Participants and study design
We studied 16 healthy participants (9 female, 7 male; mean age 26.6 years; range 20–38 years; BMI 23; range 18.8–26.4). Eleven of the participants had not received the typhoid vaccine before. Participants were recruited through local and online advertising. Inclusion criteria included: participant willing and able to give informed consent, not currently taking any medications (except the contraceptive pill) and a good sleeper determined by self-report and a sleep disorders screening interview. Exclusion criteria included: any current or previous DSM-IV Axis 1 psychiatric disorder (determined using the Standard Clinical Interview for Diagnostic and Statistical Manual for Mental Health Disorders- Fourth Edition) (Spitzer et al. 1995), diagnosis of current sleep disorder, any significant current medical condition likely to interfere with the conduct of the study or analysis of data, typhoid vaccination within the last 3 years, any vaccination within the last 6 months, history of allergies to drugs or vaccines or any component of the typhoid vaccine, congenital or acquired immune deficiency, bleeding disorder, current or recent physical illness or infection within previous 2 weeks, steroidal or non-steroidal anti-inflammatory medication within preceding 2 weeks, current substance misuse, child bearing age and not using reliable form of contraception, pregnant or breast feeding, and finally, has taken part in a psychological or medical experiment involving taking any kinds of drugs within the last 6 weeks.
All participants signed and dated the Informed Consent Form before any study specific procedures were performed. During the screening visit, in order to characterize the sample, participants completed the Eysenck Personality Questionnaire (EPQ) (Eysenck and Eysenck 1975), State Trait Anxiety Inventory (STAI) (Spielberger et al. 1970) and Beck Depression Inventory (BDI-II) (Beck et al. 1996) (Table 1). For the two study visits, participants attended the department between 1445 and 1730 hours. Following a brief interview to ensure that there had been no changes in health or medication since the screening visit that could affect eligibility, each participant was given a single 0.5-mL injection of either typhoid polysaccharide vaccine (Typhim Vi®) (Sanofi Pasteur MSD Limited, Berkshire, UK) or placebo (0.9 % sodium chloride saline solution) into the non-dominant deltoid muscle in the arm in a randomized, double blind, balanced order, cross-over design with a 7 to 14 day washout period between each study visit. Each 0.5-mL dose of typhoid vaccine contains 25 μg of the active substance, purified Vi capsular polysaccharide of Salmonella typhi (Ty2 strain). The other ingredients are phenol, sodium chloride, disodium phosphate dihydrate, sodium dihydrogen phosphate dihydrate and water. This vaccine is generally well-tolerated, but some individuals can experience temporary soreness, swelling, redness or hardness at the site of the injection. Less common side effects can include pyrexia, headache, nausea, diarrhoea, generally feeling unwell, abdominal pain and asthma with severe reactions being rare. All participants were closely monitored for 2 h post injection and a researcher was available by phone throughout the study.
Participants completed a set of subjective measures of mood state, hourly, for 4 h post injection. The Positive and Negative Affect Schedule (PANAS) is a psychometric scale, with ten descriptives for each positive and negative affect rated on a five-point Likert Scale ranging from ‘very slightly’ to ‘very much’ (Watson et al. 1988). Participants were asked to rate how they feel right now and two scores obtained; positive and negative (range 10–50). The Bond and Lader visual analogue scales (VAS) consisting of sixteen 100-mm VAS anchored by antonyms (e.g. alert–drowsy, lethargic-energetic, etc.), were combined as recommended by the authors to form three mood factors: alertness, calmness and contentedness with higher scores reflecting higher subjective feelings of stress (Bond and Lader 1974). An adverse effects profile looked at the presence and severity of common adverse effects; nausea, vomiting, dizziness, headache, loss of alertness, drowsiness and increased and decreased appetite. In addition, participants were asked to report any other symptoms that they noted.
Five milliliter blood samples were collected between February 16, 2015 and July 6, 2015 into an EDTA tube 2 h post injection, centrifuged for 10 min at 1,250×g within 30 min. Plasma was then stored at −30 °C until assay on September 23, 2015. On study completion, the plasma samples were transferred in a single batch, in unlinked anonymised form, for Human IL-6 Immunoassay using a Quantikine® HS (high sensitivity) ELISA kit (R&D Systems, Abingdon, UK). The intra-assay and inter-assay coefficients of variation are 7.4 and 7.8 %, respectively, and the limit of detection of the assay is 0.039 pg/mL.
The electrodes were fitted during each study visit and the participants then returned home to sleep as usual. On each study night, PSGs were recorded using the Embla Titanium recording system (Natus Neurology Incorporated, Middleton, WI 53562 USA). Participants retired and rose at their usual time, and this was kept constant for both study nights and preceding nights. We have previously demonstrated that the use of ambulatory home sleep recordings provides a reliable and valid method for PSG data collection and does not result in first night effects often seen in sleep laboratories (Sharpley et al. 1988, 1990). The industry standard montage, American Academy of Sleep Medicine (AASM), was used which comprises six electroencephalogram (EEG) channels (C3-M2, C4-M1, O1-M2, O2-M1, F3-M2, F4-M1), two electrooculogram (EOG) channels (E1-M2 and E2-M2), and submentalis electromyogram (EMG) channels using three electrodes (Iber et al. 2007). PSGs were staged in 30-s epochs using the Embla RemLogic sleep diagnostic software (Natus Neurology Incorporated, Middleton, WI, USA). RemLogic adheres to the AASM scoring criteria (Iber et al. 2007). AASM criteria have reclassified sleep stages and slow wave sleep (SWS) is now known as N3. Additionally, the PSGs were edited by one of the authors, an experienced sleep physiologist who was blind to treatment status (ALS). Participants were instructed not to consume alcohol, high-fat-content meals and to avoid intensive exercise for 24 h prior to each study visit. Normal caffeine intake was maintained to ensure caffeine withdrawal was not an issue. The following morning, participants removed the electrodes themselves and, within 20 min of waking, participants completed the 10-item Leeds Sleep Evaluation Questionnaire (LSEQ) (Parrott and Hindmarch, 1980). This is a standardized VAS looking at four aspects of sleep: Getting to Sleep (GTS), Quality of Sleep (QOS), Awake following Sleep (AFS) and Behaviour following Awakening (BFW) following a medication compared to the participants’ usual sleep. In addition, a final Adverse Effects profile was completed and participants were also asked to guess, with degree of certainty, whether they had received typhoid vaccine or placebo.
The first author vouches for the accuracy and completeness of the data and for the fidelity of the study to the protocol. Concealment of randomization was maintained throughout the entire study for all participants and for all researchers involved in data acquisition and analysis and the code broken only after all data had been analysed. The randomization schedule was drawn up by a member of the group not otherwise involved in the study to maintain allocation concealment.
Sixteen participants were recruited for this study. Based on previous PSG studies, this has a statistical power of 0.8 to detect a 5 % change in % sleep efficiency (SE) with <0.05 threshold. Statistical analyses were performed in SPSS version 22 (www.ibm.com/software/analytics/spss). Differences between pairs of sleep nights, LSEQ and IL-6 levels were assessed using Dependent-means (paired-samples) t test (two tailed) and effect sizes (r) calculated. Individual mood and side effect questionnaire data were analysed using a repeated measures ANOVA with two within subject factors, ‘treatment’ (placebo versus typhoid vaccine) and ‘time’ (time since treatment administration).