1. 1
Circulating mitochondrial DNA as an effect-biomarker
after exposure to halo-alkane based pesticides
Lygia T. Budnik1, Stefan Kloth1, Xaver Baur2,,
Alexandra M. Preisser 1, Heidi Schwarzenbach 3
1 Institute for Occupational and Maritime Medicine, Department of Occupational Health, School of
Medicine, University of Hamburg, Hamburg, Germany
2 Institute for Occupational Medicine, Campus Benjamin Franklin, Charité-School of Medicine,
Berlin, Germany.
3 Department of Tumor Biology. School of Medicine, University of Hamburg, Hamburg, Germany
LBudnik@uke.de
There is a need for a panel of suitable effect biomarkers for
detection of environmental chemical exposure
leading to the initiation or progression of degenerative diseases
or potentially cancer.

2. 2
Since peripheral blood contains increased levels of circulating
cell-free DNA in diseased individuals, we aimed to evaluate this
DNA as effect biomarker recognizing vulnerability after
exposure to environmental chemicals.
intox. target organes
release cell-free DNA
in to the blood
stream
zf-DNA-Extraktion
Container and other transport units are un-loaded deep within the country sides outside
the harbour areas. Workers and Bystanders can be exposed to fumigant residues in
storage rooms
Short-term high exposure Symptoms: (latency period of 2-48 h -3 days)
 Central nervous system depression, neuropsychiatry abnormalities
(headache, nausea, dizziness)
 Inflammation of the bronchi and lung, Pulmonary edema
(flu like symptoms, phlegm, chest pain, shortness of breath)
“Pesticides used as fumigants are often at least as poisono-
us to humans as to the pests against which they are used”
IMO, 2005 (International Maritime Organisation).

3. 3
164 subjects with presumed intoxication to fumigants
(plus 30 controls).
78% male and 22% female subjects at median age of 44; 66% smoker, 34% non-smoker
The exposure assessment was performed with human
biomonitoring (methyl bromide, ethylene dichloride,
dichloromethane, other halo-alkanes).
Exposure-
Biomonitoring Diagnosis
Ambient-
monitoring Effect-
Biomonitoring
Budnik et al., 2013 PLoS ONE 8(5)

4. 4
Confirmed: 86 exposures to halo-alkanes above the non-cancer
reference doses (RfD), but (mostly) lower than the occupational
exposure limits
Data from: Budnik et al., 2013 PLoS ONE 8(5)
. Cell-free DNA may provide information from intoxicated organs
Data from: Budnik et al., 2013 PLoS ONE 8(5)

5. 5
Exposure was associated with elevated serum levels of
circulating mitochondrial DNA (in size of 79 bp, mtDNA-79).
serum mtDNA-79 level serum mtDNA-79 level
All exposed vs Controls
MeBr exposed vs Controls
Data from: Budnik et al., 2013 PLoS ONE 8(5)
serum mtDNA integrity
p=0.0158
serum mtDNA-230 level
p=0.005
The increased integrity of mtDNA (mtDNA-230/mtDNA-79) in
exposed individuals implicates apoptotic processes.
Data from: Budnik et al., 2013 PLoS ONE 8(5)

6. 6
Patient subgroups
P1 current intoxication
P2 short-term past exposure (weeks, up to 4 months)
P3 long-term past exposure (interim time at least 5 months)
Patient subgroups
P1 current intoxication
P2 short-term past exposure (weeks, up to 4 months)
P3 long-term past exposure (interim time at least 5 months)
The relative amounts of mtDNA-79 in serum were positively associated with the lag-time
after intoxication of these chemicals (r=0.99, p<0.0001).
The relative amounts of mtDNA-79 in serum were positively associated with the lag-time
after intoxication of these chemicals (r=0.99, p<0.0001).
Data from: Budnik et al., 2013 PLoS ONE 8(5)
AUC=0.79
95% CI (0.64-0.95)
p=0.004
Serum mtDNA-79 level
AUC=0.77
95% CI (0.65-0.88)
p=0.003
mtDNA-Integrity
The specificity for this biomarker increased from 30% to
97% several months post-exposure in patients with
intoxication symptoms.
Specificity and sensitivity
Data from: Budnik et al., 2013 PLoS ONE 8(5)

7. 7
ConclusionsConclusions
Our findings:
 show enhanced cell free mtDNA levels in patients exposed to halogenated
hydrocarbone pesticides
 corraborate the data showing increased mtDNA in patients exposed to low levels
of benzene (<100 ppb)
Our findings:
 show enhanced cell free mtDNA levels in patients exposed to halogenated
hydrocarbone pesticides
 corraborate the data showing increased mtDNA in patients exposed to low levels
of benzene (<100 ppb)
The findings indicate that mitochondrial DNA has a potential to serve as a biomarker
recognizing vulnerable risk groups after exposure to toxic/carcinogenic chemicals
The findings indicate that mitochondrial DNA has a potential to serve as a biomarker
recognizing vulnerable risk groups after exposure to toxic/carcinogenic chemicals
But also:
 Open many questions regarding possible molecular mechanisms
But also:
 Open many questions regarding possible molecular mechanisms
Thank you