Lab Overview

Time: ~30 minutes

Goal: Practice one-way ANOVA analysis from start to finish using real public health data

Learning Objectives:

  • Understand when and why to use ANOVA instead of multiple t-tests
  • Set up hypotheses for ANOVA
  • Conduct and interpret the F-test
  • Perform post-hoc tests when appropriate
  • Check ANOVA assumptions
  • Calculate and interpret effect size (η²)

Structure:

  • Part A: Guided Example (follow along)
  • Part B: Your Turn (independent practice)

Submission: Upload your completed .Rmd file and published to Brightspace by the end of class.


PART A: GUIDED EXAMPLE

Example: Blood Pressure and BMI Categories

Research Question: Is there a difference in mean systolic blood pressure (SBP) across three BMI categories (Normal weight, Overweight, Obese)?

Why ANOVA? We have one continuous outcome (SBP) and one categorical predictor with THREE groups (BMI category). Using multiple t-tests would inflate our Type I error rate.


Step 1: Setup and Data Preparation

# Load necessary libraries
library(tidyverse)   # For data manipulation and visualization
library(knitr)       # For nice tables
library(car)         # For Levene's test
library(NHANES)      # NHANES dataset

# Load the NHANES data
data(NHANES)

Create analysis dataset:

# Set seed for reproducibility
set.seed(553)

# Create BMI categories and prepare data
bp_bmi_data <- NHANES %>%
  filter(Age >= 18 & Age <= 65) %>%  # Adults 18-65
  filter(!is.na(BPSysAve) & !is.na(BMI)) %>%
  mutate(
    bmi_category = case_when(
      BMI < 25 ~ "Normal",
      BMI >= 25 & BMI < 30 ~ "Overweight",
      BMI >= 30 ~ "Obese",
      TRUE ~ NA_character_
    ),
    bmi_category = factor(bmi_category, 
                         levels = c("Normal", "Overweight", "Obese"))
  ) %>%
  filter(!is.na(bmi_category)) %>%
  select(ID, Age, Gender, BPSysAve, BMI, bmi_category)

# Display first few rows
head(bp_bmi_data) %>% 
  kable(caption = "Blood Pressure and BMI Dataset (first 6 rows)")
Blood Pressure and BMI Dataset (first 6 rows)
ID Age Gender BPSysAve BMI bmi_category
51624 34 male 113 32.22 Obese
51624 34 male 113 32.22 Obese
51624 34 male 113 32.22 Obese
51630 49 female 112 30.57 Obese
51647 45 female 118 27.24 Overweight
51647 45 female 118 27.24 Overweight
# Check sample sizes
table(bp_bmi_data$bmi_category)
## 
##     Normal Overweight      Obese 
##       1939       1937       2150

Interpretation: We have 6026 adults with complete BP and BMI data across three BMI categories.


Step 2: Descriptive Statistics

# Calculate summary statistics by BMI category
summary_stats <- bp_bmi_data %>%
  group_by(bmi_category) %>%
  summarise(
    n = n(),
    Mean = mean(BPSysAve),
    SD = sd(BPSysAve),
    Median = median(BPSysAve),
    Min = min(BPSysAve),
    Max = max(BPSysAve)
  )

summary_stats %>% 
  kable(digits = 2, 
        caption = "Descriptive Statistics: Systolic BP by BMI Category")
Descriptive Statistics: Systolic BP by BMI Category
bmi_category n Mean SD Median Min Max
Normal 1939 114.23 15.01 113 78 221
Overweight 1937 118.74 13.86 117 83 186
Obese 2150 121.62 15.27 120 82 226

Observation: The mean SBP appears to increase from Normal (114.2) to Overweight (118.7) to Obese (121.6).

But is this difference statistically significant?


