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Original Article

Analysis of Microbiome DNA on Frequently Touched Items and

from Palm Prints
Xue Yao, Wenli Liu1, Junping Han, Guangqian Pei1, Yigang Tong1, Yaping Luo
School of Criminal Science and Technology, Peoples Public Security University of China, 1State Key Laboratory of Pathogen and Biosecurity,
Beijing Institute of Microbiology and Epidemiology. Beijing, China

Abstract
Limited by the sensitivity of laboratory techniques, conventional human DNA analysis of touch DNA on frequently used items and prints does
not always provide satisfactory results. In this study, microbiome DNA on personal computers, cell phones, and palm prints was analyzed and
compared. After sample collection, DNA extraction, polymerase chain reaction amplification, library preparation, and sequencing, data were
analyzed using the QIIME 1.8.0 software. Weighted unifrac distance between the right palm skin and the right side of a keyboard, the right
palm skin and the mouse, and the left side of the keyboard and the left palm skin was 0.258850, 0.265474, and 0.214098, respectively. Even
after palm prints were left for 1week, microbial community structures were still quite similar to those of samples collected from the palm
skin on the day they were left(weighted unifrac distance was 0.270885).
Key words: Cell phone, keyboard, metagenomics, palm prints

Introduction

Materials and Methods

The human skin microbiome refers to the entire population

of microbes that colonize the human skin including bacteria,
fungi, and viruses. Bacteria density on the human skin may
be as high as 107 cells/cm2[1] and can be readily transferred
to surfaces upon touching.[2] Nextgeneration sequencing
technology has revealed high diversity in skinassociated
bacterial communities.[37] This method may be useful for
evaluating the residual skin microbiome left on objects for
forensic identification. By calculation of distances from samples
and their donators, it will be possible to estimate whether items
or palm prints belong to one specific person or not.

Sample collection and DNA extraction

Traditional human DNA analysis of touch DNA does not

always provide satisfactory results because of the limited
content and easy degradation of human DNA. The microbiome
DNA is abundant in touch DNA, and these organisms are
much more stable because of complicated cell wall structures.
In this study, microbiome DNA on personal computers, cell
phones, and palm prints was analyzed and compared with
those collected from the palm skin. Microbial community
structures of different samples were compared to evaluate the
similarities between these frequently used items, palm prints,
and palm skin.
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DOI:
10.4103/2349-5014.184190

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In the first part of this study, samples were collected from

a personal computer and cell phone used by a 29yearold
healthy female, which she used with both palms. In the
second part of this study, samples from the palm skin and
palm prints were collected. Two pairs of palm prints were
provided by the same female. After washing hands, plastic
gloves were worn for at least 1h, and then palm prints were
made on sterile plastic sheets. One pair of palm prints was
collected and DNA was extracted within 1h. The other pair
was placed in a spare room devoid of human contact for
1week. Anote was posted to inform that anyone set foot
in the room not to touch experiment materials. In addition,
clean plastic sheets were placed to collect the microbiome
from the air. After 1week, samples from the subjects palm
skin were also collected.
Address for correspondence: Prof. Yaping Luo,
Muxidi South No.1, School of Criminal Science and Technology, Peoples
Public Security University of China, Beijing, China.
EMail:lyp6698@163.com
This is an open access article distributed under the terms of the Creative Commons
AttributionNonCommercialShareAlike 3.0 License, which allows others to remix,
tweak, and build upon the work noncommercially, as long as the author is credited and
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For reprints contact: reprints@medknow.com

How to cite this article: Yao X, Liu W, Han J, Pei G, Tong Y, Luo Y. Analysis
of Microbiome DNA on Frequently Touched Items and from Palm Prints.
J Forensic Sci Med 2016;2:74-7.

