From pre-processing to advanced dynamic modeling of pupil data.

The pupil of the eye provides a rich source of information for cognitive scientists, as it can index a variety of bodily states (e.g., arousal, fatigue) and cognitive processes (e.g., attention, decision-making). As pupillometry becomes a more accessible and popular methodology, researchers have proposed a variety of techniques for analyzing pupil data. Here, we focus on time series-based, signal-to-signal approaches that enable one to relate dynamic changes in pupil size over time with dynamic changes in a stimulus time series, continuous behavioral outcome measures, or other participants' pupil traces. We first introduce pupillometry, its neural underpinnings, and the relation between pupil measurements and other oculomotor behaviors (e.g., blinks, saccades), to stress the importance of understanding what is being measured and what can be inferred from changes in pupillary activity. Next, we discuss possible pre-processing steps, and the contexts in which they may be necessary. Finally, we turn to signal-to-signal analytic techniques, including regression-based approaches, dynamic time-warping, phase clustering, detrended fluctuation analysis, and recurrence quantification analysis. Assumptions of these techniques, and examples of the scientific questions each can address, are outlined, with references to key papers and software packages. Additionally, we provide a detailed code tutorial that steps through the key examples and figures in this paper. Ultimately, we contend that the insights gained from pupillometry are constrained by the analysis techniques used, and that signal-to-signal approaches offer a means to generate novel scientific insights by taking into account understudied spectro-temporal relationships between the pupil signal and other signals of interest.

Pleasantness of nonlinear distortion in isolated triads of synthetic timbre.

Distortion of sound is an important tool to increase the variety of timbres in musical compositions, but perceived pleasantness of distortion is understudied, and studies are limited to guitar practices in rock and metal music. This study applied a more systematic approach, using synthetic timbre and creating an audio-plugin that realized nonlinear symmetric and asymmetric distortion. Participants evaluated the perceived pleasantness of isolated triads differing in distortion (undistorted, symmetric, asymmetric), tonality (minor, major), and position (low, high, wide), taking baseline differences of tonality and position into account. Perceived pleasantness decreased by distortion, and the decrease was stronger for minor than major triads and stronger for asymmetric than symmetric distortion. Position played only a minor role in the evaluations, except for stimuli in high positions. Stimulus-based analyses showed a relation between pleasantness and the variability of roughness, mean spectral centroid, and mean sound intensity. Subject-based analyses revealed a smaller decrease in pleasantness with a preference for electronic music. Importantly, some distorted triads were rated as pleasant in absolute terms: major triads with symmetric distortion in low or wide position. That is, indeed, distortion is not always categorized as unpleasant but can be perceived as pleasant.

Deep learning models for webcam eye tracking in online experiments.

Eye tracking is prevalent in scientific and commercial applications. Recent computer vision and deep learning methods enable eye tracking with off-the-shelf webcams and reduce dependence on expensive, restrictive hardware. However, such deep learning methods have not yet been applied and evaluated for remote, online psychological experiments. In this study, we tackle critical challenges faced in remote eye tracking setups and systematically evaluate appearance-based deep learning methods of gaze tracking and blink detection. From their own homes and laptops, 65 participants performed a battery of eye tracking tasks including (i) fixation, (ii) zone classification, (iii) free viewing, (iv) smooth pursuit, and (v) blink detection. Webcam recordings of the participants performing these tasks were processed offline through appearance-based models of gaze and blink detection. The task battery required different eye movements that characterized gaze and blink prediction accuracy over a comprehensive list of measures. We find the best gaze accuracy to be 2.4° and precision of 0.47°, which outperforms previous online eye tracking studies and reduces the gap between laboratory-based and online eye tracking performance. We release the experiment template, recorded data, and analysis code with the motivation to escalate affordable, accessible, and scalable eye tracking that has the potential to accelerate research in the fields of psychological science, cognitive neuroscience, user experience design, and human-computer interfaces.

Eye blinking, musical processing, and subjective states-A methods account.

