Microbial forensics combines principles of public
health epidemiology and law enforcement to identify patterns in
a disease outbreak, determine which pathogen may be involved, and
trace
the organism to its source.
Since investigators must consider potential
prosecution and presentation of evidence in court, biocrime investigations
demand careful controls and standards for validation and evaluation
of technologies and the data they produce. Scientists can easily
evaluate new methods of detecting organisms implicated in a bioterrorist
attack, but taking the resulting evidence into a court is another
matter. Any microbial evidence, such as anthrax spores, that
links to a suspect has to meet stern standards.
Here Comes the
Nudge
We¹re
not talking about a jury of your scientific peers, we¹re
talking about lawyers, judges, juries, said Abigail Salyers,
professor of microbiology at the University of Illinois and
past president
of the American Society of Microbiology. The consequences are
not just having a paper rejected by a journal, but rather of
sending someone to jail.
Salyers said even if the anthrax perpetrator
was caught, it might not be as easy to achieve a conviction,
especially if the spores are part of the physical evidence. If
PCR based tests
on the spores found in the suspect¹s possession were the same
DNA signatures as the spores found in the letters, that¹s fine
for scientists.
But if you took that into court, all sorts
of questions arise: What does 'same¹ mean? Does it
have to be 100 percent identical? Even if it¹s 100 percent
identical, does it really prove that it¹s the same?² Salyers
said.
Salyers sees two main problems. First, although the technology
for doing DNA based and other molecular analyses is widely used
and universally
accepted in the research community, the kind of rigorous
validation and development of appropriate quality control standards
for
the use of this technology in forensics is still not well developed.
Research
is under way, but it might still be fairly easy for a defense
lawyer to raise questions about accuracy and interpretation, much
as happened
in the infamous Simpson trial, she said.
This is not necessarily
due to weaknesses in the technology but rather to the fact
that scientists had not thought about forensic uses and thus had
not developed the
validation and quality control guidelines appropriate for
legal application. This should not be a difficult goal to reach,
however, because rigorous
validation and quality control guidelines have been developed
for the use of biological technologies in hospital laboratories.
Mutation
Rate
The second problem, which is currently being investigated,
is the mutation rate. If you want to show that the strain of
bacterium that produced a spore found in the suspect¹s home or office
is the strain used in the anthrax attack and not a strain that was
originally in the soil and was tracked into the location, there is
no problem. Different strains of Bacillus anthracis differ enough
at the DNA sequence level that even a partial genome sequence of
the two strains would be sufficient to make the distinction because
there would be a number of differences, what you might call a DNA
sequence ³fingerprint,² Salyers said. ³
If one is trying
to locate the laboratory where the anthrax attack strain
came from, however, such distinctions would be more difficult since
most laboratories
shared derivatives of the same strain,² she said.
Here, natural
mutations over the past several decades that occurred in
the strain due to numerous passages in the laboratory might be
sufficiently
abundant to allow the strain obtained from one laboratory
to be distinguished from that obtained from another, but since
the differences will probably
be few in number, they will be less convincing than for
strains isolated in different geographical locations.
How much
difference is there?
This is currently being investigated. ³Scientists believe that
B. anthracis mutates very slowly, so it may be difficult to have
a differentiation that holds up in court,² Salyers said.
There
are errors that occur during the DNA sequencing process
(about 0.1% at present), but this problem can be solved by resequencing
an area
more than once. This issue has led scientists to consider
other molecules besides DNA, such as those found in traces of the
growth media used
to cultivate the strain (if those could be detected),
which might differ from one laboratory to another.
Reasonable Doubt
There are
other wrinkles in this fabric potentially attractive
to defense council. Other species of Bacillus, such as Bacillus
thurengensis (the organic
gardener¹s friend), have genome sequences strikingly similar
to that of B. anthracis, the cause of anthrax. The main difference
is two plasmids; extrachromosomal segments of DNA that make B. anthracis
capable of causing disease, whereas B. thurengensis and most other
Bacillus species are innocuous for humans. Some plasmids can be transferred
from one bacterium to another, known informally to bacteriologists
as bacterial sex.
No one knows if an innocuous strain of another
species could be rendered virulent, but you can imagine
ways in which a defense attorney might use this sort of fact to
place doubt in
the minds of jurors with no background in biology,² Salyers
said.
The reverse problem occurs in the cases of some viruses with
RNA genomes, such as HIV and influenza, which are mutating
so rapidly that it is possible to find different sequence variants
in the same
person.
Granted these differences are small in number, but
deciding what the Œsame¹ means when comparing the strain
from the presumed source with the strain that infected a victim could
raise interpretation problems,² Salyers said.
Federal Response
Traditionally, law enforcement has had the role of
crime investigation, although it is now being forced to confront
the new biocrime challenge
by partnering with the scientific community. The
anthrax attacks and subsequent public reactions revealed the need
for an
infrastructure
with analytical tools and knowledge base to rapidly
provide investigative leads and help determine who was responsible
for the crime, the source
of the agent, and how and where the weapon was produced.
While
there are a few well-developed practices for handling
and analyzing pathogenic agents, most of these assays address epidemiological
concerns
and do not provide sufficient information on the
strain or isolate to allow law enforcement to identify the source
of the evidence sample, said
the Federal Bureau of Investigation¹s Bruce Budowle in a recent
paper (Science 2003 301: 1852-1853).
Additional assays for individualization
of microbial strains is needed, Budowle said. For
example, determining the microbe sent in a letter as B. anthracis
identifies the causative
agent. At this point, anyone with access to B. anthracis
could be considered a suspect. But determining it was the Ames
strain, an
uncommon strain in nature, limits the investigation
to those who had access to that specific strain.
All of this must be defined
adequately and validated sufficiently to meet forensic
needs, Budowle
said.
The problem is there aren¹t many laboratories with adequate
biocontainment facilities to handle forensic cases. So far, there
is little guidance on the logistics and financial commitment required
to construct a microbial forensics laboratory or to retool partner
labs to perform microbial forensic work.
The FBI is leading the effort
to address these issues, in 2002 having initiated
the Scientific Working Group on Microbial Genetics and Forensics
(SWGMGF). The goal
of SWGMGF is to provide an avenue for government,
academia, and private sector scientists to develop guidelines related
to the operation
of microbial forensics.
Combating bioterrorism is a challenge
to all of us, Budowle said. To be proactive,
the U.S. government has formalized the discipline
of microbial forensics¹ to
deter and attribute perpetrators of such acts.
The FBI has
spawned scientific working groups for other forensic
disciplines, the most notable being the Scientific Working Group
on DNA Analysis
Methods, whose success can be seen by the common
use of DNA analysis in crime labs, the establishment of standards,
and the wide-spread
acceptance of DNA analysis in the courts.
Likewise, the SWGMGF aims
to contribute to the infrastructure and development
of microbial forensics. According to Budowle, the recommendations
of the SWGMGF
will be implemented in the form of a national microbial
forensics laboratory, called the National Bioforensics Analysis
Center, part
of the National Biodefense Analysis and Countermeasures
Center.
Microbial forensic experts participated in a colloquium
last year that dealt
with evidence gathering, organism identification,
organism source tracing, and investigative techniques, the findings
of which are
summarized in the report, Microbial Forensics:
A Scientific Assessment,² available online at http://www.asm.org/Academy/index.asp?bid=17994.