Step 3: Visualize the Data

# Create boxplots with individual points
ggplot(bp_bmi_data, 
  aes(x = bmi_category, y = BPSysAve, fill = bmi_category)) +
  geom_boxplot(alpha = 0.7, outlier.shape = NA) +
  geom_jitter(width = 0.2, alpha = 0.1, size = 0.5) +
  scale_fill_brewer(palette = "Set2") +
  labs(
    title = "Systolic Blood Pressure by BMI Category",
    subtitle = "NHANES Data, Adults aged 18-65",
    x = "BMI Category",
    y = "Systolic Blood Pressure (mmHg)",
    fill = "BMI Category"
  ) +
  theme_minimal(base_size = 12) +
  theme(legend.position = "none")

What the plot tells us:

  • There appears to be a trend: higher BMI categories have higher median SBP
  • The boxes overlap, but the obese group appears shifted upward
  • Variability (box heights) looks similar across groups

Step 4: Set Up Hypotheses

Null Hypothesis (H₀): μ_Normal = μ_Overweight = μ_Obese
(All three population means are equal)

Alternative Hypothesis (H₁): At least one population mean differs from the others

Significance level: α = 0.05


Step 5: Fit the ANOVA Model

# Fit the one-way ANOVA model
anova_model <- aov(BPSysAve ~ bmi_category, data = bp_bmi_data)

# Display the ANOVA table
summary(anova_model)
##                Df  Sum Sq Mean Sq F value Pr(>F)    
## bmi_category    2   56212   28106   129.2 <2e-16 ***
## Residuals    6023 1309859     217                   
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

Interpretation:

  • F-statistic: 129.24
  • Degrees of freedom: df₁ = 2 (k-1 groups), df₂ = 6023 (n-k)
  • p-value: < 2e-16 (very small)
  • Decision: Since p < 0.05, we reject H₀
  • Conclusion: There is statistically significant evidence that mean systolic BP differs across at least two BMI categories.

Step 6: Post-Hoc Tests (Tukey HSD)

Why do we need this? The F-test tells us that groups differ, but not which groups differ. Tukey’s Honest Significant Difference controls the family-wise error rate for multiple pairwise comparisons.

# Conduct Tukey HSD test
tukey_results <- TukeyHSD(anova_model)
print(tukey_results)
##   Tukey multiple comparisons of means
##     95% family-wise confidence level
## 
## Fit: aov(formula = BPSysAve ~ bmi_category, data = bp_bmi_data)
## 
## $bmi_category
##                       diff      lwr      upr p adj
## Overweight-Normal 4.507724 3.397134 5.618314     0
## Obese-Normal      7.391744 6.309024 8.474464     0
## Obese-Overweight  2.884019 1.801006 3.967033     0
# Visualize the confidence intervals
plot(tukey_results, las = 0)

Interpretation:

Comparison Mean Diff 95% CI p-value Significant?
Overweight - Normal 4.51 [3.4, 5.62] 1.98e-13 Yes
Obese - Normal 7.39 [6.31, 8.47] < 0.001 Yes
Obese - Overweight 2.88 [1.8, 3.97] 1.38e-09 Yes

Conclusion: All three pairwise comparisons are statistically significant. Obese adults have higher SBP than overweight adults, who in turn have higher SBP than normal-weight adults.


Step 7: Calculate Effect Size

# Extract sum of squares from ANOVA table
anova_summary <- summary(anova_model)[[1]]

ss_treatment <- anova_summary$`Sum Sq`[1]
ss_total <- sum(anova_summary$`Sum Sq`)

# Calculate eta-squared
eta_squared <- ss_treatment / ss_total

cat("Eta-squared (η²):", round(eta_squared, 4), "\n")
## Eta-squared (η²): 0.0411
cat("Percentage of variance explained:", round(eta_squared * 100, 2), "%")
## Percentage of variance explained: 4.11 %

Interpretation: BMI category explains 4.11% of the variance in systolic BP.

  • Effect size guidelines: Small (0.01), Medium (0.06), Large (0.14)
  • Our effect: Small

While statistically significant, the practical effect is modest—BMI category alone doesn’t explain most of the variation in blood pressure.


