Steenblock et al. Int J Neurodegener Dis 2018, 1:003
Volume 1 | Issue 1
International Journal of
• Page 1 of 4 •
Citaon: Steenblock DA, Ikrar T, Antonio ASS, Wardaningsih E, Azizi MJ (2018) Amyotrophic Lateral
Sclerosis (ALS) Linked to Intesnal Microbiota Dysbiosis & Systemic Microbial Infecon in Human
Paents: A Cross-Seconal Clinical Study. Int J Neurodegener Dis 1:003.
Accepted: September 10, 2018; Published: September 12, 2018
Copyright: © 2018 Steenblock DA, et al. This is an open-access arcle distributed under the terms
of the Creave Commons Aribuon License, which permits unrestricted use, distribuon, and
reproducon in any medium, provided the original author and source are credited.
Steenblock et al. Int J Neurodegener Dis 2018, 1:003
Amyotrophic Lateral Sclerosis (ALS) Linked to Intesnal Microbi-
ota Dysbiosis & Systemic Microbial Infecon in Human Paents:
A Cross-Seconal Clinical Study
David A Steenblock1#, Taruna Ikrar2,3,4*#, Andrew S San Antonio2,4, El Wardaningsih4 and Masoud J
1Steenblock Research Instute, USA
2Internaonal School of Biomedical Sciences, Pacic Health Sciences University, USA
3Cellcure Center, The Indonesia Army and Presidenal Central Hospital, Indonesia
4Neurodegenerave Disease Research Program, Pacic Health Sciences University, USA
#These authors (T.I. & D.A.S.) contributed equally to this work.
*Corresponding author: Prof. Dr. Taruna Ikrar, MD, M. Pharm, PhD, Cellcure Center, The Indonesia Army and Presidenal
Central Hospital, (RSPAD Gatot Subroto), Jln. Abdul Rahman Saleh No. 24, Jakarta 10410, Indonesia; Internaonal School
of Biomedical Sciences and Neurodegenerave Disease Research (NDR) Program, Pacic Health Sciences University
(PHSU), 5401 Business Park South, Suite 107, Bakerseld, California, 93309, USA; Tel: +62-21-3441008; +62-21-3840702,
+1 661-200-7210 , Ext 5005, Fax: +62-21-350619, +1-661-200-7216
by progressive degeneraon of both upper and lower
motor neurons, and manifests as progressive physical
paralysis that culminates in death (most oen due to
respiratory failure) within an average of 3-5 years from
disease onset [1-3]. ALS is the most common adult-onset
motor neuron disease (MND), with a worldwide annu-
al incidence of about 2 per 100,000 . The worldwide
prevalence of this fatal disease is expected to increase
by up to 69% over the next 25 years . ALS aects in-
dividuals of all races and ethnicies (both male and fe-
male); however, whites, males, non-Hispanics, individu-
als aged > 50 years, and those with a family history of
ALS are more likely to develop this disease [6-8]. There
is currently no cure for ALS, and its eology remains elu-
sive. The only 2 drugs approved by the FDA to treat this
disease are riluzole (a glutamate inhibitor) and edara-
vone (an an-oxidant), both of which fail to halt the pro-
gression of this disease and fail to consistently prolong
healthy lifespan (healthspan) of ALS paents beyond
several months. Thus, there is an urgent need for the
ethical scienc discovery, development, and delivery of
therapeucs that consistently extends the healthy lifes-
Amyotrophic Lateral Sclerosis (ALS) or Lou Gehrig’s dis-
ease is a neurodegenerative and neuromuscular disorder
characterized by a progressive death of motor neurons &
motor paralysis that culminates in death, usually within 3-5
years of diagnosis from respiratory failure due to paralysis.
Currently approved ALS therapies are not curative and fail
to increase healthy lifespan. The worldwide prevalence of
ALS is expected to increase by 69% over the next 25 years,
yet its etiology remains scientically unveried, complicat-
ing the discovery and development of effective therapies.
