Diagnostic strategy used to establish etiologies of encephalitis in a prospective cohort of patients in England.

The laboratory diagnostic strategy used to determine the etiology of encephalitis in 203 patients is reported. An etiological diagnosis was made by first-line laboratory testing for 111 (55%) patients. Subsequent testing, based on individual case reviews, resulted in 17 (8%) further diagnoses, of which 12 (71%) were immune-mediated and 5 (29%) were due to infection. Seventy-five cases were of unknown etiology. Sixteen (8%) of 203 samples were found to be associated with either N-methyl-d-aspartate receptor or voltage-gated potassium channel complex antibodies. The most common viral causes identified were herpes simplex virus (HSV) (19%) and varicella-zoster virus (5%), while the most important bacterial cause was Mycobacterium tuberculosis (5%). The diagnostic value of testing cerebrospinal fluid (CSF) for antibody was assessed using 139 samples from 99 patients, and antibody was detected in 46 samples from 37 patients. Samples collected at 14 to 28 days were more likely to be positive than samples taken 0 to 6 days postadmission. Three PCR-negative HSV cases were diagnosed by the presence of virus-specific antibody in the central nervous system (CNS). It was not possible to make an etiological diagnosis for one-third of the cases; these were therefore considered to be due to unknown causes. Delayed sampling did not contribute to these cases. Twenty percent of the patients with infections with an unknown etiology showed evidence of localized immune activation within the CNS, but no novel viral DNA or RNA sequences were found. We conclude that a good standard of clinical investigation and thorough first-line laboratory testing allows the diagnosis of most cases of infectious encephalitis; testing for CSF antibodies allows further cases to be diagnosed. It is important that testing for immune-mediated causes also be included in a diagnostic algorithm.

Causality in acute encephalitis: defining aetiologies.

Defining the causal relationship between a microbe and encephalitis is complex. Over 100 different infectious agents may cause encephalitis, often as one of the rarer manifestations of infection. The gold-standard techniques to detect causative infectious agents in encephalitis in life depend on the study of brain biopsy material; however, in most cases this is not possible. We present the UK perspective on aetiological case definitions for acute encephalitis and extend them to include immune-mediated causes. Expert opinion was primarily used and was supplemented by literature-based methods. Wide usage of these definitions will facilitate comparison between studies and result in a better understanding of the causes of this devastating condition. They provide a framework for regular review and updating as the knowledge base increases both clinically and through improvements in diagnostic methods. The importance of new and emerging pathogens as causes of encephalitis can be assessed against the principles laid out here.

Implications for HIV testing policy derived from combining data on voluntary confidential testing with viral sequences and serological analyses.

OBJECTIVES: Laboratory, clinical and sequence-based data were combined to assess the differential uptake of voluntary confidential HIV testing (VCT) according to risk and explore the occurrence of HIV transmission from individuals with recently acquired HIV infection, before the diagnostic opportunity. METHODS: Between 1999 and 2002, nearly 30,000 anonymous tests for previously undiagnosed HIV infection were conducted among men who have sex with men (MSM) attending 15 sentinel sexually transmitted infection (STI) clinics in England, Wales and Northern Ireland. Using a serological testing algorithm, undiagnosed HIV-infected men were categorised into those with recent and non-recent infection. VCT uptake was compared between HIV-negative, recently HIV-infected and non-recently HIV-infected men. A phylogenetic analysis of HIV pol sequences from 127 recently HIV-infected MSM was conducted to identify instances in which transmission may have occurred before the diagnostic opportunity. RESULTS: HIV-negative MSM were more likely to receive VCT at clinic visits compared with undiagnosed HIV-infected MSM (56% (14,020/24,938) vs 31% (335/1072); p<0.001). Recently HIV-infected MSM were more likely to receive VCT compared with those with non-recent infections (42% (97/229) vs 28% (238/844); p<0.001). 22% (95/425) of undiagnosed HIV-infected MSM with STI received VCT. Phylogenetic analysis revealed at least seven transmissions may have been generated by recently HIV-infected MSM: a group that attended STI clinics soon after seroconversion. CONCLUSIONS: The integration of clinical, laboratory and sequence-based data reveals the need for specific targeting of the recently HIV exposed, and those with STI, for VCT. VCT promotion alone may be limited in its ability to prevent HIV transmission.

Genetic variants of parvovirus B19 identified in the United Kingdom: implications for diagnostic testing.

BACKGROUND: Discrepant results in diagnostic parvovirus B19 PCR assays have been observed with strains showing nucleotide sequence variation. OBJECTIVES AND STUDY DESIGN: To perform phylogenetic analysis on two parvovirus B19 strains that gave discrepant PCR results. RESULTS: One strain was found to be genotype 2; the second strain was genotype 3. CONCLUSIONS: Parvovirus B19 genotypes 2 and 3 strains were identified in diagnostic samples of UK origin following the investigation of discrepant PCR results. More structured investigations are needed to estimate the prevalence of these variants. In the meantime, diagnostic PCR results should be interpreted cautiously when they are at variance with serological testing. Manufacturers of PCR kits for the detection of B19 sequences will need to consider re-designing their primers.

The behaviour of microbes.

Viruses and bacteria have complex interactions with their hosts, beyond mere replication in them. They range from those that are detrimental, to others that may be non-pathogenic or even beneficial. Molecular techniques can help to unravel these interactions, sometimes revealing phenomena that benefit host as well as microbial populations.

Virulence typing of Escherichia coli using microarrays.

We describe a microarray based broad-range screening technique for Escherichia coli virulence typing. Gene probes were amplified by PCR from a plasmid bank of characterised E. coli virulence genes and were spotted onto a glass slide to form an array of capture probes. Genomic DNA from E. coli strains which were to be tested for the presence of these virulence gene sequences was labelled with fluorescent cyanine dyes by random amplification and then hybridised against the array of probes. The hybridisation, washing and data analysis conditions were optimised for glass slides, and the applicability of the method for identifying the presence of the virulence genes was determined using reference strains and clinical isolates. It was found to be a sensitive screening method for detecting virulence genes, and a powerful tool for determining the pathotype of E. coli. It will be possible to expand and automate this microarray technique to make it suitable for rapid and reliable diagnostic screening of bacterial isolates.

Contribution of influenza and respiratory syncytial virus to community cases of influenza-like illness: an observational study.

BACKGROUND: Respiratory syncytial virus (RSV) is an important cause of lower-respiratory-tract infection in children and elderly people, but its effect in other age-groups is uncertain. We did a community-based observational study of RSV infection in community-dwelling individuals of all ages who presented to general practices in the UK with influenza-like illnesses during three successive winters (1995-96, 1996-97, and 1997-98). METHODS: Nasopharyngeal swabs routinely submitted for virological surveillance were examined by multiplex reverse transcription PCR for influenza A and B viruses and RSV A and B, and findings were related to the clinical incidence of influenza-like illness and acute bronchitis at that time. RSV strains identified were compared with those obtained from hospital admissions. FINDINGS: 480 RSV and 709 influenza viruses were identified from a total of 2226 swabs submitted. Both types of virus were found in all age-groups for between 12 and 20 weeks in each winter. RSV A accounted for 60% of RSV detections. Similar strains of RSV were present in hospital and community patients within the same year, but there were different lineages each year. INTERPRETATION: In individuals diagnosed with influenza-like illness, there is a substantial potential for confusion between illnesses caused by influenza and those caused by RSV. The burden of illness attributable to each needs to be clarified to define optimum management routines.