Time trends in herpesvirus seroepidemiology among Swedish adults.

BACKGROUND: Human herpesviruses are widespread among the human population. The infections often occur unnoticed, but severe disease as well as long-term sequelae are part of the symptom spectrum. The prevalence varies among subpopulations and with time. The aim of this study was to describe the seroprevalence of Immunoglobulin G against Herpes simplex 1, Herpes simplex 2, Epstein-Barr virus and Cytomegalovirus in the adult Swedish population over a time period of several decades. METHODS: Serum samples (n = 892) from biobanks, originating from 30-year-old women, 50-year-old men and 50-year-old women sampled between 1975 and 2018, were analyzed for presence of anti-herpesvirus antibodies. Linear regression analysis was used to test for a correlation between birth year and seroprevalence. Multiple linear regression analysis was used to differentiate between other factors such as age and gender. RESULTS: Birth year correlated negatively with the prevalence of immunoglobulin G against Herpes simplex 1 and Epstein-Barr virus (p = 0.004 and 0.033), and positively with Immunoglobulin G against Cytomegalovirus (p = 0.039). When participant categories were analyzed separately, birth year correlated negatively with the prevalence of Immunoglobulin G against Herpes simplex 1 and Herpes simplex 2 (p = 0.032 and 0.028) in 30-year-old women, and with the prevalence of Immunoglobulin G against Cytomegalovirus in 50-year-old men (p = 0.011). CONCLUSIONS: The prevalence of Immunoglobulin G against Herpes simplex 1, Herpes simplex 2 and Epstein-Barr virus decreases in later birth cohorts. This indicates a trend of declining risk of getting infected with these viruses as a child and adolescent.

Herpes Simplex Viral Infection Doubles the Risk of Dementia in a Contemporary Cohort of Older Adults: A Prospective Study.

Background: Evidence indicates that herpes simplex virus (HSV) participates in the pathogenesis of Alzheimer's disease (AD). Objective: We investigated AD and dementia risks according to the presence of herpesvirus antibodies in relation to anti-herpesvirus treatment and potential APOE ɛ4 carriership interaction. Methods: This study was conducted with 1002 dementia-free 70-year-olds living in Sweden in 2001-2005 who were followed for 15 years. Serum samples were analyzed to detect anti-HSV and anti-HSV-1 immunoglobulin (Ig) G, anti-cytomegalovirus (CMV) IgG, anti-HSV IgM, and anti-HSV and anti-CMV IgG levels. Diagnoses and drug prescriptions were collected from medical records. Cox proportional-hazards regression models were applied. Results: Cumulative AD and all-cause dementia incidences were 4% and 7%, respectively. Eighty-two percent of participants were anti-HSV IgG carriers, of whom 6% received anti-herpesvirus treatment. Anti-HSV IgG was associated with a more than doubled dementia risk (fully adjusted hazard ratio = 2.26, p = 0.031). No significant association was found with AD, but the hazard ratio was of the same magnitude as for dementia. Anti-HSV IgM and anti-CMV IgG prevalence, anti-herpesvirus treatment, and anti-HSV and -CMV IgG levels were not associated with AD or dementia, nor were interactions between anti-HSV IgG and APOE ɛ4 or anti-CMV IgG. Similar results were obtained for HSV-1. Conclusions: HSV (but not CMV) infection may be indicative of doubled dementia risk. The low AD incidence in this cohort may have impaired the statistical power to detect associations with AD.

Longitudinal Effects of Herpesviruses on Multiple Cognitive Outcomes in Healthy Elderly Adults.

BACKGROUND: Herpesviruses have been proposed to be involved in Alzheimer's disease development as potentially modifiable pathology triggers. OBJECTIVE: To investigate associations of serum antibodies for herpes simplex virus (HSV)-1 and cytomegalovirus (CMV) and anti-herpesvirus treatment with cognitive outcomes in relation to interactions with APOE ɛ4. METHODS: The study included 849 participants in the population-based Prospective Investigation of the Vasculature in Uppsala Seniors study. Cognitive performance at the ages of 75 and 80 years was assessed using the Mini-Mental State Examination (MMSE), trail-making test (TMT) A and B, and 7-minute screening test (7MS). RESULTS: Anti- HSV-1 IgG positivity was associated cross-sectionally with worse performance on the MMSE, TMT-A, TMT-B, 7MS, enhanced free recall, and verbal fluency tests (p = 0.016, p = 0.016, p < 0.001, p = 0.001, p = 0.033, and p < 0.001, respectively), but not orientation or clock drawing. Cognitive scores did not decline over time and longitudinal changes did not differ according to HSV-1 positivity. Anti- CMV IgG positivity was not associated cross-sectionally with cognition, but TMT-B scores declined more in anti- CMV IgG carriers. Anti- HSV-1 IgG interacted with APOE ɛ4 in association with worse TMT-A and better enhanced cued recall. Anti- HSV IgM interacted with APOE ɛ4 and anti-herpesvirus treatment in association with worse TMT-A and clock drawing, respectively. CONCLUSION: These findings indicate that HSV-1 is linked to poorer cognition in cognitively healthy elderly adults, including impairments in executive function, memory, and expressive language. Cognitive performance did not decline over time, nor was longitudinal decline associated with HSV-1.