Through this cross-sectional clinical study employing micro-
biological and cellular analyses of fecal and blood samples
isolated from human patients with and without ALS, we
achieved to conrm recently emerging pre-clinical and clini-
cal evidence linking ALS with intestinal dysbiosis & system-
ic microbial infection, and thus provide rationale for future
scientic investigations of intestinal dysbiosis as a potential
therapeutic target for improving and extending the healthy
lifespan of human patients diagnosed with ALS.
Amyotrophic Lateral Sclerosis (ALS) or Lou Gehrig’s
disease is a neurodegenerave disorder characterized
Steenblock et al. Int J Neurodegener Dis 2018, 1:003 • Page 2 of 4 •
82 human paents included in this study, 54 (26 male
& 28 female) were diagnosed with ALS per the revised
El Escorial criteria, and 28 (14 male & 14 female) did
not have ALS or ALS-related symptoms and served as
a control group in this study. At the me of biological
(blood and feces) sample collecon & analyses, the
mean age of human paents with ALS (ALS Paents)
was 63.5 ± 12.3 years, and those without ALS (Non-ALS
Paents) was 59.4 ± 13.4 years.
Fecal & blood sample analyses
Concentraon of fecal secretory-IgA was determined
by an enzyme-linked immunosorbent assay (ELISA). Se-
rum white blood cell dierenal was determined by au-
tomated cell counng. Screening for a panel of 5 intes-
nal pathogenic bacteria was conducted per culture-based
microbiological methodology. LabCorp, Doctor’s Data,
and Great Palms Laboratory completed all afore men-
oned fecal & blood samples analyses per contemporary
best pracces in clinical laboratory science.
pan (healthspan) of ALS paents beyond what is current-
ly possible. The eology of ALS has not been fully elu-
cidated in the peer-reviewed scienc literature; how-
ever, evidence is emerging to indicate that disturbances
in the intesnal microbiota are involved in the cause &
progression of ALS and other neurodegenerave diseas-
es . Here, we provide clinical evidence linking intes-
nal dysbiosis & systemic microbial infecon with ALS
and discuss raonale for further scienc invesgaon
of this link and it’s potenal as a therapeuc target.
Materials and Methods
Data from the blood and fecal analyses of 82 human
paents were used in this retrospecve cross-seconal
study. The Steenblock Research Instute (SRI) & its Eth-
ics Commiee approved of this retrospecve cross-sec-
onal clinical study. All human paents were > 18 years
of age & were able to provide informed consent. Of the
F I G U RE 1 : FE C A L S E C R ET O R Y
I M M U NO G L O B UL I N A
A L S VS . N O N- A L S H U M A N P A T I EN T S
( C O N T R O L )
Figure 1: Mean concentration of fecal secretory-IgA is consistently & signicantly higher in ALS relative to Non-ALS Patients
(323.96 ± 30.73 mg/dL vs. 106.71 ± 5.56 mg/dL; P < 0.001).
FIGURE 2: PREVALENCE OF INTENSTINAL PATHOGENIC
BACTERIA IN 54 HUMAN PATIENTS WITH ALS
Figure 2: Fecal carriage of one or more bacterial species was detected in 100% of the 54 Human ALS Patients in this study.
Steenblock et al. Int J Neurodegener Dis 2018, 1:003 • Page 3 of 4 •
alence of pseudomonas aeruginosa and salmonella spp
is found to be 83.3% and 66.7%, respecvely (Figure 2),
thereby indicang an increased suscepbility in the ALS
Paents’ intesne to harbor pathogenic bacterial spe-
cies relave to that of healthy non-clinical populaons.
This study’s nding of intesnal dysbiosis as a feature in
ALS is in agreement with smaller cross-seconal clinical
studies conducted in 2016  and 2017 , both of
which evaluated the intesnal microbiome of ALS Pa-
ents through dierent methodologies to confer status
of intesnal dysbiosis.
