Text: Wouter Oude Groothuis
Translation: Word’s Worth | Photography: Marco Vellinga
Forensic scientists can now determine who has touched a given object by sampling its surface. The technique, known as microbial profiling, is a big advance in metagenomics.
Text: Wouter Oude Groothuis
Translation: Word’s Worth | Photography: Marco Vellinga
By comparing the microbial profiles taken from a suspect’s hand and from an object found at a crime scene, it is possible to assess the probability that the suspect was present there when the crime was committed. Humans, on average, have 150 types of microbes on their hands. This population is unique enough to be used as a means of identification. Moreover, microbial profiles show the origins of the tissue found in a trace. Peter de Knijff, professor of population and evolution genetics at Leiden University Medical Center’s department of Human Genetics, explains why this is such an exciting breakthrough. “The main problem in forensics is usually the lack of DNA. A human body cell contains just 6 picograms of DNA. The success of DNA testing depends largely on the amount of cell material on the surface and the time that has elapsed since it was last touched.” As De Knijff, who also heads the forensic lab for DNA testing (FLDO), points out, “microbial profiling is a totally different kettle of fish, because there’s usually plenty of bacteria available at the crime scene.”
“Microbial profiling’s main advantage over DNA testing is that bacteria, unlike DNA, are usually abundant at a crime scene”Professor Peter de Knijff, expert in forensic metagenomics at Leiden University Medical Center
Currently, the microbial DNA sequence—the nucleic acid sequence—of 450 samples can be analyzed in parallel. This number is expected to rise to 1,000 samples in the near future. The genome contains all the information needed for forensic analysis; not only the DNA—the genes—but also the parts that do not code for amino acids. “In genome sequencing, it’s important to differentiate between novo sequencing and re-sequencing,” says De Knijff. “In novo sequencing, you’re sequencing an unknown genome for the first time. In resequencing, you already have a reliable genome to compare your own results to.” Novo sequencing requires at least 60 times the genome to arrive at a somewhat reliable assembly. First, researchers cut up all the DNA from a pure culture into random pieces ranging in size from a single base pair to several thousand base pairs. Next, they determine the nucleic acid sequence of DNA fragments that are 6 to 10 times the length of the genome. This reveals overlaps that can be used to determine the order of the fragments in the whole genome and to verify the accuracy of the sequencing. For re-sequencing, only 10 to 15 times the genome is usually needed, depending on the goal, De Knijff estimates.
“The main problem in forensics is usually the lack of DNA material”Professor Peter de Knijff, expert in forensic metagenomics at Leiden University Medical Center
According to De Knijff, the future of forensic science lies with metagenomic shotgun sequencing. This technique allows scientists to first isolate and then parallel sequence every gene of every organism in a complex sample. Known as Massive (or Massively) Parallel Sequencing (MPS), this high-throughput method is used to sequence all DNA molecules: microbes, viruses, human DNA, plants, pollen, and so on. This is not possible using capillary sequencing or polymerase chain reaction (PCR) techniques. De Knijff explains why this works well in forensics: “Because you use trace material, you’re often dealing with degraded DNA, which will probably always give you a less complete and reliable human genome. But that’s irrelevant, as long as you get the parts of the genome that you’re most interested in.”
MPS is still a thing of the future, De Knijff says. “We’re limiting ourselves to human DNA and microbes, or microbiome analysis, as it’s called in the field. Over time, it will become an all-in-one analysis.”
Possible sequencing techniques are methylation sequencing and Next Generation Sequencing (NGS) technologies using DNA, RNA. However, De Knijff is less than enthusiastic about these methods. “For methylation sequencing, you need truckloads of DNA, which makes it unsuitable for forensics for the time being. What we most likely are going to do is DNA and RNA sequencing, either directly, or by way of a cDNA1 translation. No doubt those two techniques will provide us with lots of useful information, even if the results are incomplete.”
The microbial profile of a hand smear is highly specific. In one test, researchers tried to identify who had used a particular computer mouse. They took a hand smear from 270 individuals, and identified the mouse user with 70-90% accuracy. As De Knijff says: “In forensics, we’re always dealing with a fait accompli. All we have is one trace. It’s no use trying to figure out how we can get more trace material, because we have no influence on that. Therefore, it makes far more sense to study how we can squeeze as many results as possible from a minimum amount of trace material.”
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1: cDNA = copy or complementary DNA acquired through reverse transcription from mRNA and therefore contains neither introns (portions of genes that do not code for amino acids) nor signal peptides (the specific amino acid sequence).
