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] 3.82e-12 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) %>% 
  kable(caption = "Physical Activity and Mental Health Dataset (first 6 rows)")
Physical Activity and Mental Health Dataset (first 6 rows)
ID Age Gender DaysMentHlthBad PhysActive activity_level
51624 34 male 15 No None
51624 34 male 15 No None
51624 34 male 15 No None
51630 49 female 10 No None
51647 45 female 3 Yes Vigorous
51647 45 female 3 Yes Vigorous
#check sample sizes
cat("sample size:", nrow(mental_health_data), "adults aged 18+")
## sample size: 5757 adults aged 18+

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
summary_stats <- mental_health_data %>%
group_by(activity_level) %>%
  summarise(
    n=n(),
    mean= mean(DaysMentHlthBad, na.rm= TRUE),
    SD= sd(DaysMentHlthBad, na.rm= TRUE),
    Median= median(DaysMentHlthBad, na.rm=TRUE),
    min= min(DaysMentHlthBad, na.rm= TRUE),
    max= max(DaysMentHlthBad, na.rm= TRUE),
  )
  
summary_stats %>%
  kable( digits= 3,
         caption= "Descriptive statistics for Bad Mental Health Days by physical activity level")
Descriptive statistics for Bad Mental Health Days by physical activity level
activity_level n mean SD Median min max
None 3139 5.084 9.010 0 0 30
Moderate 768 3.811 6.873 0 0 30
Vigorous 1850 3.537 7.171 0 0 30

YOUR TURN - Interpret:

  • Which group has the highest mean number of bad mental health days? The group with no physical activity (activity level=none) has the highest mean number of bad mental health days, mean= 5.08
  • Which group has the lowest? The group with a vigorous activity level has the lowest mean number of bad mental health days, mean= 3.54 —

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 = 0.7) +
  geom_jitter(width = 0.2, alpha = 0.3, size = 0.5) +
  scale_fill_brewer(palette = "Set2") +
  labs(
    title = "Bad Mental Health Days by Physical Activity Level",
    subtitle = "NHANES 2017-2018, Adults aged 18+",
    x = "Physical Activity Level",
    y = "Bad Mental Health Days",
    fill = "Activity Level"
  ) +
  theme_minimal(base_size = 12) +
  theme(legend.position = "none")

YOUR TURN - Describe what you see:

  • Do the groups appear to differ?

Yes the groups appear to differ from each other. The graph shows that the vigorous physical activity group had, on average, a lower amount of bad mental health days. There is a lot of overlap between all three groups but the IQR for vigorous is smaller indicating that more individuals in this group reported very few/zero bad mental health days. The none group had more concentrated data points at the higher numbers of bad mental health days (20-30 days) while the vigourous group had some scattered at this area but most data is concentrated at zero.

  • Are the variances similar across groups?

The variances are similar across groups but not the same. The none and moderate groups look to have greater variability with a wider spread and more higher values. The vigorous group is more tightly concentrated towards zero even though there are higher values present which indicates the variance is lower than the other two groups.

Step 4: Set Up Hypotheses

YOUR TURN - Write the hypotheses:

Null Hypothesis (H₀): μ_None = μ_Moderate = μ_Vigorous
(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

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

# Display the ANOVA table
summary(anova_model)
##                  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: df1= 2, df2= 5754
  • p-value: 9.52e-11 (very small)
  • Decision (reject or fail to reject H₀): since p < 0.05 we reject H0
  • Statistical conclusion in words: There is statistically significant evidence that mean systolic BP differs across at least two BMI categories

Step 6: Post-Hoc Tests

# YOUR TURN: Conduct Tukey HSD test
# Only if your ANOVA p-value < 0.05
tukey_results <- TukeyHSD(anova_model)
print(tukey_results)
##   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(tukey_results, las = 0)

YOUR TURN - Complete the table:

Comparison Mean Difference 95% CI Lower 95% CI Upper p-value Significant?
Moderate - None -1.27 -2.05 -0.50 0.0003 Yes
Vigorous - None -1.55 -2.11 -0.098 0.000 Yes
Vigorous - Moderate -0.27 -1.10 0.55 0.716 No

Interpretation:

Which specific groups differ significantly?