Step 8: Check Assumptions

ANOVA Assumptions:

  1. Independence: Observations are independent (assumed based on study design)
  2. Normality: Residuals are approximately normally distributed
  3. Homogeneity of variance: Equal variances across groups
# Create diagnostic plots
par(mfrow = c(2, 2))
plot(anova_model)

par(mfrow = c(1, 1))

Diagnostic Plot Interpretation:

  1. Residuals vs Fitted: Points show random scatter around zero with no clear pattern → Good!
  2. Q-Q Plot: Points follow the diagonal line reasonably well → Normality assumption is reasonable
  3. Scale-Location: Red line is relatively flat → Equal variance assumption is reasonable
  4. Residuals vs Leverage: No points beyond Cook’s distance lines → No highly influential outliers
# Levene's test for homogeneity of variance
levene_test <- leveneTest(BPSysAve ~ bmi_category, data = bp_bmi_data)
print(levene_test)
## Levene's Test for Homogeneity of Variance (center = median)
##         Df F value  Pr(>F)  
## group    2  2.7615 0.06328 .
##       6023                  
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

Levene’s Test Interpretation:

  • p-value: 0.0633
  • If p < 0.05, we would reject equal variances
  • Here: Equal variance assumption is met

Overall Assessment: With n > 2000, ANOVA is robust to minor violations. Our assumptions are reasonably satisfied.


Step 9: Report Results

Example Results Section:

We conducted a one-way ANOVA to examine whether mean systolic blood pressure (SBP) differs across BMI categories (Normal, Overweight, Obese) among 6,026 adults aged 18-65 from NHANES. Descriptive statistics showed mean SBP of 114.2 mmHg (SD = 15) for normal weight, 118.7 mmHg (SD = 13.9) for overweight, and 121.6 mmHg (SD = 15.3) for obese individuals.

The ANOVA revealed a statistically significant difference in mean SBP across BMI categories, F(2, 6023) = 129.24, p < 0.001. Tukey’s HSD post-hoc tests indicated that all pairwise comparisons were significant (p < 0.05): obese adults had on average 7.4 mmHg higher SBP than normal-weight adults, and 2.9 mmHg higher than overweight adults.

The effect size (η² = 0.041) indicates that BMI category explains 4.1% of the variance in systolic blood pressure, representing a small practical effect. These findings support the well-established relationship between higher BMI and elevated blood pressure, though other factors account for most of the variation in SBP.


PART B: YOUR TURN - INDEPENDENT PRACTICE

Practice Problem: Physical Activity and Depression

Research Question: Is there a difference in the number of days with poor mental health across three physical activity levels (None, Moderate, Vigorous)?

Your Task: Complete the same 9-step analysis workflow you just practiced, but now on a different outcome and predictor.


Step 1: Data Preparation

# Prepare the dataset
set.seed(553)

mental_health_data <- NHANES %>%
  filter(Age >= 18) %>%
  filter(!is.na(DaysMentHlthBad) & !is.na(PhysActive)) %>%
  mutate(
    activity_level = case_when(
      PhysActive == "No" ~ "None",
      PhysActive == "Yes" & !is.na(PhysActiveDays) & PhysActiveDays < 3 ~ "Moderate",
      PhysActive == "Yes" & !is.na(PhysActiveDays) & PhysActiveDays >= 3 ~ "Vigorous",
      TRUE ~ NA_character_
    ),
    activity_level = factor(activity_level, 
                           levels = c("None", "Moderate", "Vigorous"))
  ) %>%
  filter(!is.na(activity_level)) %>%
  select(ID, Age, Gender, DaysMentHlthBad, PhysActive, activity_level)

# YOUR TURN: Display the first 6 rows and check sample sizes
# First 6 rows:
head(mental_health_data)
## # A tibble: 6 × 6
##      ID   Age Gender DaysMentHlthBad PhysActive activity_level
##   <int> <int> <fct>            <int> <fct>      <fct>         
## 1 51624    34 male                15 No         None          
## 2 51624    34 male                15 No         None          
## 3 51624    34 male                15 No         None          
## 4 51630    49 female              10 No         None          
## 5 51647    45 female               3 Yes        Vigorous      
## 6 51647    45 female               3 Yes        Vigorous
#Check sample sizes
summary(mental_health_data$activity_level)
##     None Moderate Vigorous 
##     3139      768     1850