Affective sciences often make use of self-reports to assess subjective states. Seeking a more implicit measure for states and emotions, our study explored spontaneous eye blinking during music listening. However, blinking is understudied in the context of research on subjective states. Therefore, a second goal was to explore different ways of analyzing blink activity recorded from infra-red eye trackers, using two additional data sets from earlier studies differing in blinking and viewing instructions. We first replicate the effect of increased blink rates during music listening in comparison with silence and show that the effect is not related to changes in self-reported valence, arousal, or to specific musical features. Interestingly, but in contrast, felt absorption reduced participants' blinking. The instruction to inhibit blinking did not change results. From a methodological perspective, we make suggestions about how to define blinks from data loss periods recorded by eye trackers and report a data-driven outlier rejection procedure and its efficiency for subject-mean analyses, as well as trial-based analyses. We ran a variety of mixed effects models that differed in how trials without blinking were treated. The main results largely converged across accounts. The broad consistency of results across different experiments, outlier treatments, and statistical models demonstrates the reliability of the reported effects. As recordings of data loss periods come for free when interested in eye movements or pupillometry, we encourage researchers to pay attention to blink activity and contribute to the further understanding of the relation between blinking, subjective states, and cognitive processing.

In touch: Cardiac and respiratory patterns synchronize during ensemble singing with physical contact.

Musical ensemble performances provide an ideal environment to gain knowledge about complex human interactions. Network structures of synchronization can reflect specific roles of individual performers on the one hand and a higher level of organization of all performers as a superordinate system on the other. This study builds on research on joint singing, using hyperscanning of respiration and heart rate variability (HRV) from eight professional singers. Singers performed polyphonic music, distributing their breathing within the same voice and singing without and with physical contact: that is touching each other's shoulder or waist. The idea of singing with touch was motivated by historical depictions of ensemble performances that showed singers touching each other. It raises the question of the potential benefit of touch for group performances. From a psycho-physiological point of view, physical contact should increase the synchronization of singing coordination. The results confirm previous findings on synchronization of respiration and HRV during choir singing and extend those findings to a non-homophonic musical repertoire while also revealing an increase in synchronization in respiration during physical contact. These effects were significant across different frequency ranges. The effect of physical contact was stronger when all singers were singing in comparison to the partial ensemble. Importantly, the synchronization could not be fully explained by the singing action (i.e., singing the same voice, or singing vs. listening) or by the standing position or touch. This finding suggests a higher level of organization of all singers, forming a superordinate system.

Multisensory integration of musical emotion perception in singing.

We investigated how visual and auditory information contributes to emotion communication during singing. Classically trained singers applied two different facial expressions (expressive/suppressed) to pieces from their song and opera repertoire. Recordings of the singers were evaluated by laypersons or experts, presented to them in three different modes: auditory, visual, and audio-visual. A manipulation check confirmed that the singers succeeded in manipulating the face while keeping the sound highly expressive. Analyses focused on whether the visual difference or the auditory concordance between the two versions determined perception of the audio-visual stimuli. When evaluating expressive intensity or emotional content a clear effect of visual dominance showed. Experts made more use of the visual cues than laypersons. Consistency measures between uni-modal and multimodal presentations did not explain the visual dominance. The evaluation of seriousness was applied as a control. The uni-modal stimuli were rated as expected, but multisensory evaluations converged without visual dominance. Our study demonstrates that long-term knowledge and task context affect multisensory integration. Even though singers' orofacial movements are dominated by sound production, their facial expressions can communicate emotions composed into the music, and observes do not rely on audio information instead. Studies such as ours are important to understand multisensory integration in applied settings.

Manual motor reaction while being absorbed into popular music.