At-home sampling to meet geographical challenges for serological assessment of SARS-CoV-2 exposure in a rural region of northern Sweden, March to May 2021: a retrospective cohort study.

BackgroundThe current SARS-CoV-2 pandemic has highlighted a need for easy and safe blood sampling in combination with accurate serological methodology. Venipuncture for testing is usually performed by trained staff at healthcare centres. Long travel distances to healthcare centres in rural regions may introduce a bias of testing towards relatively large communities with closer access. Rural regions are therefore often not represented in population-based data.AimThe aim of this retrospective cohort study was to develop and implement a strategy for at-home testing in a rural region of Sweden during spring 2021, and to evaluate its role to provide equal health care for its inhabitants.MethodsWe developed a sensitive method to measure antibodies to the S-protein of SARS-CoV-2 and optimised this assay for clinical use together with a strategy of at-home capillary blood sampling.ResultsWe demonstrated that our ELISA gave comparable results after analysis of capillary blood or serum from SARS-CoV-2-experienced individuals. We demonstrated stability of the assay under conditions that reflected temperature and humidity during winter or summer. By assessment of capillary blood samples from 4,122 individuals, we could show both feasibility of the strategy and that implementation shifted the geographical spread of testing in favour of rural areas.ConclusionImplementation of at-home sampling enabled citizens living in remote rural areas access to centralised and sensitive laboratory antibody tests. The strategy for testing used here could therefore enable disease control authorities to get rapid access to information concerning immunity to infectious diseases, even across vast geographical distance.

PILRA polymorphism modifies the effect of APOE4 and GM17 on Alzheimer's disease risk.

PILRA (rs1859788 A > G) has been suggested to be a protective variant for Alzheimer's disease (AD) and is an entry co-receptor for herpes simplex virus-1. We conducted a nested case-control study of 360 1:1-matched AD subjects. Interactions between the PILRA-A allele, APOE risk variants (ε3/ε4 or ε4/ε4) and GM17 for AD risk were modelled. The associations were cross-validated using two independent whole-genome sequencing datasets. We found negative interactions between PILRA-A and GM17 (OR 0.72, 95% CI 0.52-1.00) and between PILRA-A and APOE risk variants (OR 0.56, 95% CI 0.32-0.98) in the discovery dataset. In the replication cohort, a joint effect of PILRA and PILRA × GM 17/17 was observed for the risk of developing AD (p .02). Here, we report a negative effect modification by PILRA on APOE and GM17 high-risk variants for future AD risk in two independent datasets. This highlights the complex genetics of AD.

Long-term time trends in reactivated herpes simplex infections and treatment in Sweden.

BACKGROUND: Our aim was to describe the annual prevalence of herpes simplex virus (HSV) reactivation in relation to solar ultraviolet (UV) radiation and antiviral drug use in the Swedish adult population. METHODS: The study comprised 2879 anti-HSV-1 immunoglobulin (Ig) G positive subjects from five different cohorts who had donated serum from 1988 to 2010. The sera were analyzed for anti-HSV IgM using enzyme-linked immunosorbent assay. Associations between the presence of anti-HSV IgM antibodies, the apolipoprotein E ε4 allele and the serum sampling year were assessed by logistic regression. Seasonality of anti-HSV IgM was evaluated in a UV radiation model. Data of antiviral drugs for the entire Swedish population were compiled from two different nationwide databases: the Swedish Prescribed Drug Register and the Swedish Association of the Pharmaceutical Industry. RESULTS: Cross-sectional and longitudinal analyses indicated that the prevalence of anti-HSV IgM antibodies declined between 1988 and 2010 (odds ratio [OR] = 0.912, p < .001), while the total annual use of antiviral drugs in Sweden gradually increased from 1984 to 2017. Higher UV radiation was associated with higher prevalence of anti-HSV IgM antibodies (OR = 1.071, p = .043). CONCLUSION: The declining time trend of HSV reactivation in a Swedish cohort coincides with a steady increase of antiviral drug use in the Swedish general population.