YOUR TURN - Answer these questions:

  • How many people are in each physical activity group?
    • None: 3139
    • Moderate: 768
    • Vigorous: 1850

Step 2: Descriptive Statistics

# YOUR TURN: Calculate summary statistics by activity level
# Hint: Follow the same structure as the guided example
# Variables to summarize: n, Mean, SD, Median, Min, Max
activity_summary_total <- mental_health_data %>%
  group_by(activity_level) %>%
    summarize(
      N = n(),
      Mean = mean(DaysMentHlthBad),
      SD = sd(DaysMentHlthBad),
      Median = median(DaysMentHlthBad),
      Min = min(DaysMentHlthBad),
      Max = max(DaysMentHlthBad)
        )


print(activity_summary_total)
## # A tibble: 3 × 7
##   activity_level     N  Mean    SD Median   Min   Max
##   <fct>          <int> <dbl> <dbl>  <dbl> <int> <int>
## 1 None            3139  5.08  9.01      0     0    30
## 2 Moderate         768  3.81  6.87      0     0    30
## 3 Vigorous        1850  3.54  7.17      0     0    30

YOUR TURN - Interpret:

  • Which group has the highest mean number of bad mental health days? Activity Level: None
  • Which group has the lowest? Activity level: Vigorous

Step 3: Visualization

# YOUR TURN: Create boxplots comparing DaysMentHlthBad across activity levels
# Hint: Use the same ggplot code structure as the example
# Change variable names and labels appropriately

ggplot(mental_health_data, 
  aes(x = activity_level, y = DaysMentHlthBad, fill = activity_level)) +
  geom_boxplot(alpha = 0.5, outlier.shape = NA,) +
  geom_jitter(width = 0.2, alpha = 0.1, size = 0.5) +
 # scale_fill_brewer(palette = "Set2") +
  labs(
    title = "Days of Reported Poor Mental Health by Activity Level",
    subtitle = "NHANES Data, Adults aged 18-65",
    x = "Activity Level",
    y = "Days Reporting Bad Mental Health",
    fill = "Activity Level"
  ) +
  theme_classic(base_size = 12) +
  theme(legend.position = "none")

YOUR TURN - Describe what you see:
Do the groups appear to differ?
The groups do appear to differ, somewhat. The medians of each group by activity level remains around 0, though the variances differ somewhat between the three groups.
Are the variances similar across groups?
The variance of Days Reporting Bad Mental Health Days for people reporting vigorous activity are smaller than the cariances seen in “None” and “Moderate” activity levels, though those groups are similar.


Step 4: Set Up Hypotheses

YOUR TURN - Write the hypotheses:

Null Hypothesis (H₀): μ_None = μ_Moderate = μ_Vigorous (All three population means of # of bad mental health days are equal)

Alternative Hypothesis (H₁): At least one population mean differs from the others.

Significance Level: α = 0.05


Step 5: Fit the ANOVA Model

# YOUR TURN: Fit the ANOVA model
# Outcome: DaysMentHlthBad
# Predictor: activity_level
# Fit the one-way ANOVA model
activity_anova <- aov(DaysMentHlthBad ~ activity_level, data = mental_health_data)

# Display the ANOVA table
summary(activity_anova)
##                  Df Sum Sq Mean Sq F value   Pr(>F)    
## activity_level    2   3109  1554.6   23.17 9.52e-11 ***
## Residuals      5754 386089    67.1                     
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

YOUR TURN - Extract and interpret the results:
- F-statistic: 23.17
- Degrees of freedom: 2
- p-value: <0.001
- Decision (reject or fail to reject H₀): Reject
- Statistical conclusion in words: There is statistically significant evidence that mean number of poor health days differs across at least activity level categories.

