Population exposure to a novel influenza A virus over three waves of infection.

BACKGROUND: The influenza A(H1N1)2009 virus has been spreading throughout the world since April 2009. Since then, several studies have been undertaken to measure the frequency of antibodies that react against the virus. Microneutralisation assays have regularly been used for these analyses, and titres of ≥40 have conventionally been taken to represent significant levels of antibodies (this significance is derived from it being four times the minimum level of antibodies that the assay can detect rather an established correlate of protection). However a microneutralisation titre that correlates with protection against influenza A(H1N1)2009 has not been established. OBJECTIVES: Analysing influenza A(H1N1)2009 antibody seroprevalence in Scotland at multiple timepoints, and in different age groups and geographical locations, and comprehensively describing the spread of the virus in Scotland (taken alongside previously published data). This study presents for the first time the effects of a novel influenza virus on a naïve population that has been followed from the initial outbreak to a time when the majority of the population have reactive antibodies. STUDY DESIGN: A microneutralisation titre ≥10 represents the minimum level of antibodies detectable by the assay. Blood samples (taken in April 2009 and April 2010 in Edinburgh (n=400 each year), and in February 2011 in Aberdeen, Edinburgh, Glasgow, and Inverness (n=1600)) were tested for the presence of influenza A(H1N1)2009 antibodies at this titre. This represents an effective indicator of the proportion of a population who have been exposed to the virus. RESULTS: Following the 2010/2011 influenza season, there is evidence of exposure to influenza A(H1N1)2009 in approximately four fifths of the Scottish population. CONCLUSIONS: This study provides impetus to the call for further research in establishing robust correlates of susceptibility to influenza infection and the development of clinical illness, provides useful information for future outbreaks, and is relevant to public health policy in planning for future influenza seasons.

Influenza A(H1N1)2009 antibody seroprevalence in Scotland following the 2010/11 influenza season.

Following the 2010/11 influenza season, we determined the age- and location-specific seroprevalence of antibodies against the influenza A(H1N1)2009 virus in Scotland. Samples were analysed by microneutralisation assay. Age/seropositivity profiles varied significantly between cities. The increases in seroprevalence relative to the previous influenza season (2009/10) were similar across age groups and geographic locations. However, the increased seropositivity in older adults appeared to be driven by exposure to vaccination, indicating significantly lower levels of infection than in younger age groups.

BK viremia surveillance after kidney transplant: single-center experience during a change from cyclosporine-to lower-dose tacrolimus-based primary immunosuppression regimen.

BACKGROUND: The aim was to report our experience of BK viremia surveillance after kidney transplant during a period of change from cyclosporine (CyA)-to lower-dose tacrolimus (Tac)-based primary immunosuppression regimens. METHODS: In a prospective single-center observational cohort study, 68 consecutive patients received renal transplant during the period when we used a CyA-based primary immunosuppression regimen and 66 after we changed to a lower-dose Tac-based regimen. Testing for BK viremia by quantitative polymerase chain reaction assay was performed at least monthly for a minimum of 1 year. RESULTS: Thirty-nine (29.1%) patients developed BK viremia and 2 (1.5%) developed BK nephropathy. The actuarial time to BK viremia was shorter in patients receiving CyA/mycophenolate mofetil (MMF)/prednisolone (Pred) compared with Tac/MMF/Pred (P=0.04) and primary immunosuppression with CyA/MMF/Pred was the only independent predictor of BK viremia (hazard ratio 1.95; P=0.047). Comparing patients who experienced BK viremia and those who did not, there was no difference in incidence of acute rejection (20.5% vs. 25.3%; P=0.56) or estimated glomerular filtration rate at 12 months (48.8 vs. 49.9 mL/min/1.73 m(2)), but the incidence of ureteric stenosis was higher (10.3% vs. 1.1%; P=0.01). CONCLUSIONS: Our data demonstrate a lower incidence of BK viremia in patients on lower-dose Tac compared with CyA-based primary immunosuppression in contrast to previous studies, and provide further support for the association between BK virus and ureteric complications.

2009 pandemic influenza A(H1N1) virus in Scotland: geographically variable immunity in Spring 2010, following the winter outbreak.