In three experiments, we investigated the behavioral consequences of being absorbed into music on performance in a concurrent task. We tested two competing hypotheses: Based on a cognitive load account, captivation of attention by the music and state absorption might slow down reactions in the decisional task. Alternatively, music could induce spontaneous motor activity, and being absorbed in music might result in a more autonomous, flow-driven behavior with quicker motor reactions. Participants performed a simple, visual, two-alternative forced-choice task while listening to popular musical excerpts. Subsequently, they rated their subjective experience using a short questionnaire. We presented music in four tempo categories (between 80 and 140 BPM) to account for a potential effect of tempo and an interaction between tempo and absorption. In Experiment 1, absorption was related to decreased reaction times (RTs) in the visual task. This effect was small, as expected in this setting, but replicable in Experiment 2. There was no effect of the music's tempo on RTs but a tendency of mind wandering to relate to task performance. After slightly changing the study setting in Experiment 3, flow predicted decreased RTs, but absorption alone - as part of the flow construct - did not predict RTs. To sum up, we demonstrated that being absorbed in music can have the behavioral consequence of speeded manual reactions in specific task contexts, and people seem to integrate the music into an active, flow-driven and therefore enhanced performance. However, shown relations depend on task settings, and a systematic study of context is necessary to understand how induced states and their measurement contribute to the findings.

Hepatitis E virus: Efficacy of pasteurization of plasma-derived VWF/FVIII concentrate determined by pig bioassay.

BACKGROUND: Hepatitis E virus (HEV) is the leading cause of acute hepatitis throughout the world. Increasing blood component transfusion-associated HEV infections highlight the need for reliable virus inactivation procedures for plasma derivatives from pooled plasma donations. STUDY DESIGN AND METHODS: An animal infection study was conducted to evaluate the efficiency of HEV inactivation by pasteurization during the manufacturing process of the von Willebrand Factor/Factor VIII (VWF/FVIII) concentrate Haemate P/Humate-P (CSL Behring, Marburg, Germany). For this purpose, groups of pigs were inoculated with stabilized VWF/FVIII intermediate spiked with HEV-positive liver homogenate and exposed to increasing incubation times of 0, 3, 6, and 10 h at 60°C. Animals were evaluated for virus replication over 27 days and in a subsequent trial over 92 days. RESULTS: Virus replication was detected in animals up to the 6-h pasteurization group. In contrast, pasteurization for 10 h did not reveal virus detection when the observation period was 27 days. In an additional experiment using the 10-h pasteurized material, two individuals started virus excretion and seroconverted when the observation period was extended to 92 days. Based on the total infection rate (2 of 12) of the animals inoculated with the sample pasteurized for 10 h, a virus reduction factor of at least 4.7 log10 is calculated. CONCLUSION: This study demonstrates that pasteurization at 60°C for 10 h of an HEV-positive plasma derivative leads to the effective reduction of infectivity, resulting in a VWF/FVIII product with an appropriate margin of safety for HEV.

Gaze-based and attention-based rehearsal in spatial working memory.

How do we maintain information about spatial configurations in mind? Many working memory (WM) models assume that rehearsal processes are used to counteract forgetting in WM. Here, we investigated the contributions of gaze-based and attention-based rehearsal for protecting spatial representations from time-based forgetting. Participants memorized 6 locations selected from a grid of 30 scattered dots. Memory was tested after 1.5 or 4.5 s, and this interval was either blank or the grid remained onscreen (which is assumed to provide rehearsal support). In 2 experiments, we monitored eye movements during the retention phase, or asked participants to fixate the screen center. In 3 subsequent experiments, we tested spatial WM under dual-task conditions inhibiting shifts of visuospatial attention or central attention to the memoranda. Memory was better and more resistant to time-based forgetting in the grid than blank condition. Recording of fixations showed more frequent and efficient gaze-based rehearsal in the presence of the grid. Fixations toward distractor locations occurred at a similar frequency in the blank and grid conditions, and it did not predict incorrect recalls. Inhibition of eye-movements or shifts of visuospatial attention impaired memory overall, but it did not change the grid benefit nor the rate of time-based forgetting. In contrast, distracting central attention increased time-based forgetting regardless of grid presence. These results indicate that (a) the grid benefit is only partially explained by rehearsal; (b) gaze-errors (i.e., distractor fixations) do not lead to more forgetting; and (c) the maintenance of spatial representations over time depends on central processing. (PsycInfo Database Record (c) 2020 APA, all rights reserved).