VALZ-Pilot: High-dose valacyclovir treatment in patients with early-stage Alzheimer's disease.

Introduction: Herpes simplex virus (HSV) may be involved in Alzheimer's disease (AD) pathophysiology. The antiviral valacyclovir inhibits HSV replication. Methods: This phase-II pilot trial involved valacyclovir administration (thrice daily, 500 mg week 1, 1000 mg weeks 2-4) to persons aged ≥ 65 years with early-stage AD, anti-HSV immunoglobulin G, and apolipoprotein E ε4. Intervention safety, tolerability, feasibility, and effects on Mini-Mental State Examination (MMSE) scores and cerebrospinal fluid (CSF) biomarkers were evaluated. Results: Thirty-two of 33 subjects completed the trial on full dosage. Eighteen percent experienced likely intervention-related mild, temporary adverse events. CSF acyclovir concentrations were mean 5.29 ± 2.31 µmol/L. CSF total tau and neurofilament light concentrations were unchanged; MMSE score and CSF soluble triggering receptor expressed on myeloid cells 2 concentrations increased (P = .02 and .03). Discussion: Four weeks of high-dose valacyclovir treatment was safe, tolerable, and feasible in early-stage AD. Our findings may guide future trial design.

Serological assessment of SARS-CoV-2 exposure in northern Sweden by the use of at-home sampling to meet geographical challenges in rural regions.

The current SARS-CoV-2 pandemic has highlighted a need for easy and safe blood sampling in combination with accurate serological methodology. Venipuncture is usually performed by trained staff at health care centers. Long travel distances may introduce a bias of testing towards relatively large communities with close access to health care centers. Rural regions may thus be overlooked. Here, we demonstrate a sensitive method to measure antibodies to the S-protein of SARS-CoV-2. We adapted and optimized this assay for clinical use together with capillary blood sampling to meet the geographical challenges of serosurveillance. Finally, we tested remote at-home capillary blood sampling together with centralized assessment of S-specific IgG in a rural region of northern Scandinavia that encompasses 55,185 sq kilometers. We conclude that serological assessment from capillary blood sampling gives comparable results as analysis of venous blood. Importantly, at-home sampling enabled citizens living in remote rural areas access to centralized and sensitive laboratory antibody tests.

Antiviral treatment associated with reduced risk of clinical Alzheimer's disease-A nested case-control study.

INTRODUCTION: In this nested case-control study, we investigated if antiviral treatment given prior to onset of Alzheimer's disease (AD) could influence incident AD. METHODS: From a large population-based cohort study in northern Sweden, 262 individuals that later developed AD were compared to a non-AD matched control group with respect to prescriptions of herpes antiviral treatment. All included subjects were herpes simplex virus 1 (HSV1) carriers and the matching criteria were age, sex, apolipoprotein E genotype (ε4 allele carriership), and study sample start year. RESULTS: Among those who developed AD, 6 prescriptions of antivirals were found, compared to 20 among matched controls. Adjusted for length of follow-up, a conditional logistic regression indicated a difference in the risk for AD development between groups (odds ratio for AD with an antiviral prescription 0.287, P = .018). DISCUSSION: Antiviral treatment might possibly reduce the risk for later development of HSV1-associated AD.

Plasma Amyloid-ß in Relation to Antibodies Against Herpes Simplex Virus, Cytomegalovirus, and Chlamydophila pneumoniae.

BACKGROUND: Amyloid-ß (Aß), the key constituent of Alzheimer's disease (AD) plaques, has antimicrobial properties. OBJECTIVE: To investigate the association between plasma Aß and antibodies against the AD-related pathogens herpes simplex virus (HSV), cytomegalovirus (CMV), and C. pneumoniae. METHODS: Plasma from 339 AD cases, obtained on average 9.4 years (±4.00) before diagnosis, and their matched controls were analyzed for Aß40 and Aß42 concentrations with Luminex xMAP technology and INNOBIA plasma Aß-form assays. Enzyme-linked immunosorbent assays were utilized for analyses of anti-HSV immunoglobulin (Ig) G, anti-HSV1 IgG, anti-HSV2 IgG, anti-CMV IgG, and anti-C. pneumoniae IgG. Follow-up samples were available for 150 of the cases. RESULTS: Presence and levels of anti-HSV1 IgG, anti-HSV2 IgG, anti-CMV IgG, and anti-C. pneumoniae IgG did not correlate with concentrations of Aß42 or Aß40 in cases or controls. CONCLUSION: Levels of plasma Aß were not associated with antibodies against different AD-related pathogens.

