Page 10 - World Aquaculture Magazine - March 2011

8 March 2011
causing disease. Today, laser-based capture microdissection is an
emerging technology enhancing histopathology to allow research-
ers to precisely isolate specific pathogens from tissue sections, even
with mixed infections. These then, can be isolated for nucleic acid
extraction and molecular diagnostic, genetic and proteomic anal-
ysis (Player et al. 2004, Domazet et al. 2008, Small et al. 2008; Fig-
ure 1). The implementation of histology-based virtual microscopy
is also an emerging technique. Virtual Microscopy allows storage
of a complete clinical and pathology workup consisting of sev-
eral images that are stored in a dedicated server database. This
facilitates rapid effective case management and communication
for teaching or for off site diagnostic review (Figure 2). The use of
digital slides also represents a powerful tool for the assessment of
diagnostic accuracy and quality control programs for diagnostic
laboratories in different parts of the world (UE research founded
program BEQUALM athttp://www.bequalm.org/fishdisease.htm;
Dee 2009, Rocha et al. 2009, Spitsbergen et al. 2009).
Digital imaging has also been applied to transmission electron
microscopy and scanning electron microscopy, replacing the need
for labor intensive films. It is also noteworthy that automated plat-
forms for DNA and RNA extraction are replacing labor-inten-
sive manual methods in some aquaculture molecular diagnostic
laboratories. Time consuming conventional methods for bacterial
identification are being replaced by a strip-concept of dehydrated
biochemical tests (enzymatic and assimilation) in miniaturized mi-
crotubes (API 20 E). Moreover, a fully automated microbial identi-
fication and susceptibility system (VITEK® System) has been intro-
duced for busier clinical laboratories and aquaculture certification
programs (Dodson et al. 1999, Kuen 2007). Whenever the bacteria
strain is difficult to identify, a combination of traditional pheno-
type and genotype (16S rRNA sequencing) tests will continue to be
the gold standard for bacterial identification, but it is also expected
that the emerging DNA-chip technology may progress toward be-
ing a powerful tool reducing the time needed for identification of
bacteria based on amplified highly conservative genes.
An emerging platform combines end-point nucleic acid amplifica-
tion, such as PCR or loop-mediated isothermal amplification, with
dot-blot hybridization or ICT. These emerging methods are allowing
the development of highly specific, sensitive, rapid and cost effective
methodologies for detection of pathogenic microorganisms, which
are less prone to contamination. In addition, these methods can be
applied in resource-poor and ‘point-of-care’ diagnostic settings (et al.
2007, Srisala et al. 2008, Kiatpathomchai et al. 2008, Andrade and
Lightner 2009, Soliman and El-Matbouli 2010; Figure 3). New di-
mensions are being opened for diagnostics with powerful multiplex-
ing platforms for simultaneously testing for multiple different patho-
gens using emerging Luminex xMAPTM and microarray technology.
Although these technologies are just beginning to be applied to aqua-
culture, they are likely to become more widely used in aquatic animal
diagnostic laboratories in the future (Adams and Thompson 2009).
All of these applications have the potential to greatly improve aqua-
culture animal health and may soon be commercialized, but at pres-
ent only one commercial kit for a semi-nested WSSV test has been
evaluated by OIE for application in aquaculture. Thus, it is vitally
important that these novel and emerging diagnostic methods should
be certified by a governmental accreditation program to assure that
they are ‘accurately fit for purpose’ before they can be recommended
to OIE (Lightner et al. 2009) .
Fig. 1. The laser-based capture microdissection is an
emerging technology coming to the aid of histopa-
thology to allow researchers to precisely isolate spe-
cific pathogens from H&E tissues sections mounted
on a thermoplastic film slide. (Photo by Thales P. D.
Andrade).
Fig. 2. The implementation of digitalized slides into
histology is also an emerging technique in aquaculture.
H&E stained sections of hepatopancreas of the shrimp
Litopenaeus vannamei at 400x (left) and gills of the fish
Rachycentrum canadum at 100X (right). Photos: per-
sonal virtual database of Thales P. D. Andrade.
Fig. 3. Schematic diagram of RT-LAMP-NALF assay for
detection of infectious myonecrosis virus (Andrade and
Lightner 2009).