On-item fixations during serial encoding do not affect spatial working memory.

Ample evidence suggests that there is overlap between the eye-movement system and spatial working memory. Such overlapping structures or capacities may result in interference on the one hand and beneficial support on the other. We investigated eye-movement control during encoding of verbal or spatial information, keeping the display the same between tasks. Saccades to to-be-encoded items were scarce during spatial encoding in comparison with verbal encoding. However, despite replicating this difference across different tasks (serial, free recall) and presentation modalities (simultaneous, sequential presentation), we found no relation between item fixations and memory performance-that is, no costs or benefits. Inducing a change from covert to overt encoding did not affect spatial memory performance as well. In contrast, regressive fixations on prior items, that were no longer on the screen, were associated with increased spatial memory performance. Regressions occurred mainly at the end of the encoding period and were targeted at the first presented item. Our results suggest a dissociation between two types of fixations that accompany serial spatial memory: On-item fixations are epiphenomenal; regressions indicate rehearsal or output preparation.

The application of eye-tracking in music research.

Though eye-tracking is typically a methodology applied in the visual research domain, recent studies suggest its relevance in the context of music research. There exists a communityof researchers interested in this kind of research from varied disciplinary backgrounds scattered across the globe. Therefore, in August 2017, an international conference was held at the Max Planck Institute for Empirical Aesthetics in Frankfurt, Germany,to bring this research community together. The conference was dedicated to the topic of music and eye-tracking, asking the question: what do eye movements, pupil dilation, and blinking activity tell us about musical processing? This special issue is constituted of top-scoring research from the conference and spans a range of music-related topics. From tracking the gaze of performers in musical trios to basic research on how eye movements are affected by background music, the contents of this special issue highlight a variety of experimental approaches and possible applications of eye-tracking in music research.

High sensitivity of domestic pigs to intravenous infection with HEV.

BACKGROUND: Hepatitis E virus (HEV) is one major cause of acute clinical hepatitis among humans throughout the world. In industrialized countries an increasing number of autochthonous HEV infections have been identified over the last years triggered by food borne as well as - to a much lower degree - by human to human transmission via blood transfusion. Pigs have been recognised as main reservoir for HEV genotype 3 (HEV-3), and zoonotic transmission to humans through undercooked/raw meat is reported repeatedly. The minimal infectious dose of HEV-3 for pigs is so far unknown. RESULTS: The minimum infectious dose of HEV-3 in a pig infection model was determined by intravenous inoculation of pigs with a dilution series of a liver homogenate of a HEV infected wild boar. Seroconversion, virus replication and shedding were determined by analysis of blood and faeces samples, collected over a maximum period of 91 days. A dose dependent incubation period was observed in faecal shedding of viruses employing a specific and sensitive PCR method. Faecal viral shedding and seroconversion was detected in animals inoculated with dilutions of up to 10- 7. This correlates with an intravenously (i.v.) administered infectious dose of only 6.5 copies in 2 ml (corresponding to 24 IU HEV RNA/ml). Furthermore the first detectable shedding of HEV RNA in faeces is clearly dose dependent. Unexpectedly one group infected with a 10- 4 dilution exhibited prolonged virus shedding for more than 60 days suggesting a persistent infection. CONCLUSION: The results indicate that pigs are highly susceptible to i.v. infection with HEV and that the swine model represents the most sensitive infectivity assay for HEV so far. Considering a minimum infectious dose of 24 IU RNA/ml our findings highlights the potential risk of HEV transmission via blood and blood products.

The rhythm of cognition - Effects of an auditory beat on oculomotor control in reading and sequential scanning.

