Data Preprocessing

Preparing data collected during animal experiment and ANOVA

Libraries

Load needed libraries and packages.

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library(tidyverse)
library(here)
library(pals)

Custom Functions

Load customized functions.

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source(here("R/utilities.R"))
source_dir(here("R"))

Read Data

Data collected during animal experiment.

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antem_df <- read_csv(here("local/bov_antemortem_2024.csv"))

# minimum date, donor inoculation
min_date <- min(antem_df$date)

# add variables 
antem_df <- antem_df %>%
  mutate(hpdi = as.numeric(difftime(date, min_date, units = "hours")), # hours post donor inoculation 
         hpe = dpe*24, # convert dpe to hpe
         exp_type = if_else(group == "donor", "inoc", "cont"), # exposed by inoculation or direct contact
         fever = if_else(temp >= 104, "fever", "no_fever"), # temp >= 104 constitutes fever
         censor_status = if_else(group == "Group 1" | animal == "BR23-24", 0, 1), # No obs symptoms (0) in these 
  )

Plots to Check Data

Donors

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plot_donors(antem_df)

Contact Groups

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plot_contact_groups(antem_df)

ANOVA

To evaluate the differences in nasal, serum, and lesion score among groups and over time, repeated measures ANOVAs were applied using the ezANOVA() function from the ez R-package. The analyses included within-subjects (days post-exposure, dpe) and between-subjects (group) factors, with nasal, serum, and lesion scores as the dependent variables. Mauchly’s test was conducted to test the assumption of sphericity. Where violations were found, Greenhouse-Geisser and Huynh-Feldt corrections were applied.

Prepare Data

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contact_groups <- antem_df %>%
  filter(group %in% c("Group 2", "Group 3", "Group 4")) %>%
  select(animal, group, nasal, serum, dpe, score) %>%
  mutate(
    score = replace_na(score, 0)) %>%
  drop_na() %>%
  mutate(
    score = replace_na(score, 0),          
    nasal = replace(nasal, nasal == 45, 0), # 45 indicates no detection
    serum = replace(serum, serum == 45, 0),
    dpe = as.factor(dpe),                   
    animal = as.factor(animal),            
    group = as.factor(group)                
  ) %>%
  as.data.frame()

Results indicate that nasal virus quantity differed significantly between the groups and across the days post-exposure, with an interaction effect suggesting that the temporal pattern of virus quantity varied among the groups.

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nasal_anova <- ez::ezANOVA(data = contact_groups, dv = .(nasal), wid = .(animal), within = .(dpe), between = .(group))
nasal_anova

$ANOVA
     Effect DFn DFd         F            p p<.05       ges
2     group   2   9  9.223642 6.619789e-03     * 0.2693092
3       dpe   6  54 33.985994 1.195097e-16     * 0.7559307
4 group:dpe  12  54  5.531737 4.748785e-06     * 0.5020495

$`Mauchly's Test for Sphericity`
     Effect           W           p p<.05
3       dpe 0.002507464 0.007421478     *
4 group:dpe 0.002507464 0.007421478     *

$`Sphericity Corrections`
     Effect       GGe        p[GG] p[GG]<.05       HFe        p[HF] p[HF]<.05
3       dpe 0.4698995 7.240063e-09         * 0.7063488 2.378893e-12         *
4 group:dpe 0.4698995 1.041644e-03         * 0.7063488 9.172502e-05         *

Results suggest that while there were no significant differences between groups, there were significant changes over time, and the pattern of these changes differed significantly among the groups.

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serum_anova <- ez::ezANOVA(data = contact_groups, dv = .(serum), wid = .(animal), within = .(dpe), between = .(group))
serum_anova

$ANOVA
     Effect DFn DFd         F            p p<.05       ges
2     group   2   9  2.789541 1.141052e-01       0.1386202
3       dpe   6  54 25.182482 5.082150e-14     * 0.6744439
4 group:dpe  12  54  7.951312 2.761527e-08     * 0.5667711

$`Mauchly's Test for Sphericity`
     Effect            W            p p<.05
3       dpe 1.265279e-05 6.228482e-08     *
4 group:dpe 1.265279e-05 6.228482e-08     *

$`Sphericity Corrections`
     Effect       GGe        p[GG] p[GG]<.05       HFe        p[HF] p[HF]<.05
3       dpe 0.2446423 7.610504e-05         * 0.2805428 2.727829e-05         *
4 group:dpe 0.2446423 2.894414e-03         * 0.2805428 1.633022e-03         *

Results indicate that lesion scores differed significantly between the groups and across the days post-exposure, with an interaction effect suggesting that the temporal pattern of lesion severity varied among the groups.

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score_anova <- ez::ezANOVA(data = contact_groups, dv = .(score), wid = .(animal), within = .(dpe), between = .(group))
score_anova

$ANOVA
     Effect DFn DFd         F            p p<.05       ges
2     group   2   9  5.860606 2.345433e-02     * 0.2550442
3       dpe   6  54 45.873199 1.823174e-19     * 0.7897890
4 group:dpe  12  54  3.038905 2.531711e-03     * 0.3323484

$`Mauchly's Test for Sphericity`
     Effect            W            p p<.05
3       dpe 6.163627e-19 5.763765e-47     *
4 group:dpe 6.163627e-19 5.763765e-47     *

$`Sphericity Corrections`
     Effect       GGe        p[GG] p[GG]<.05       HFe        p[HF] p[HF]<.05
3       dpe 0.4739793 2.829807e-10         * 0.7157848 1.639730e-14         *
4 group:dpe 0.4739793 2.364298e-02         * 0.7157848 8.310038e-03         *