This study’s analysis of serum white blood cell con-
centraon indicates that human ALS Paents have a
signicantly elevated neutrophil-to-lymphocyte rao
(Figure 3), which is an indicaon of systemic inamma-
on in response to systemic microbial infecon [16,17].
Intesnal dysbiosis has been shown to cause intesnal
barrier dysfuncon or intesnal epithelial hyperper-
meability in a mouse model of ALS [18,19], thereby
providing an anatomical pathway for the migraon of
pathogenic intesnal microbes and/or their metabol-
ic byproducts into the systemic circulaon via the gut
circulatory system to elicit systemic microbial infecon.
Given the co-occurrence of intesnal dysbiosis and sys-
temic microbial infecon in 100% of the ALS Paents
enrolled in this study, it is plausible that a similar physio-
logical phenomenon is occurring in human ALS Paents.
This study’s nding of systemic microbial infecon as a
feature of ALS Paents is consistent with that of a pre-
vious smaller study reporng the presence of serum
pathogenic microbial byproducts & systemic inamma-
on in human ALS Paents .
In a recent pre-clinical study ulizing a mouse mod-
el of ALS, it was demonstrated that intesnal dysbio-
sis precedes the development and progression of ALS
Data expression & analysis
Dierences between groups (ALS Paents vs. Non-
ALS Paents) were determined by ANOVA with Stu-
dent’s t-test. Data are expressed as mean ± standard
error of the mean (SEM). A P < 0.05 was considered as
Figure 1, Figure 2 and Figure 3.
Intesnal dysbiosis is widely dened as an abnormal
elevaon in the concentraon of pathogenic microbial
species that reside within the intesnal lumen and has
recently been associated with the eology and progres-
sion of many neurodegenerave diseases . Secretory
immunoglobulin A (SIgA), an endogenous anmicrobial
factor measurable in feces, is naturally found in the ex-
ternal secreons bathing mucosal surfaces of the intes-
ne at a concentraon that is posively proporonal to
the concentraon of pathogenic microbial species that
reside within the intesnal lumen . This cross-sec-
onal clinical study found that the mean concentraon
of fecal secretory immunoglobulin A (SIgA) is signicant-
ly higher (> 300%) in ALS Paents relave to Non-ALS
Paents (Figure 1), thereby indicang that ALS Paents
harbor a signicantly higher concentraon of patho-
genic microbial species in their intesnal lumen and are
thus in a state of intesnal dysbiosis. Furthermore, fecal
carriage prevalence of one or more pathogenic bacterial
species was conrmed in 100% of ALS Paents enrolled
in this study (Figure 2). Among healthy human popula-
ons, fecal carriage prevalence of the pathogenic bac-
teria pseudomonas aeruginosa and salmonella spp is
8.2% and 2-3%, respecvely [11-13]. However, in this
study’s populaon of ALS Paents, fecal carriage prev-
Concentraon (1x10^9 Cells Per uL)
FIGURE 3: SERUM WHITE BLOOD CELL DIFFERENTIAL
ALS VS. NON-ALS PATIENTS (CONTROL)
Figure 3: Mean serum concentration of Neutrophils (10.09 × 109 Cells vs. 2.78 × 109 Cells, P = 0.036), Lymphocytes (4.63 ×
109 Cells vs. 2.19 × 109 Cells, P = 0.015), and Eosinophils (1.95 × 109 Cells vs. 0.14 × 109 Cells, P = 0.023) are signicantly
higher in ALS relative to Non-ALS Human Patients.
Steenblock et al. Int J Neurodegener Dis 2018, 1:003 • Page 4 of 4 •
7. Mehta P, Antao V, Kaye W, Sanchez M, Williamson D, et al.
(2014) Prevalence of amyotrophic lateral sclerosis-United
States, 2010-2011. MMWR Suppl 63: 1-14.
8. Mehta P, Kaye W, Bryan L, Larson T, Copeland T, et al.
(2016) Prevalence of amyotrophic lateral sclerosis-United
States, 2012-2013. MMWR Suppl 65: 1-12.