Herpesvirus infections, antiviral treatment, and the risk of dementia-a registry-based cohort study in Sweden.

INTRODUCTION: Herpesviruses, including Herpes simplex virus type 1 (HSV1) and varicella zoster-virus (VZV), have been implicated in Alzheimer's disease (AD) development. Likewise, antiviral treatment has been suggested to protect against dementia development in herpes-infected individuals. METHODS: The study enrolled 265,172 subjects aged ≥ 50 years, with diagnoses of VZV or HSV, or prescribed antiviral drugs between 31 December 2005 and 31 December 2017. Controls were matched in a 1:1 ratio by sex and birth year. RESULTS: Antiviral treatment was associated with decreased risk of dementia (adjusted hazard ratio [HR] 0.89, 95% confidence interval [CI] 0.86 to 0.92), while herpes infection without antiviral drugs increased the risk of dementia (adjusted HR 1.50, 95% CI 1.29 to 1.74). DISCUSSION: Antiviral treatment was associated with a reduced long-term risk of dementia among individuals with overt signs of herpes infection. This is consistent with earlier findings indicating that herpesviruses are involved in the pathogenesis of AD.

An Ig γ Marker Genotype Is a Strong Risk Factor for Alzheimer Disease, Independent of Apolipoprotein E ε4 Genotype.

Increasing evidence implicates HSV type 1 (HSV1) in the pathogenesis of late-onset Alzheimer disease (AD). HSV1 has evolved highly sophisticated strategies to evade host immunosurveillance. One strategy involves encoding a decoy Fcγ receptor (FcγR), which blocks Fc-mediated effector functions, such as Ab-dependent cellular cytotoxicity. Ig γ marker (GM) allotypes, encoded by highly polymorphic IGHG genes on chromosome 14q32, modulate this immunoevasion strategy, and thus may act as effect modifiers of the HSV1-AD association. In this nested case-control human study, 365 closely matched case-control pairs-whose blood was drawn on average 9.6 y before AD diagnosis-were typed for GM alleles by a TaqMan genotyping assay. APOE genotype and a genetic risk score based on nine additional previously known AD risk genes (ABCA7, BIN1, CD33, CLU, CR1, EPHA1, MS4A4E, NECTIN2, and PICALM) were extracted from a genome-wide association study analysis. Antiviral Abs were measured by ELISA. Conditional logistic regression models were applied. The distribution of GM 3/17 genotypes differed significantly between AD cases and controls, with higher frequency of GM 17/17 homozygotes in AD cases as compared with controls (19.8 versus 10.7%, p = 0.001). The GM 17/17 genotype was associated with a 4-fold increased risk of AD (odds ratio 4.142, p < 0.001). In conclusion, the results of this study demonstrate that Ig GM 17/17 genotype contributes to the risk of later AD development, independent of apolipoprotein ε4 genotype and other AD risk genes, and explain, at least in part, why every HSV1-infected person is not equally likely to develop HSV1-associated AD.

Maternal blood folate status during early pregnancy and occurrence of autism spectrum disorder in offspring: a study of 62 serum biomarkers.