Eye-movement behavior is inherently rhythmic. Even without cognitive input, the eyes never rest, as saccades are generated 3 to 4 times per second. Based on an embodied view of cognition, we asked whether mental processing in visual cognitive tasks is also rhythmic in nature by studying the effects of an external auditory beat (rhythmic background music) on saccade generation in exemplary cognitive tasks (reading and sequential scanning). While in applied settings background music has been demonstrated to impair reading comprehension, the effect of musical tempo on eye-movement control during reading or scanning has not been investigated so far. We implemented a tempo manipulation in four steps as well as a silent baseline condition, while participants completed a text reading or a sequential scanning task that differed from each other in terms of underlying cognitive processing requirements. The results revealed that increased tempo of the musical beat sped up fixations in text reading, while the presence (vs. absence) of the auditory stimulus generally reduced overall reading time. In contrast, sequential scanning was unaffected by the auditory pacemaker. These results were supported by additionally applying Bayesian inference statistics. Our study provides evidence against a cognitive load account (i.e., that spare resources during low-demand sequential scanning allow for enhanced processing of the external beat). Instead, the data suggest an interpretation in favor of a modulation of the oculomotor saccade timer by irrelevant background music in cases involving highly automatized oculomotor control routines (here: in text reading).

Microsaccade-rate indicates absorption by music listening.

The power of music is a literary topos, which can be attributed to intense and personally significant experiences, one of them being the state of absorption. Such phenomenal states are difficult to grasp objectively. We investigated the state of musical absorption by using eye tracking. We utilized a load related definition of state absorption: multimodal resources are committed to create a unified representation of music. Resource allocation was measured indirectly by microsaccade rate, known to indicate cognitive processing load. We showed in Exp. 1 that microsaccade rate also indicates state absorption. Hence, there is cross-modal coupling between an auditory aesthetic experience and fixational eye movements. When removing the fixational stimulus in Exp. 2, saccades are no longer generated upon visual input and the cross-modal coupling disappeared. Results are interpreted in favor of the load hypothesis of microsaccade rate and against the assumption of general slowing by state absorption.

Protection of pigs against pandemic swine origin H1N1 influenza A virus infection by hemagglutinin- or neuraminidase-expressing attenuated pseudorabies virus recombinants.

Influenza is an important respiratory disease of pigs, and may lead to novel human pathogens like the 2009 pandemic H1N1 swine-origin influenza virus (SoIV). Therefore, improved influenza vaccines for pigs are required. Recently, we demonstrated that single intranasal immunization with a hemagglutinin (HA)-expressing pseudorabies virus recombinant of vaccine strain Bartha (PrV-Ba) protected pigs from H1N1 SoIV challenge (Klingbeil et al., 2014). Now we investigated enhancement of efficacy by prime-boost vaccination and/or intramuscular administration. Furthermore, a novel PrV-Ba recombinant expressing codon-optimized N1 neuraminidase (NA) was included. In vitro replication of this virus was only slightly affected compared to parental virus. Unlike HA, the abundantly expressed NA was efficiently incorporated into PrV particles. Immunization of pigs with the two PrV recombinants, either singly or in combination, induced B cell proliferation and the expected SoIV-specific antibodies, whose titers increased substantially after boost vaccination. After immunization of animals with either PrV recombinant H1N1 SoIV challenge virus replication was significantly reduced compared to PrV-Ba vaccinated or naïve controls. Protective efficacy of HA-expressing PrV was higher than of NA-expressing PrV, and not significantly enhanced by combination. Despite higher serum antibody titers obtained after intramuscular immunization, transmission of challenge virus to naïve contact animals was only prevented after intranasal prime-boost vaccination with HA-expressing PrV-Ba.

A 2.5-kilobase deletion containing a cluster of nine microRNAs in the latency-associated-transcript locus of the pseudorabies virus affects the host response of porcine trigeminal ganglia during established latency.