Step 6: Post-Hoc Tests

# YOUR TURN: Conduct Tukey HSD test
# Only if your ANOVA p-value < 0.05

# Conduct Tukey HSD test
activity_tukey <- TukeyHSD(activity_anova)
print(activity_tukey)
##   Tukey multiple comparisons of means
##     95% family-wise confidence level
## 
## Fit: aov(formula = DaysMentHlthBad ~ activity_level, data = mental_health_data)
## 
## $activity_level
##                         diff       lwr        upr     p adj
## Moderate-None     -1.2725867 -2.045657 -0.4995169 0.0003386
## Vigorous-None     -1.5464873 -2.109345 -0.9836298 0.0000000
## Vigorous-Moderate -0.2739006 -1.098213  0.5504114 0.7159887
# Visualize the confidence intervals
plot(activity_tukey, las = 0)

YOUR TURN - Complete the table:

Comparison Mean Difference 95% CI Lower 95% CI Upper p-value Significant?
Moderate - None -1.2725867 -2.045657 -0.4995169 0.0003386 Yes
Vigorous - None -1.5464873 -2.109345 -0.9836298 0.0000000 Yes
Vigorous - Moderate -0.2739006 -1.098213 0.5504114 0.7159887 No

Interpretation:
Which specific groups differ significantly?
None differs significantly from vigorous and moderate, but vigorous and moderate do not differ from each other.


Step 7: Calculate Effect Size

# YOUR TURN: Calculate eta-squared
# Hint: Extract Sum Sq from the ANOVA summary
# Extract sum of squares from ANOVA table
activity_summary <- summary(activity_anova)[[1]]

ss_treatment <- activity_summary$`Sum Sq`[1]
ss_total <- sum(activity_summary$`Sum Sq`)

# Calculate eta-squared
eta_squared <- ss_treatment / ss_total

cat("Eta-squared (η²):", round(eta_squared, 4), "\n")
## Eta-squared (η²): 0.008
cat("Percentage of variance explained:", round(eta_squared * 100, 2), "%")
## Percentage of variance explained: 0.8 %
cat("Effect Size:", 
    ifelse(eta_squared < 0.06, 
            "Small", ifelse(eta_squared < 0.14, 
                            "Medium", 
                            "Large")
           )
    )
## Effect Size: Small

YOUR TURN - Interpret:

- η² = 0.008
- Percentage of variance explained: = 0.8%
- Effect size classification = Small
- What does this mean practically? While statistically significant, the practical effect is minimal— activity level alone doesn’t explain most of the variation in the number of poor mental health days.


Step 8: Check Assumptions

# YOUR TURN: Create diagnostic plots
par(mfrow = c(2, 2))
plot(activity_anova)

par(mfrow = c(1, 1))

YOUR TURN - Evaluate each plot:

  1. Residuals vs Fitted: Points show three lines representing each of the group means, but in a way that does not move the line substantially away from flat, so this is fine to use - centered around the group means.
  2. Q-Q Plot: Honestly this doesn’t follow the diagonal well at all, it skews up and then flattens out of the top. I think I can still assume CLT applies because n > 5,700k.
  3. Scale-Location: This red line is relatively flat → Equal variance assumption is reasonable
  4. **Residuals vs Leverage:* There are no points beyond Cook’s distance lines → No highly influential outliers
# YOUR TURN: Conduct Levene's test

activity_levene <- leveneTest(DaysMentHlthBad ~ activity_level, data = mental_health_data)
print(levene_test)
## Levene's Test for Homogeneity of Variance (center = median)
##         Df F value  Pr(>F)  
## group    2  2.7615 0.06328 .
##       6023                  
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

YOUR TURN - Overall assessment:
- Are assumptions reasonably met? We fail to reject the null,indicating that we can assume homogeneiity of variance.
- Do any violations threaten your conclusions? No, regular ANOVA should be fine.


