96
observed at 50 ng/L for methyl mercury chloride, 0.34 μg/L for phenyl mercury
acetate, and 2.3 mg/L for dimethyl mercury. Willardson et al. (1998) developed
whole-cell biosensor system to sense toluene, and the results were cross-checked
with GC-MS.
4.5.3 Genetically Modified Bacterial Biosensor in Polycyclic
Aromatic Hydrocarbons
Polycyclic aromatic hydrocarbons are found ubiquitous in the environment due to
natural and anthropogenic activities. The anthropogenic activities include petrogenic and pyrogenic activities. It occurs in the environment due to incomplete combustion of fossil fuels, burning of wood and coal, and the natural activities including
volcanic eruption, forest fires, etc. Polycyclic aromatic hydrocarbons are mutagenic
and carcinogenic. Soil can be contaminated with 1 μg/kg to 300 g/kg of polycyclic
aromatic hydrocarbons. There are various methods used for the detection of polycyclic aromatic hydrocarbon, but still they have some limitations. Kuncova et al.
(2011) reported P. putida TVAS with a tod-luxCDABE is sensitive for the bioavailability of organic pollutants. The bioreporter was responsive to 23 organic pollutants, whereas tod reporter mostly identified benzene, ethylbenzene, xylene, and
toluene from wastewater and groundwater samples with 0.5–120 mg/L detection
limit. Using similar strategies, Kohlmeier et al. (2008) reported a highly sensitive
naphthalene bioavailability sensor based on a nahR-gfp fusion in P. putida. The
strain was applied for the detection of naphthalene in water and gas phase. It turned
out that the detection limit of the compound is 50 nM in gas medium and 0.5 μM in
water. Shin (2010) reported that the mutagenesis of the effector binding sites of
regulators involved in the degradation of naphthalene, and salicylate increases the
sensitivity and specificity to polycyclic aromatic hydrocarbons. To increase the sensitivity of the biosensor, eight single mutants (N169A, N169C, N169K, N169S,
R248H, R248M, R248Q, and R248Y) were made at residues 169 central inducer
recognition domain and 248 C-terminal multimerization domains in nahR regulatory gene. The substitution of the amino acids leads into drastic changes in the
microbial response to salicylate, including the increase sensitivity of biosensor. The
effects of these mutations were examined by monitoring expression of a luc reporter
gene under the control of nahR. The biosensors showed response to toxic concentration up to 5 mM. Thus, the mutagenesis method could be overcoming the draw of
low sensitivity in developing microbial biosensor.
S. Ganesan and N. Vasudevan
observed at 50 ng/L for methyl mercury chloride, 0.34 μg/L for phenyl mercury
acetate, and 2.3 mg/L for dimethyl mercury. Willardson et al. (1998) developed
whole-cell biosensor system to sense toluene, and the results were cross-checked
with GC-MS.
4.5.3 Genetically Modified Bacterial Biosensor in Polycyclic
Aromatic Hydrocarbons
Polycyclic aromatic hydrocarbons are found ubiquitous in the environment due to
natural and anthropogenic activities. The anthropogenic activities include petrogenic and pyrogenic activities. It occurs in the environment due to incomplete combustion of fossil fuels, burning of wood and coal, and the natural activities including
volcanic eruption, forest fires, etc. Polycyclic aromatic hydrocarbons are mutagenic
and carcinogenic. Soil can be contaminated with 1 μg/kg to 300 g/kg of polycyclic
aromatic hydrocarbons. There are various methods used for the detection of polycyclic aromatic hydrocarbon, but still they have some limitations. Kuncova et al.
(2011) reported P. putida TVAS with a tod-luxCDABE is sensitive for the bioavailability of organic pollutants. The bioreporter was responsive to 23 organic pollutants, whereas tod reporter mostly identified benzene, ethylbenzene, xylene, and
toluene from wastewater and groundwater samples with 0.5–120 mg/L detection
limit. Using similar strategies, Kohlmeier et al. (2008) reported a highly sensitive
naphthalene bioavailability sensor based on a nahR-gfp fusion in P. putida. The
strain was applied for the detection of naphthalene in water and gas phase. It turned
out that the detection limit of the compound is 50 nM in gas medium and 0.5 μM in
water. Shin (2010) reported that the mutagenesis of the effector binding sites of
regulators involved in the degradation of naphthalene, and salicylate increases the
sensitivity and specificity to polycyclic aromatic hydrocarbons. To increase the sensitivity of the biosensor, eight single mutants (N169A, N169C, N169K, N169S,
R248H, R248M, R248Q, and R248Y) were made at residues 169 central inducer
recognition domain and 248 C-terminal multimerization domains in nahR regulatory gene. The substitution of the amino acids leads into drastic changes in the
microbial response to salicylate, including the increase sensitivity of biosensor. The
effects of these mutations were examined by monitoring expression of a luc reporter
gene under the control of nahR. The biosensors showed response to toxic concentration up to 5 mM. Thus, the mutagenesis method could be overcoming the draw of
low sensitivity in developing microbial biosensor.
S. Ganesan and N. Vasudevan