We determined the age- and location-specific seroprevalence of antibodies against 2009 pandemic influenza A(H1N1) virus in Scotland following the first two waves of infection. Serum samples collected following the winter outbreak were analysed by microneutralisation assay. The proportion of positive sera varied significantly between cities and, in the case of Inverness, between age groups (with younger adults more likely to be positive than older individuals). This study demonstrates that older people are no longer more likely to have antibodies against the virus than younger adults.

Prevalence of chronic viral hepatitis in people of south Asian ethnicity living in England: the prevalence cannot necessarily be predicted from the prevalence in the country of origin.

The prevalence of hepatitis B and hepatitis C in immigrant communities is unknown. Immigrants from south Asia are common in England and elsewhere, and the burden of viral hepatitis in these communities is unknown. We aimed to determine the prevalence of viral hepatitis in immigrants from south Asia living in England, and we therefore undertook a community-based testing project in such people at five sites in England. A total of 4998 people attending community centres were screened for viral hepatitis using oral fluid testing. The overall prevalence of anti-hepatitis C virus (HCV) in people of south Asian origin was 1.6% but varied by country of birth being 0.4%, 0.2%, 0.6% and 2.7% in people of this ethnic group born in the UK, India, Bangladesh and Pakistan, respectively. The prevalence of hepatitis B surface antigen was 1.2%-0.2%, 0.1%, 1.5% and 1.8% in people of this ethnic group born in the UK, India, Bangladesh and Pakistan, respectively. Analysis of risk factors for HCV infection shows that people from the Pakistani Punjab and those who have immigrated recently are at increased risk of infection. Our study suggests that migrants from Pakistan are at highest risk of viral hepatitis, with those from India at low risk. As prevalence varies both by country and region of origin and over time, the prevalence in migrant communities living in western countries cannot be easily predicted from studies in the country of origin.

Hepatitis B virus: origin and evolution.

The pathogenesis of hepatitis B virus (HBV) is complex and it appears that molecular variants play a role in this process. HBV undergoes numerous rounds of error prone production within an infected host. The resulting quasispecies are heterogeneous and in the absence of archaeological records of past infection, the evolution of HBV can only be inferred indirectly from its epidemiology and by genetic analysis. This review gathered the controversies about the HBV origin and factors influencing its quasispecies. Also, it provided some evidence on how HBV genotypes correlated with human history and patterns of migration. It is our belief that this topic deserves further attention and thus it is likely that more critical research work will be performed to elucidate the unknown mechanisms and processes in this area.

Development of a multiplex real-time RT-PCR that allows universal detection of influenza A viruses and simultaneous typing of influenza A/H1N1/2009 virus.

On June 11, 2009, the World Health Organization declared that the influenza A/H1N1/2009 virus had become the first influenza pandemic of the 21st century. Rapid detection and differentiation from seasonal and avian influenza would be beneficial for patient management and infection control. It was the aim of this study to develop a real-time RT-PCR that can detect all influenza A viruses and offer simultaneous typing for influenza A/H1N1/2009. This would be a useful addition to existing diagnostic protocols for influenza A. Its routine use would allow laboratories to screen out influenza A/H1N1/2009 positive samples rapidly and would reduce overall testing costs.

Simultaneous detection and quantitation of cytomegalovirus, Epstein-Barr virus, and adenovirus by use of real-time PCR and pooled standards.

Quantitative real-time PCR has become the most widely used preemptive approach for managing cytomegalovirus (CMV), Epstein-Barr virus (EBV), and adenovirus infections in immunosuppressed patients. These three assays are normally available as separate tests, each using five quantitation standards that are tested in duplicate. We have developed an adenovirus-CMV-EBV triplex assay that uses one set of five pooled quantitative standards, tested singly rather than in duplicate. This test demonstrated a sensitivity and an accuracy of quantitation equivalent to those of our previous single tests and was shown to be able to detect mixed infections with no loss in sensitivity. This assay is now in routine use in our laboratory and has considerably simplified the work flow of the laboratory, with a resultant improvement in sample turnaround time and significantly reduced costs.

Using multiplex real time PCR in order to streamline a routine diagnostic service.