BACKGROUND: Autism spectrum disorder (ASD) evolves from an interplay between genetic and environmental factors during prenatal development. Since identifying maternal biomarkers associated with ASD risk in offspring during early pregnancy might result in new strategies for intervention, we investigated maternal metabolic biomarkers in relation to occurrence of ASD in offspring using both univariate logistic regression and multivariate network analysis. METHODS: Serum samples from 100 women with an offspring diagnosed with ASD and 100 matched control women with typically developing offspring were collected at week 14 of pregnancy. Concentrations of 62 metabolic biomarkers were determined, including amino acids, vitamins (A, B, D, E, and K), and biomarkers related to folate (vitamin B9) metabolism, lifestyle factors, as well as C-reactive protein (CRP), the kynurenine-tryptophan ratio (KTR), and neopterin as markers of inflammation and immune activation. RESULTS: We found weak evidence for a positive association between higher maternal serum concentrations of folate and increased occurrence of ASD (OR per 1 SD increase: 1.70, 95% CI 1.22-2.37, FDR adjusted P = 0.07). Multivariate network analysis confirmed expected internal biochemical relations between the biomarkers. Neither inflammation markers nor vitamin D3 levels, all hypothesized to be involved in ASD etiology, displayed associations with ASD occurrence in the offspring. CONCLUSIONS: Our findings suggest that high maternal serum folate status during early pregnancy may be associated with the occurrence of ASD in offspring. No inference about physiological mechanisms behind this observation can be made at the present time because blood folate levels may have complex relations with nutritional intake, the cellular folate status and status of other B-vitamins. Therefore, further investigations, which may clarify the potential role and mechanisms of maternal blood folate status in ASD risk and the interplay with other potential risk factors, in larger materials are warranted.

Chlamydia trachomatis and Anti-MUC1 Serology and Subsequent Risk of High-Grade Serous Ovarian Cancer: A Population-Based Case-Control Study in Northern Sweden.

BACKGROUND: Chlamydia trachomatis salpingitis causes inflammatory damage to the fallopian tube and could potentially cause initiation and progression of high-grade serous ovarian cancer (HGSC). Furthermore, C. trachomatis infection may stimulate mucin 1 (MUC1) protein production, possibly affecting anti-MUC1 antibody levels. The aim of this study was to examine if serology indicating past infection with C. trachomatis as well as anti-MUC1 production was associated with subsequent risk of HGSC. MATERIALS AND METHODS: In a prospective nested case-control study within the Northern Sweden Health and Disease Study and the Northern Sweden Maternity Cohort, the prevalence of chlamydial and anti-MUC1 antibodies was analyzed in blood samples drawn more than one year before diagnosis from 92 women with HGSC and 359 matched controls. Matching factors were age, date at blood draw, and sampling cohort. Plasma C. trachomatis IgG was analyzed using commercial micro-immunofluorescence test; chlamydial Heat Shock Protein 60 IgG (cHSP60) and anti-MUC1 IgG were analyzed with ELISA technique. RESULTS: The prevalence of C. trachomatis IgG and cHSP60 IgG antibodies, as well as the level of anti-MUC1 IgG was similar in women with HGSC and controls (16.3% vs. 17.0%, P = 0.87; 27.2% vs. 28.5%, P = 0.80; median 0.24 vs. 0.25, P = 0.70). Anti-MUC1 IgG and cHSP60 IgG levels were correlated (r = 0.169; P < 0.001). CONCLUSIONS: The findings of this prospective nested case-control study did not support an association between C. trachomatis infection, as measured by chlamydial serology, or anti-MUC1 IgG antibodies, and subsequent risk of HGSC.

Urea dilution of serum for reproducible anti-HSV1 IgG avidity index.

BACKGROUND: Herpes simplex virus type 1 (HSV1), establishes life-long latency and can cause symptoms during both first-time infection and later reactivation. The aim of the present study was to describe a protocol to generate a reliable and discriminative avidity index (AI) for anti-HSV1 IgG content in human sera. METHODS: Human serum from two distinct cohorts; one a biobank collection (Betula) (n = 28), and one from a clinical diagnostics laboratory at Northern Sweden University Hospital (NUS) (n = 18), were assessed for presence of IgG antibodies against HSV1 by a commercially available ELISA-kit. Addition of urea at the incubation step reduces effective binding, and the ratio between urea treated sample and non-treated sample was used to express an avidity index (AI) for individual samples. RESULTS: AI score ranged between 43.2 and 73.4% among anti-HSV1 positive biobank sera. Clinical samples ranged between 36.3 and 74.9%. Reproducibility expressed as an intraclass correlation coefficient (ICC) was estimated at 0.948 (95% CI: 0.900-0.979) and 0.989 (95% CI 0.969-0.996) in the biobank and clinical samples, respectively. CONCLUSION: The method allows for AI scoring of anti-HSV1 IgG from individual human sera with a single measurement. The least significant change between two measurements at the p < 0.05 level was estimated at 5.4 and 3.2 points, respectively, for the two assessed cohorts.

Herpes Simplex Virus, APOEɛ4, and Cognitive Decline in Old Age: Results from the Betula Cohort Study.

