What subtle change does Scout notice in her father? What is the cause of this difference?

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"I was beginning to notice subtle changes in my father these days, that came out when he talked to with Aunt Alexandra"

This is a quote from chapter 16. Scout is talking and beginning to see the first signs of stress on her father.

Aunt Alexandra has come to town and does not think the same way Atticus does. She has no problems letting him know this and tries to get him to fire Calpurina. Atticus refuses and says that Cal is a member of the family. Atticus is also starting to feel anger at the people in the town. They are all showing prejudice against Tom, just because he is a black man. Atticus is silently thinking to himself that he just might have a chance of winning the case. Scout sees him working harder and harder. He is spending less time at home. Most of the care is now being done by Calpurnia and Aunt Alexandra. 

Scout has always seen her father with the utmost composure and the patience of a saint. He is now starting to show signs of anger and impatience. He is angry at the town for the way they think and he is angry at Aunt Alexandra for her ignorance. Atticus is a man who fights for the rights of all people. The stress of the case is taking its toll on him. This is what Scout sees in her father. She sees how tired he is and she is starting to see the worry he carries.

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