48 4 ART for Antibacterial Infection
cells release the bactericidal NO. Second, although NO production in G− bacteria
is unaffected by ART, G+ bacteria among cohabitats can be influenced by ART.
Third, ART can inhibit bacterial CAT, either G+ or G−, to exert an enhanced apop-
totic role via H 2 O 2.
To induce the gastrointestinal bacterial infection, we fed mice daily with live
bacteria, either B. Licheniformis or E. coli collected from overnight bacterial cul-
tures. To estimate the degree of bacterial infection, we determined mouse serum
NO levels and counted the colony numbers on solid media plating with diluted
stool cultures. These simple measurement methods enable the convenient inves-
tigation on the effect of ART on antibiotics’ bactericidal capacity. However, the
available data of serum NO levels and bacterial colony numbers were all derived
from both endogenous (host residing) and exogenous (daily feeding) bacteria. To
distinguish the endogenous bacteria from the exogenous bacteria, we introduced
a plasmid carrying a selective marker (antibiotic resistance) into bacteria, which
could mimic antibiotic resistant bacteria.
4.3.2 Results and Analysis
4.3.2.1 Indirect and Direct Monitoring of Bacterial Infection
in Gastrointestinal Tracts of Mice Fed with Live Bacteria
Except for the production of NO from mouse gut microbiota, daily fed live bac-
teria can also activate mouse mucosa immune systems to trigger a burst of NO
into mouse gastrointestinal tracts. In consequences, only a low-level (4–6 μM)
NO was measured in the serum of control mice, whereas a high-level (22–25 μM)
NO was determined in the serum of mice after feeding with E. coli for three days.
These results implied that live bacterial feeding, even nonpathogenic, can lead to
bacterial infection, macrophage activation, and NO production. Therefore, a raised
serum NO level may represent an indirect index indicating bacterial infection.
To directly monitor the gastrointestinal bacterial infection, we established a cor-
relation of bacterial infection with colony formation upon plating a fecal dilution
of mice fed with bacteria. After cultured overnight on the plate containing AMP
(100 μg/ml), we did observe a few colonies occurring on the fecal plate prepared
from bacteria-fed mice, whereas no colony was formed on the fecal plate prepared
from control mice. The combined use of a plasmid-harboring bacterial strain with
selective cultural plates ensures only fed bacteria rather than host bacteria forming
colonies because the plasmid carries AMP resistance gene (Ampr).
4.3.2.2 In Vivo Study in Mice Fed with E. Coli
By overnight incubation, no colony occurs on the plate covering a layer of fecal
dilution prepared from E. coli-fed mice injected with 100 μg/ml AMP + 60 μg/ml