Cry1 pre-induced by NO in vitro and in vivo is
important for V. cholerae blue light protection.
Since we demonstrated that cry1 is induced in the presence of NO
(Fig. 1C), we assessed the potential role of NO pre-induction on the
protection of blue light in V. cholerae . Wildtype andΔcry1 were first cultured overnight in LB with or without the
addition of NO, then diluted into artificial seawater and exposed to
blue light for 8 hours. We found that pre-incubation with NO increased
blue light survival rate approximately 10-fold in wildtype, but not inΔcry1 mutants (Fig. 3A). When Δcry1 was complemented with
a plasmid containing an IPTG-induced
Ptac -cry1 , this strain also displayed blue
light resistance regardless of NO preinduction (Fig. 3A), suggesting
that induction of cry1 enhances blue light survivability inV. cholerae .
To further explore the impact of Cry1 on the environmental survival ofV. cholerae after exiting the host, we co-colonized Δcry1and wildtype in streptomycin-treated adult mice, in which bacteria
experience host-generated oxidative and nitrosative stress (Speeset al. , 2013, Wang et al. , 2018, Stern et al. ,
2012). To mitigate host-derived RNS, we orally administrated
aminoguanidine (AG), an iNOS inhibitor that is known to reduce NO
production in mice (Cross et al. , 1994, Winter et al. ,
2013). The experimental procedure is detailed in Fig. 3B. 2 days
post-infection, fecal samples were collected from each respective group,
purified, and introduced into seawater with or without exposure to blue
light. Our findings revealed that the colonization of Δcry1 was
comparable to that of wildtype strains in both regular mice and
AG-treated mice (Fig. 3C, orange bars). However, the survival rate ofΔcry1 mutants, once outside regular mice (without AG treatment)
and exposed to blue light, was significantly reduced compared to that of
the wildtype strains (Fig. 3C, left panel, gray circles and blue
squares). Notably, no substantial competitive disadvantage
was evident for Δcry1 mutants originating from AG-treated mice
(Fig. 3C, right panel, gray circles and blue squares). Taken together,
these data suggest that during infection, host-derived RNS induces Cry1
production, and this induction appears to confer protection uponV. cholerae against the deleterious effects of blue light
subsequent to its exit from the host.