2016 Journal of Forensic Science and Medicine | Published by Wolters Kluwer - Medknow

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Yao, etal.: Analysis of Microbiome DNA on Frequently Touched Items and Palm Prints

During sample collection, sharp cotton swabs were dipped with

0.15 M NaCl and 0.1% Tween20 solution and then swabbed
repeatedly to obtain as much DNA as possible. Next, the swabs
were cut and placed into a 1.5mL centrifuge tube containing
600 L 0.15 M NaCl and 0.1% Tween20 solution. After
vortexing for 2min, swabs were discarded and the solution
was ultrasonicated at 18% power(ultrasonication for 2 s, pause
for 3 s) for 5min(JY88II DN Ultrasonic signal generator;
Nanjing Xinchen Biotechnology Co., Ltd., Nanjing, Jiangsu,
China). After pretreatment, DNA was extracted using a Roche
High Pure PCR Template Preparation Kit(Roche Diagnostics
GMbH, RischRotkreuz, Switzerland). Finally, DNA was
dissolved in 100 L ddH2O and concentration was measured
using Qubit 2.0 Fluorometer(Life Technologies Corporation,
Carlsbad, CA, America).

Polymerase chain reaction amplification, sequencing,

and analysis

Primers targeted to the V1V2 region of 16S ribosomal RNA,

F27 and R338, were used in this study, with 4bp tags connected
to the 5 ends to differentiate between samples. The polymerase
chain reaction(PCR) system contained 1 L(10pM) of each
forward and reverse primer, 25 L of Q5 HighFidelity 2X
Master Mix(New England Biolabs, Ipswich, MA, USA), and
ddH2O containing DNA to 50 L(approximately 3ng DNA in
each PCR reaction). Anegative control with no DNA template
was used to maintain the sensitivity of the analysis. Samples
were initially denatured at 95C for 5min and then amplified
for 30cycles at 95C for 30 s, 55C for 30 s, and 72C for 30 s.
Afinal extension for 7min at 72C was conducted at the end
of the program to ensure complete amplification of the target
region. Next, the libraries were prepared and sequenced on
an Ion Torrent Personal Genome Machine(Life Technologies
Corporation, Carlsbad, CA, USA). Sequencing results were
analyzed using QIIME 1.8.0.[8] Only those sequences>200bp
in length with an average quality score of at least 20 and
no ambiguous characters were included in the analysis.
Similar sequences were clustered into operational taxonomic
units(OTUs) with a minimum identity of 97%.

Results and Analysis

Microbial community structures of frequently touched
items
Overall conditions of sequencing results
The total number of sequences after filtering was 74,815;
average OTU number per sample was 1005, and average
species observed per sample was 415.

Alpha diversity of different samples

The diversity indices of each sample are calculated and the
relative abundance of different samples at the genus level
is shown in Figure1. In Figure1, different colors represent
different genus and lengths of bars represent percentage
of different genus. Microbial community structures on the
downside of the cell phone are quite different from that in
other samples.

Beta diversity of different samples

Weighted unifrac distances between samples were calculated;
the smallest the value of distances between samples indicates
the most similar microbial structure. The most similar samples
and their distances are listed in Table1.
Weighted unifrac distance between the right side of the
keyboard and righthand skin was 0.258850. Weighted unifrac
distance between the mouse and righthand skin was 0.265474.
Microbial community structures between the left side of the
keyboard and her left palm skin were also similar(weighted
unifrac distance was 0.214098). However, microbial
community structures from the downside of the phone were
quite different between both hands(weighted unifrac distances
were 0.414164 and 0.455530). An unweighted pair group
method with arithmetic mean(UPGMA) clustering tree based
on weighted unifrac distances is shown in Figure2.

Microbial community structures of palm prints

Overall conditions of sequencing results

The total number of sequences after filtering was 270,312;
average OTU number per sample was 1071, and average

Figure1: Relative abundance at genus level in the first part of the

experiment. LP: Left palm skin, RP: Right palm skin, UP: Upside of
phone, DP: Downside of phone, LK: Left side of keyboard, RK: Right
side of keyboard, M: Mouse

Table1: Most similar samples weighted unifrac distance

in the first part of experiment
Most similar samples
Left side of keyboard, right
side of keyboard
Mouse, right side of keyboard
Downside of phone, right
side of keyboard
Left palm, right palm
Upside of phone, mouse