An increasing number of virology laboratories are now utilising in house real time PCR assays as the frontline diagnostic tests. As the number of tests on offer increases the natural progression from this will be to rationalise their service via multiplexing. Since 2003 we have introduced a large number of qualitative and quantitative multiplex real time PCR assays into our routine testing service. This paper describes the development of the multiplex assays, the problems encountered and the resultant benefits to the routine service.

The effectiveness of national influenza vaccination policies for at-risk populations over 5 seasons in a Scottish general practice.

AIM: To determine the effectiveness of serial influenza vaccination. SCOPE: Studied in a Scottish GP population, the overall seroresponse rate increased with annual influenza vaccinations and after 5 years it increased from 45.1% to 93.3% for influenza virus A (H1) and from 48.4% to 98.3% for influenza virus A (H3). However, there was little boosting effect with further doses after becoming a seroresponder. The pre-vaccination titres were significantly higher in previous year's seroresponders compared to non-responders. CONCLUSIONS: The policy of annual vaccination is supported by our data in order to increase the disappointing response rate after one dose. However, the lack of a boosting response with subsequent doses and the significant residual immunity after becoming a seroresponder suggests a prior serological immunity check in order to better direct the vaccine supply (in the years of no antigenic drift), to those who need it most.

Emergence of a new norovirus variant in Scotland in 2006.

For each month between January 2005 and August 2006, a representative number of outbreaks was examined using nucleic acid sequence analysis. Using this method, we showed that an increase in norovirus activity coincided with the emergence of a new GII genotype 4 variant, which by March 2006 was detected in the majority of health boards in Scotland.

Contamination with PCR detectable virus in a virus isolation quality assurance panel.

External quality control schemes are an essential part of the quality assurance of all diagnostic virology laboratories. There are a few providers of nucleic acid detection quality control panels. Consequently, diagnostic laboratories may test panels that are developed specifically for virus isolation by nucleic acid detection methods. Here, we report on a recent simulated eye swab panel from a National external quality assessment service, which was meant for virus isolation but which we tested by polymerase chain reactions assays. The results suggest that the samples had been contaminated at source leading to difficulty in interpretation of the panel and a poor score.

Hospital transmission of hepatitis B virus in the absence of exposure prone procedures.

In February and in June 1998, two people developed acute hepatitis B following in-patient care in a district general hospital. Initial enquiries indicated their infections were not attributable to staff undertaking exposure-prone procedures (EPPs). We report the findings and implications of the subsequent investigation: a multi-disciplinary, multi-agency investigation, including molecular epidemiological analysis. Occupational Health records showed that staff involved in EPPs with the patients were HBsAg negative. No contact between the patients was identified nor were there failures in sterilization. The patients' HBV strains were identical, indicating a common source. A total of 231 out of 232 staff who might have treated either patient were tested for HBsAg; the remaining doctor, working abroad, was HBsAg- and HBeAg-positive and had the same HBV strain as the patients. On two occasions the doctor's hand had been cut while breaking glass vials, but there was no documentation linking these events to the two patients. The doctor had been vaccinated in 1993 and tested for anti-HBs prior to commencing work in 1997. The doctor was recalled to Occupational Health but did not attend and was not followed up. In total, 4948 patients potentially treated by the doctor received an explanatory letter and 3150 were tested for HBsAg. Only one was positive, and HBV sequencing showed no link to the doctor. Occasionally transmission of HBV from heath-care workers can occur in a non-EPP setting and the implications of this require examination by those setting national policy. Occupational Health Services should investigate clinical heath-care workers who do not respond to vaccination. They should ensure HBV carriers are identified and offer them appropriate advice to prevent transmission to patients.

Practical experience of high throughput real time PCR in the routine diagnostic virology setting.

The advent of PCR has transformed the utility of the virus diagnostic laboratory. In comparison to traditional gel based PCR assays, real time PCR offers increased sensitivity and specificity in a rapid format. Over the past 4 years, we have introduced a number of qualitative and quantitative real time PCR assays into our routine testing service. During this period, we have gained substantial experience relating to the development and implementation of real-time assays. Furthermore, we have developed strategies that have allowed us to increase our sample throughput while maintaining or even reducing turn around times. The issues resulting from this experience (some of it bad) are discussed in detail with the aim of informing laboratories that are only just beginning to investigate the potential of this technology.