UNLABELLED: The alphaherpesvirus pseudorabies virus (PrV) establishes latency primarily in neurons of trigeminal ganglia when only the transcription of the latency-associated transcript (LAT) locus is detected. Eleven microRNAs (miRNAs) cluster within the LAT, suggesting a role in establishment and/or maintenance of latency. We generated a mutant (M) PrV deleted of nine miRNA genes which displayed properties that were almost identical to those of the parental PrV wild type (WT) during propagation in vitro. Fifteen pigs were experimentally infected with either WT or M virus or were mock infected. Similar levels of virus excretion and host antibody response were observed in all infected animals. At 62 days postinfection, trigeminal ganglia were excised and profiled by deep sequencing and quantitative RT-PCR. Latency was established in all infected animals without evidence of viral reactivation, demonstrating that miRNAs are not essential for this process. Lower levels of the large latency transcript (LLT) were found in ganglia infected by M PrV than in those infected by WT PrV. All PrV miRNAs were expressed, with highest expression observed for prv-miR-LLT1, prv-miR-LLT2 (in WT ganglia), and prv-miR-LLT10 (in both WT and M ganglia). No evidence of differentially expressed porcine miRNAs was found. Fifty-four porcine genes were differentially expressed between WT, M, and control ganglia. Both viruses triggered a strong host immune response, but in M ganglia gene upregulation was prevalent. Pathway analyses indicated that several biofunctions, including those related to cell-mediated immune response and the migration of dendritic cells, were impaired in M ganglia. These findings are consistent with a function of the LAT locus in the modulation of host response for maintaining a latent state. IMPORTANCE: This study provides a thorough reference on the establishment of latency by PrV in its natural host, the pig. Our results corroborate the evidence obtained from the study of several LAT mutants of other alphaherpesviruses encoding miRNAs from their LAT regions. Neither PrV miRNA expression nor high LLT expression levels are essential to achieve latency in trigeminal ganglia. Once latency is established by PrV, the only remarkable differences are found in the pattern of host response. This indicates that, as in herpes simplex virus, LAT functions as an immune evasion locus.

From pre-attentive processes to durable representation: an ERP index of visual distraction.

Visual search and oddball paradigms were combined to investigate memory for to-be-ignored color changes in a group of 12 healthy participants. The onset of unexpected color change of an irrelevant stimulus evoked two reliable ERP effects: a component of the event-related potential (ERP), similar to the visual mismatch negativity response (vMMN), with a latency of 120-160 ms and a posterior distribution over the left hemisphere and Late Fronto-Central Negativity (LFCN) with a latency of 320-400 ms, apparent at fronto-central electrodes and some posterior sites. Color change of that irrelevant stimulus also slowed identification of a visual target, indicating distraction. The amplitude of this color-change vMMN, but not LFCN, indexed this distraction effect. That is, electrophysiological and behavioral measures were correlated. The interval between visual scenes approximated 1s (611-1629 ms), indicating that the brain's sensory memory for the color of the preceding visual scenes must persist for at least 600 ms. Therefore, in the case of the neural code for color, durable memory representations are formed in an obligatory manner.

Natural and experimental hepatitis E virus genotype 3-infection in European wild boar is transmissible to domestic pigs.

Hepatitis E virus (HEV) is the causative agent of acute hepatitis E in humans in developing countries, but sporadic and autochthonous cases do also occur in industrialised countries. In Europe, food-borne zoonotic transmission of genotype 3 (gt3) has been associated with domestic pig and wild boar. However, little is known about the course of HEV infection in European wild boar and their role in HEV transmission to domestic pigs. To investigate the transmissibility and pathogenesis of wild boar-derived HEVgt3, we inoculated four wild boar and four miniature pigs intravenously. Using quantitative real-time RT-PCR viral RNA was detected in serum, faeces and in liver, spleen and lymph nodes. The antibody response evolved after fourteen days post inoculation. Histopathological findings included mild to moderate lymphoplasmacytic hepatitis which was more prominent in wild boar than in miniature pigs. By immunohistochemical methods, viral antigens were detected mainly in Kupffer cells and liver sinusoidal endothelial cells, partially associated with hepatic lesions, but also in spleen and lymph nodes. While clinical symptoms were subtle and gross pathology was inconspicuous, increased liver enzyme levels in serum indicated hepatocellular injury. As the faecal-oral route is supposed to be the most likely transmission route, we included four contact animals to prove horizontal transmission. Interestingly, HEVgt3-infection was also detected in wild boar and miniature pigs kept in contact to intravenously inoculated wild boar. Given the high virus loads and long duration of viral shedding, wild boar has to be considered as an important HEV reservoir and transmission host in Europe.