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Weighted unifrac distance

0.178067
0.192727
0.294587
0.153575
0.280976

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Yao, etal.: Analysis of Microbiome DNA on Frequently Touched Items and Palm Prints

species observed per sample was 367. Among all samples

sequenced, samples collected from the blank plastic sheet to
collect the microbiome in the air recovered only 98 OTUs,
which is significantly fewer than for other samples.
Alpha diversity of different samples
The diversity indices of each sample are calculated and the
relative abundance of different samples at the genus level is
shown in Figure3. The figure shows that the relative abundance
of samples collected from blank plastic sheets at the genus
level is quite different from that of other samples.

Figure2: Unweighted pair group method with arithmetic mean clustering

tree based on weighted unifrac distance in the first part of experiment

Beta diversity of different samples

Weighted unifrac distances between samples are calculated,
and the most similar samples are listed in Table2. The distance
between samples collected from the blank plastic sheet and
other samples was significantly higher(0.4195920.780260).
AUPGMA clustering tree based on weighted unifrac distance
is shown in Figure4. Even after palm prints were left for
1week, microbial community structures remained quite similar
to samples collected from the palm skin(YR0) on the day they
were obtained.

Discussion and Conclusion

In this study, microbial community structures on a keyboard
and mouse successfully matched the corresponding palm skin.
However, microbial community structures collected from the
downside of the cell phone were quite different from the palm
skin or computer keyboard. This may be because the microbial
community was transferred from various locations onto the
phone. Even after the palm prints were left for 1week, the
microbial community structures remained quite similar to
samples collected from the palm skin on the day they were
left, and the microbiome in the air did not significantly change
the microbial community structures of palm prints over a
1week period.
This study showed a potential method to compare microbial
community structures between palm skin, items, and palm
prints. After largescale sample analysis, threshold values can
be obtained for forensic application. For example, if samples
from 100 persons palm skin and cell phone were collected,
statistical analysis of weighted unifrac distances can be made
according to this study. If most(for example, 90%) of the
distances between ones palm skin and his/her cell phone are
below 0.35 (for instance), while distances between ones palm
skin and others cell phone are above 0.5(for instance), then

Figure3: Relative abundance at genus level in the second part of the

experiment. YL0: Samples collected from left palm skin on the day when
palm prints were printed, YR0: Samples collected from right palm skin
on the day when palm prints were printed, YLP0: Samples collected from
left palm print on the day when palm prints were printed, YRP0: Samples
collected from right palm print on the day when palm prints were printed,
YL1: Samples collected from left palm skin 1week after palm prints were
printed, YR1: Samples collected from right palm skin 1week after palm
prints were printed, YLP1: Samples collected from left palm print 1week
after palm prints were printed, YRR1: Samples collected from right palm
print 1week after palm prints were printed, AIR1: Samples collected from
blank plastic sheet 1week after palm prints were printed

Table2: Most similar samples weighted unifrac distance

in the second part of experiment
Most similar samples
YRP1, YLP1
YRP0, YLP0
YR1, YL1
YL0, YL1
YR0, YRP1

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Weighted unifrac distance

0.197390
0.165754
0.111076
0.219494
0.270885

Figure4: Unweighted pair group method with arithmetic mean clustering

tree based on weighted unifrac distance
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Yao, etal.: Analysis of Microbiome DNA on Frequently Touched Items and Palm Prints

these values can be used as threshold. When a cell phone was

discovered from a crime scene that may belong to one suspect
and weighted unifrac distance between the cell phone and the
suspect was 0.2, then chances are 90% of it belonged to this
suspect.
Additional studies are required to determine how microbial
community structures change over longer time periods, and
a large number of subjects should be evaluated. Successful
implementation of this technology is beneficial for promoting
the key evidence usage ratio and improving the technical
support capabilities in criminal investigations.

Financial support and sponsorship

This work has received funding from the State Key Laboratory
of Pathogen and Biosecurity (xxhz201510), and Shanghai
Research Institute of Criminal Science and Technology
(2014XCWZK14).

Conflicts of interest

There are no conflicts of interest.

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