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45 Biosensors for Sensitive Detection of Agricultural Contaminants, Pathogens and Food-Borne Toxins 873the most toxic and over 20 different analogues have been de-
scribed (Fusetani and Kern 2009), with a range of associated
toxicity levels. PSPs are known to be produced byAlexandrium,
GymnodiniumandPyrodiniumspecies. Infected organisms can
include clams, crabs, cockles, cods, gastropods, herrings, lob-
sters, mackerel, mussels, oysters, pufferfish and scallops. Symp-
toms can be neurological and, less frequently, gastrointestinal,
including nausea, vomiting and diarrhoea. These symptoms can
appear from 15 minutes to 10 hours after ingestion of contam-
inated shellfish, any may also be associated with the onset of a
tingling, a prickly feeling in the toes and fingertips, a burning
sensation of the lips and skin, dizziness, numbness in the mouth
and extremities, headache, ataxia and fever. Individuals may also
experience a feeling of calmness and serenity. Severe poisoning
can lead to lack of coordination and respiratory distress that can
lead to death within 2–25 hours of intoxication.
Fonfr ́ıa et al. (2007) developed a SPR inhibition assay using
an anti-GTX2/3 antibody and a STX-immobilised CM5 Biacore
surface. Here, STX in solution was detected by competition for
binding to the anti-GTX2/3 antibody. The biosensor detected
a number of different saxitoxin analogues, including dcSTX,
GTX2/3, dcGTX2/3, GTX5 and C1/2 (with slightly higher sen-
sitivity than STX). Significantly, the antibody had low cross-
reactivity for neoSTX, GTX1/4 or decarbamoyl neosaxitoxin.
The biosensor platform was subsequently shown to be effective
in a number of different shellfish matrices, including mussels,
clams, cockles, scallops and oysters, with a detection limit be-
tween 2 and 50 ng/mL observed. More recently, Taylor et al.
(2008) devised a novel SPR immunosensor for the low molec-
ular weight toxin, tetrodotoxin (TTX), using a polyclonal anti-
TTX antibody as a biorecognition element. TTX was chemically
immobilised on a gold film with a mixed SAM, which reduced
non-specific binding at the surface. The inhibitory concentra-
tion causing 20% displacement (IC20)for the assay was approxi-
mately 0.3 ng/mL, and the corresponding concentration for 50%
displacement (IC 50 ) was approximately 6 ng/mL. In summary,
these examples clearly demonstrate how biosensors can be used
to monitor potentially fatal contaminants in water and marine
life, which has a direct impact on the food industry.LEGISLATION
In this chapter, a variety of different biosensor-based platforms
used to detect a range of different molecules such as pesti-
cides, herbicides, mycotoxins and phycotoxins are described.
It is important to consider the legislation associated with the
monitoring of quality. The Food and Agriculture Organisation
of the United Nations (FAOSTAT) (http://faostat.fao.org/) is an
international body whose overall aim is to protect the health
of consumers. FAOSTAT has an online database (The Codex
Alimentarius: Pesticide Residues in Food Maximum Residue
Limits; see ‘Websites of Interest’), which details the MRLs for
pesticides in commodities such as fruit and vegetables. The es-
tablishment of a MRL (usually expressed as mg/kg) is dependent
on good agricultural practice for pesticides. This relates to where
the highest detectable residues anticipated (when a pesticide-
containing product is applied to a commodity to remove con-taminants) are intended to be toxicologically acceptable. Under
these arrangements, the important point is that residue levels do
not pose unacceptable risks for consumers by being present in
values that exceed these levels.
Similar legislation exists within the European Union (EU).
Within the EU Directive (91/414/EEC), the Plant Protection
Products Directive (‘The Authorisations Directive’: 1991) was
proposed with the aim of harmonising the overall arrangements
for authorisation of plant protection products within the EU.
However, legislation relating to MRLs of pesticides in cere-
als, fruit, vegetables, foods of animal origin and feeding stuffs
was substantially amended several times since this legislation
was developed. A single act has replaced the amended original
Directive Regulation (EC) No. 396/2005. This establishes max-
imum levels of pesticide residues permitted in or on food and
feed of plant and animal origin. These MRLs include levels that
are specific to particular foodstuffs that are intended for human
or animal consumption, and a general limit that applies where
no specific MRL has been established.
As infants have developing immune systems, fruit and
vegetable-based baby foods need to be rigorously monitored,
as the presence of low concentrations of pesticides could be of
significance. The EU has set an MRL for any given pesticide in
infant foods of a concentration not exceeding 10μg/mL. This
information is very significant for the use and future applications
of biosensor-based detection methods.
The EU has established an upper safe limit of 20 mg/kg
for total ASP toxin content in the edible part of the mollusc
(EEC 1991), with HLPC selected as a reference method. The
European Commission have set tolerable levels for OA, DTXs
and PTXs present in edible tissues at 160μg OA equivalent/kg
shellfish and a maximal level of YTXs of 1 mg YTX equiv-
alent/kg shellfish (European Commission Decision 2002). The
mouse or rat bioassay is the reference method for DSP detec-
tion. The EU directive 91/492/EEC allows a maximum level of
PSP toxin of 80μg STX eq/100g shellfish meat. The European
Commission also decided in 2002 (2002/226/EC) to allow a con-
ditional harvesting of scallops with a concentration of domoic
acid (DA) in the whole body exceeding 20 mg/kg but lower than
250 mg/kg, based on the limit of ASP, imparted by Council Di-
rective 91/492/EEC (1991). More recently (2009), the European
Food Safety Authority (EFSA) recommended that it was safe to
consume shellfish containing less than 4.5 mg domoic acid/kg,
so as not to exceed the acute reference dose of 30μg DA/kg
body weight (equivalent to 1.8 mg DA per portion for a 60 kg
adult) (EFSA 2009a, EFSA 2009b).CONCLUSION
This chapter has provided a concise review of how biosensors
may be applied to monitoring the quality of fruit and vegetable
produce, and also for the detection of pathogens (bacterial and
fungal) and associated toxins. Examples are given where such
applications have been successfully employed, while recent de-
velopments in the area have focused on the development of low
and high-density arrays capable of multiple analyte monitoring
and portable or disposable platforms for in situ monitoring. With