As mentioned, the central theme of the novel is definitive female courage in the midst of adversity/tribulation.
In a literary work, the setting often includes descriptions of the weather, central locations, details about social culture, and descriptions about pertinent surroundings. In the novel, Brooks uses all of these elements to highlight the courage of the novel's female characters.
In Part Two Chapter 3, Brooks describes how Edward Cooper and Jamie Frith are caught playing with dead rats. It is Edward's mother, Mary, who catches the boys and alerts Jamie's mother (Anna) to the boys' exploits.
In the same chapter, the barber-surgeon (who tends to Edward) indirectly confirms Anna's suspicions that Edward is suffering from symptoms of the Black Plague. Meanwhile, the damp weather, which provides the right conditions for a flea infestation in the village, directly confirms the presence of the Black Plague.
History tells us that the plague was caused by the bacterium Yersinia pestis, which infected the Oriental rat flea. The rat flea began its murderous work by first infecting rodents such as mice and rats. Then, when the animals perished, the infected rat fleas sought their new home in the bodies of humans. This is how the Plague came to London in 1665. Note that the story is set in the fall of 1666: this period constituted the height of the Plague's domination in London.
The novel's dark setting is further reinforced in Part 2 Chapter 4, when the people's fears lead them to viciously target the female faith healers Mem and Anys Gowdie. Grieving villagers throw the brutally tortured Mem into the flooded mine and declare that she will be designated a witch unless she floats. Of course, Mem sinks straightaway. Despite the murderous throng surrounding the hapless Mem and the dangers the watery mine posed, Anna (our heroine) courageously works to save her friend.
The only one who steps in to assist Anna is Anys Gowdie, Mem's niece. The two eventually manage to pull Mem out, and the older woman revives after Anys performs CPR on her. However, the superstitious villagers now think that Anys used witchcraft to save Mem, and they turn on her. To protect Anys, Anna wards off blows from violently angry male villagers who are intent upon lynching the beleaguered faith healer. In the end, despite her best efforts, Anna is no match for the attackers. Anys is hanged and dies from her wounds.
The dark setting definitely reinforces Anna's courage. For more about the setting, you may want to discuss how the villagers feel circumscribed by the "wide green prison" of their existence (Part 2 Chapter 6). For more examples about Anna's courage, please refer to Part 2 Chapter 7, which discusses how Anna takes Maggie Cantwell (the Bradfords' servant) in, despite the fact that she now suffers from symptoms of the Plague. Meanwhile, in Part 3 Chapter 1, Anna saves Mrs. Elizabeth Bradford's illegitimate baby, by agreeing to raise the child as her own in another town. For their part, the Bradfords hide their evil intentions from Anna. Fortunately, Mr. Mompellion warns Anna (just in time) that the couple may be planning to kill her. Essentially, Anna puts her life in danger for the sake of an innocent child.
One of the strongest symbols that highlights the central theme is the medicinal herbs utilized by Mem, Anys, and Anna. You can find more information about this at Anna's Herbal: An Education In The Healing Power of Herbs.
Wormwood and feverfew: In Part 2 Chapter 4, Anys uses these herbs to try to bring Jamie's fever down. Despite the obvious risk to their lives, both Anys and Mem practice their herbal craft to benefit and comfort their fellow villagers. Their actions reinforce the central theme of the novel.
Comfrey: In Part 2 Chapter 5, this is the herb Elinor Mompellion and Anna use on Mem's wounded face. The herb is known to greatly aid the healing of wounds and is also beneficial as a skin tonic. Both Anna and Elinor willingly place their lives at risk in order to nurse Mem during the last days of her life. Again, the actions of these women reinforce the main theme of the novel.
The herbs that the women wield for the benefit of their fellow villagers symbolize the power of women as healers. For more, please refer to the links I have provided.