Proof of principle: non-invasive sampling for early detection of foot-and-mouth disease virus infection in wild boar using a rope-in-a-bait sampling technique.

In this study we describe the use of a rope-in-a-bait sampling method ("pSWAB": pathogen sampling wild animals with baits) for non-invasive saliva sampling aimed at the detection of foot-and-mouth disease (FMD) viral genome in wild boar. The pSWABs are produced in the form of a standardized product by embedding a 10 cm long cotton rope in a cereal-based bait matrix. To assess the general suitability of this novel sampling technique an animal experiment was conducted to detect FMD viral genome in saliva of infected wild boar. Two juvenile animals were inoculated in the bulb of the heel with a recent wild boar FMD virus isolate and kept together with three noninoculated wild boar of the same age. Over a period of 29 days, the animals were sampled by using five pSWABs per day in addition to the collection of blood and conventional saliva swabs taken every three to four days. Viral RNA in pSWABs was identified already 24 h after infection during the incubation period and until 23 dpi. Comparison of the results of pSWAB sampling with those of conventional saliva swabs or serum samples showed satisfactory sensitivity. These experimental data demonstrate the suitability of non-invasive sampling of wild boar by using pSWABs as a sensitive, cheap and feasible sample collection technique independent of hunting activities. In addition, the use of non-invasive sampling in an appropriate surveillance strategy is discussed.

Dietary Enterococcus faecium NCIMB 10415 and zinc oxide stimulate immune reactions to trivalent influenza vaccination in pigs but do not affect virological response upon challenge infection.

Swine influenza viruses (SIV) regularly cause significant disease in pigs worldwide. Since there is no causative treatment of SIV, we tested if probiotic Enterococcus (E.) faecium NCIMB 10415 or zinc (Zn) oxide as feed supplements provide beneficial effects upon SIV infection in piglets. Seventy-two weaned piglets were fed three different diets containing either E. faecium or different levels of Zn (2500 ppm, Zn(high); 50 ppm, Zn(low)). Half of the piglets were vaccinated intramuscularly (VAC) twice with an inactivated trivalent SIV vaccine, while all piglets were then infected intranasally with H3N2 SIV. Significantly higher weekly weight gains were observed in the E. faecium group before virus infection, and piglets in Zn(high) and E. faecium groups gained weight after infection while those in the control group (Zn(low)) lost weight. Using ELISA, we found significantly higher H3N2-specific antibody levels in the E. faecium+VAC group 2 days before and at the day of challenge infection as well as at 4 and 6 days after challenge infection. Higher hemagglutination inhibition (HI) titers were also observed in the Zn(high)+VAC and E. faecium+VAC groups at 0, 1 and 4 days after infection. However, there were no significant differences in virus shedding and lung lesions between the dietary groups. Using flow cytometry analysis significantly higher activated T helper cells and cytotoxic T lymphocyte percentages in the PBMCs were detected in the Zn(high) and E. faecium groups at single time points after infection compared to the Zn(low) control group, but no prolonged effect was found. In the BAL cells no influence of dietary supplementation on immune cell percentages could be detected. Our results suggest that feeding high doses of zinc oxide and particularly E. faecium could beneficially influence humoral immune responses after vaccination and recovery from SIV infection, but not affect virus shedding and lung pathology.