clinical specimens using PCR-based methodologies without an organism being isolated. A nested/seminested multilocus sequence typing (MLST) system was developed at the Health Protection Agency Meningococcal Reference Unit to allow strain characterization beyond the serogroup for cases confirmed by PCR only. This system was evaluated on a panel of 20 meningococcus-positive clinical specimens (3 cerebrospinal fluid and 17 blood samples) from different patients containing various concentrations of meningococcal DNA that had corresponding N. meningitidis isolates. In each case, the sequence type generated from the clinical specimens matched that produced from the corresponding N. meningitidis isolate; the sensitivity of the MLST system was determined to be less than 12 genome copies per PCR. The MLST system was then applied to 15 PCR meningococcus-positive specimens (2 cerebrospinal fluid and 13 blood samples), each from a different patient, involved in three case clusters (two serogroup B and one serogroup W135) for which no corresponding N. meningitidis organisms had been isolated. In each case, an MLST sequence type was generated, allowing the accurate assignment of individual cases within each of the case clusters. In summary, the adaptation of the N. meningitidis MLST to a sensitive nested/seminested format for strain characterization directly from clinical specimens provides an important tool for surveillance and management of meningococcal infection.

To determine the epidemiological relationship between pneumococci of serotype 9V, with reduced susceptibility to ciprofloxacin, penicillin and erythromycin, referred to the Reference Laboratory during 1997-2001.Isolates were characterized by multilocus sequence typing (MLST), PFGE, and sequencing of parC and gyrA. Relevant clinical data were sought.Forty-eight isolates were received from nine laboratories in England, but 35 (73%) were from one laboratory in Birmingham, and were mostly from elderly patients receiving ofloxacin or ciprofloxacin for respiratory infections. There were two quinolone resistance phenotypes, with ciprofloxacin, moxifloxacin and gemifloxacin MICs of 8-32, 0.5-1 and 0.125-0.25 mg/L, and 64-256, 4-16 and 1-4 mg/L, respectively. Each of three isolates from the former group had mutations in parC, whereas each of nine isolates from the more resistant group had mutations in both parC and gyrA. Several also had increased quinolone efflux. Typing of 27 quinolone-resistant isolates showed that eight were indistinguishable from the epidemic Spain9V-3 (ST156) clone, while the remainder belonged to a novel but related type (ST609), that differed from Spain9V-3 at 2/7 alleles (2 bp changes in aroE and 1 bp change in gdh). Both MLST types were represented among isolates with high- and low-level quinolone resistance. Three of five serotype 9V isolates from Birmingham, with reduced susceptibility to penicillin and erythromycin, and ciprofloxacin MICs of 1-2 mg/L, belonged to MLST type ST609, while another was indistinguishable from the Spain9V-3 clone. Review of records of 32 patients from Birmingham indicated that some isolates were nosocomial, whereas others were acquired in the community.In the late 1990s, a quinolone-resistant strain, clonally related to Spain9V-3, emerged in England, principally in Birmingham.

Between January 2000 and March 2001,Streptococcus pneumoniaewere isolated from the blood of 56 patients admitted to a single district general hospital in the South-East of England. The serotype and antibiotic susceptibility were determined for all isolates and, for those resistant to erythromycin, the presence or absence of themef(A) anderm(B) genes was determined by PCR. Multi-locus sequence typing, along with PFGE, was undertaken on all isolates resistant to penicillin or erythromycin and a group of antibiotic-susceptible isolates, to identify whether globally distributed pneumococcal clones, as described by the Pneumococcal Molecular Epidemiology Network (PMEN), were present in the study population. Three serotype 9V penicillin-resistant isolates were identified as belonging to the Spain9V-3 clone, while 14 erythromycin-resistant isolates of serotype 14 belonged to the England14-9 clone. A single multi-resistant isolate of serotype 6B, was found to be a single-locus variant of the Spain6B-2 clone. All 14 erythromycin-resistant serotype 14 isolates possessed themef(A) gene, while the single multi-resistant isolate possessed theerm(B) gene. These findings confirm the wide distribution and clinical impact of PMEN clones, which accounted for all of the penicillin and erythromycin resistance observed amongst invasive isolates in a district general hospital over a 15-month period.