The moderate and vigorous physical activity groups differed significantly from the none group, meaning that the two groups both had significantly fewer bad mental health days in comparison to those who had no physical activity. There was no significant difference between the vigorous and moderate groups.

Step 7: Calculate Effect Size

# YOUR TURN: Calculate eta-squared
# Hint: Extract Sum Sq from the ANOVA summary

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): 0.008
cat("Percentage of variance explained:", round(eta_squared * 100, 2), "%")
## Percentage of variance explained: 0.8 %

YOUR TURN - Interpret:

  • η² = 0.008
  • Percentage of variance explained: 0.8%
  • Effect size classification (small/medium/large): small
  • What does this mean practically? While statistically significant, the small effect size suggests that there are other factors also affecting bad mental health days.

Step 8: Check Assumptions

# YOUR TURN: Create diagnostic plots

par(mfrow = c(2, 2))
plot(anova_model)

par(mfrow = c(1, 1))

**YOUR TURN - Evaluate each plot:**

1. **Residuals vs Fitted:** The red line is slightly diagonal and slanted away from zero which is not a good sign

2. **Q-Q Plot:** The points do not follow the dotted line well, they curve upward far away from the line. Normality assumption is not reasonable 

3. **Scale-Location:** The line is not flat and slightly diagonal. Equal variance assumption is not reasonable

4. **Residuals vs Leverage:** No points cross Cook's distance lines indicating no highly influential outliers


``` r
# YOUR TURN: Conduct Levene's test
levene_test <- 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  23.168 9.517e-11 ***
##       5754                      
## ---
## Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

YOUR TURN - Overall assessment:

  • Are assumptions reasonably met? We reject equal variances since p < 0.05 so no our assumptions are not reasonably met
  • Do any violations threaten your conclusions? It is unlikely that any violations will affect the validity of the conclusions

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:

I conducted a one-way ANOVA to test whether bad mental health days differed across three physical activity levels: none, moderate, and vigorous. There were differences in mean bad mental health days across the activity groups. The no physical activity group (none) was 5.1 while the moderate groups mean was 3.8 and vigorous was 3.1.

The F-statistic was F(2, 5754)= 23.17, this means that the between-group variation is 23 times larger than the within-group variation. The p-value is less than .0001 which indicates that the difference between groups is unlikely to have happened by chance if all groups had the same mean.

The post-hoc tukey HSD tests revealed that there was no statistically significant difference between the moderate and vigorous groups because p=0.716 which suggests that the benefits of vigorous activity is similar to that of moderate physical activity. There was a difference between moderate and none and vigorous and none.

While statistically significant, the effect size is very small, 0.008, which indicates that physical activity only explains .8% of the variance in bad mental health days. It is likely that other outside factors affect bad mental health days more than physical activity.

Reflection Questions

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

Effect size tells us how much the factor we are looking at effects the variance in the problem we are examining, in this case how much physical activity effects the amount of bad mental health days. This helps us understand the practical significance because even though statistical significance can occur, the effect size helps determine if the differences we see across different activity levels is meaningful in real world contexts.

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

Checking ANOVA assumptions is imprtant because it helps ensure that the conclusions being drawn are reliable. If they are violated then it can be assumed that p-values are wrong and error may be occuring.

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

Anova may be chosen when looking at mental health outcomes across dietary patterns or sleep patterns. These patterns would have three or more groups to be evaluated for ANOVA to be chosen, such as varying amounts of calorie intake per day.

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

During this lab the most challenging part was checking assumptions and levenes test, while it was easy to run the code I struggled a bit to understand the meaning of the output/graphs to properly relate them to the research question.


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)