Step 9: Write Up Results

YOUR TURN - Write a complete 2-3 paragraph results section:

Include: 1. Sample description and descriptive statistics 2. F-test results 3. Post-hoc comparisons (if applicable) 4. Effect size interpretation 5. Public health significance

Your Results Section:

We conducted a one-way ANOVA to examine whether mean number of days reporting poor mental health differs across levels of activity (None, Moderate, Vigorous) among 5,757 adults aged 18-65 from NHANES. Descriptive statistics showed a mean number of days reporting poor mental health of 5.1 days (SD = 9) for those with no activity, a mean number of days of 3.8 days (SD = 6.9)) for those with moderate activity, and a mean of 3.5 days (SD = 7.2) for those with vigorous activity,

The ANOVA revealed a statistically significant difference in mean days reporting poor mental health across activity categories, F(2, 5754) = 23.17, p < 0.001.

Tukey’s HSD post-hoc tests indicated that only the comparisons between those with no activity and moderate and vigorous respectively differed, while moderate and vigorous did not differ between each other.

The effect size (η² = 0.008) indicates that activity level explains 0.8% of the variance days reporting poor mental health representing a small practical effect. These findings indicate that most variance is not accounted for by activity level.


Reflection Questions

1. How does the effect size help you understand the practical vs. statistical significance?

Effect size describes how much of the variance is accounted for by this measure.

2. Why is it important to check ANOVA assumptions? What might happen if they’re violated?

If these are violated we should be instead using modifications of the ANOVA that account for the specific violations.

3. In public health practice, when might you choose to use ANOVA?

Use anova if comparing means across more than two groups. Any categories that aren’t binary.

4. What was the most challenging part of this lab activity?

I think learning to interpret the diagnostic plots was the most challenging because a lot of the examples online were more dramatic and intentionally different deviations, whereas actual data felt more subject to interpretation in the way that you learn over time.


Submission Checklist

Before submitting, verify you have:

To submit: Upload both your .Rmd file and the HTML output to Brightspace.


Lab completed on: February 05, 2026


GRADING RUBRIC (For TA Use)

Total Points: 15

Category Criteria Points Notes
Code Execution All code chunks run without errors 4 - Deduct 1 pt per major error
- Deduct 0.5 pt per minor warning
Completion All “YOUR TURN” sections attempted 4 - Part B Steps 1-9 completed
- All fill-in-the-blank answered
- Tukey table filled in
Interpretation Correct statistical interpretation 4 - Hypotheses correctly stated (1 pt)
- ANOVA results interpreted (1 pt)
- Post-hoc results interpreted (1 pt)
- Assumptions evaluated (1 pt)
Results Section Professional, complete write-up 3 - Includes descriptive stats (1 pt)
- Reports F-test & post-hoc (1 pt)
- Effect size & significance (1 pt)

Detailed Grading Guidelines

Code Execution (4 points):

  • 4 pts: All code runs perfectly, produces correct output
  • 3 pts: Minor issues (1-2 small errors or warnings)
  • 2 pts: Several errors but demonstrates understanding
  • 1 pt: Major errors, incomplete code
  • 0 pts: Code does not run at all

Completion (4 points):

  • 4 pts: All sections attempted thoughtfully
  • 3 pts: 1-2 sections incomplete or minimal effort
  • 2 pts: Several sections missing
  • 1 pt: Only partial completion
  • 0 pts: Little to no work completed

Interpretation (4 points):

  • 4 pts: All interpretations correct and well-explained
  • 3 pts: Minor errors in interpretation
  • 2 pts: Several interpretation errors
  • 1 pt: Significant misunderstanding of concepts
  • 0 pts: No interpretation provided

Results Section (3 points):

  • 3 pts: Publication-quality, complete results section
  • 2 pts: Good but missing some elements
  • 1 pt: Incomplete or poorly written
  • 0 pts: No results section written

Common Deductions

  • -0.5 pts: Missing sample sizes in write-up
  • -0.5 pts: Not reporting confidence intervals
  • -1 pt: Incorrect hypothesis statements
  • -1 pt: Misinterpreting p-values
  • -1 pt: Not checking assumptions
  • -0.5 pts: Poor formatting (no tables, unclear output)