BACKGROUND: Herpes simplex virus (HSV) has been suggested to play a role in Alzheimer's disease (AD) development. OBJECTIVE: The aim of the present study was to investigate the early AD-related symptom episodic memory decline in relation to HSV and carriage of allele 4 of the apolipoprotein E gene (APOEɛ4) in a large population-based cohort with a long follow-up time. METHODS: The study included 3,413 persons, with longitudinal data available for 1,293 persons with a mean follow-up time of 11.6 years. The associations between HSV carriage, APOEɛ4 carriage, and episodic memory was investigated at baseline, as well as in longitudinal analyses where individuals with and without HSV antibodies (HSV1/2 non-specific) were matched and episodic memory decline compared. RESULTS: Cross-sectional analyses revealed an age-dependent association of HSV carriage with lower episodic memory function, particularly among APOEɛ4 carriers (p = 0.008). Longitudinal analyses showed an increased risk of episodic memory decline in HSV carriers (≥65 years: p < 0.001, all ages: non-significant), and a significant interaction between HSV and APOEɛ4 for episodic memory decline (p < 0.001). CONCLUSION: In this large population-based cohort study, both cross-sectional and longitudinal results support an association between HSV carriage and declining episodic memory function, especially among APOEɛ4 carriers. The results strengthen the hypothesis that HSV is associated with AD development.

A genetic signature including apolipoprotein Eε4 potentiates the risk of herpes simplex-associated Alzheimer's disease.

INTRODUCTION: Herpes simplex virus type 1 (HSV1) in combination with genetic susceptibility has previously been implicated in Alzheimer's disease (AD) pathogenesis. METHODS: Plasma from 360 AD cases, obtained on average 9.6 years before diagnosis, and their age- and sex-matched controls, were analyzed for anti-HSV1 immunoglobulin (Ig) G with enzyme-linked immunosorbent assays (ELISAs). A POE genotype and nine other selected risk genes for AD were extracted from a genome-wide association study analysis by deCODE genetics, Reykjavik, Iceland. RESULTS: The interaction between APOEε4 heterozygosity (APOEε2/ε4 or ε3/ε4) and anti-HSV1 IgG carriage increased the risk of AD (OR 4.55, P = .02). A genetic risk score based on the nine AD risk genes also interacted with anti-HSV1 IgG for the risk of developing AD (OR 2.35, P = .01). DISCUSSION: The present findings suggest that the APOEε4 allele and other AD genetic risk factors might potentiate the risk of HSV1-associated AD.

Preparedness and response against diseases with epidemic potential in the European Union: a qualitative case study of Middle East Respiratory Syndrome (MERS) and poliomyelitis in five member states.

BACKGROUND: EU Decision 1082/2013/EU on serious cross-border health threats provides a legal basis for collaboration between EU Member States, and between international and European level institutions on preparedness, prevention, and mitigation in the event of a public health emergency. The Decision provides a context for the present study, which aims to identify good practices and lessons learned in preparedness and response to Middle East Respiratory Syndrome (MERS) (in UK, Greece, and Spain) and poliomyelitis (in Poland and Cyprus). METHODS: Based on a documentary review, followed by five week-long country visits involving a total of 61 interviews and group discussions with experts from both the health and non-health sectors, this qualitative case study has investigated six issues related to preparedness and response to MERS and poliomyelitis: national plans and overall preparedness capacity; training and exercises; risk communication; linking policy and implementation; interoperability between the health and non-health sectors; and cross-border collaboration. RESULTS: Preparedness and response plans for MERS and poliomyelitis were in place in the participating countries, with a high level of technical expertise available to implement them. Nevertheless, formal evaluation of the responses to previous public health emergencies have sometimes been limited, so lessons learned may not be reflected in updated plans, thereby risking mistakes being repeated in future. The nature and extent of inter-sectoral collaboration varied according to the sectors involved, with those sectors that have traditionally had good collaboration (e.g. animal health and food safety), as well as those that have a financial incentive for controlling infectious diseases (e.g. agriculture, tourism, and air travel) seen as most likely to have integrated public health preparedness and response plans. Although the formal protocols for inter-sectoral collaboration were not always up to date, good personal relations were reported within the relevant professional networks, which could be brought into play in the event of a public health emergency. Cross-border collaboration was greatly facilitated if the neighbouring country was a fellow EU Member State. CONCLUSIONS: Infectious disease outbreaks remain as an ongoing threat. Efforts are required to ensure that core public health capacities for the full range of preparedness and response activities are sustained.