9. Marizzoni M, Provasi S, Cattaneo A, Frisoni GB (2017) Mi-
crobiota and neurodegenerative diseases. Curr Opin Neu-
rol 30: 630-638.
10. Brandtzaeg P (2013) Secretory IgA: Designed for anti-mi-
crobial defense. Front Immunol 4: 222.
11. Estepa V, Rojo-Bezares B, Torres C, Sáenz Y (2014) Fae-
cal carriage of Pseudomonas aeruginosa in healthy hu-
mans: antimicrobial susceptibility and global genetic lineag-
es. FEMS Microbiol Ecol 89: 15-19.
12. L Garedew-Kifelew, N Wondafrash, A Feleke (2014) Identi-
cation of drug-resistant Salmonella from food handlers at
the University of Gondar, Ethiopia. BMC Res Notes 7: 545.
13. B Abera, G Yitayew, H Amare (2016) Salmonella seroty-
petyphi, shigella, and intestinal parasites among food han-
dlers at bahir dar university, Ethiopia. J Infect Dev Ctries
14. Fang X, X Wang, S Yang, F Meng, X Wang,
et al. (2016) Evaluation of the microbial diversity in amy-
otrophic lateral sclerosis using high throughput sequenc-
ing. Front Microbiol 7: 1479.
15. Rowin J, Xia Y, Jung B, Sun J (2017) Gut inammation and
dysbiosis in human motor neuron disease. Physiol Rep 5.
16. Naess A, Nilssen SS, Mo R, Eide GE, Sjursen H (2017)
Role of neutrophil to lymphocyte and monocyte to lympho-
cyte ratios in the diagnosis of bacterial infection in patients
with fever. Infection 45: 299-307.
17. Lowsby R, Gomes C, Jarman I, Lisboa P, Nee PA, et
al. (2015) Neutrophil to lymphocyte count ratio as an early
indicator of blood stream infection in the emergency depart-
ment. Emerg Med J 32: 531-534.
18. Wu S, J Yi, YG Zhang, J Zhou, J Sun (2015) Leaky intestine
and impaired microbiome in an amyotrophic lateral sclero-
sis mouse model. Physiol Rep 3.
19. Zhang YG, S Wu, J Yi, Y Xia, D Jin, et al. (2017) Target In-
testinal microbiota to alleviate disease progression in amy-
otrophic lateral sclerosis. Clin Ther 39: 322-336.
20. Zhang R, RG Miller, R Gascon, S Champion, J Katz, et al.
(2009) Circulating endotoxin and systemic immune acti-
vation in sporadic amyotrophic lateral sclerosis (sALS). J
Neuroimmunol 206: 121-124.
symptoms, and that reversal of intesnal dysbiosis with
a dietary probioc intervenon alleviates ALS symptoms
and progression, reverses intesnal barrier dysfuncon,
and prolongs healthy lifespan . This strongly indi-
cates that intesnal dysbiosis may be a modiable cause
of ALS. Through this cross-seconal clinical study, we
achieved to conrm recently emerging pre-clinical and
clinical evidence linking ALS with intesnal dysbiosis &
systemic microbial infecon, and thus provide raonale
for future scienc invesgaon of intesnal dysbiosis as
a potenal therapeuc target for extending the healthy
lifespan of human paents diagnosed with ALS.
T.I. and D.A.S. performed data collecon & paent
examinaons. T.I. performed data analysis, prepared
the gures, & wrote the manuscript. M.J.A composed &
revised the manuscript per reviewer feedback. A.S.S.A.
& E.W. revised the manuscript.
Conﬂict of Interest Statement
The authors declare that the research was conduct-
ed in the absence of any commercial or nancial rela-
onships that could be construed as a potenal conict
of interest. Authors claim no conicts of interest related
to the research described in the manuscript.
1. Matthew C Kiernan, Steve Vucic, Benjamin C Cheah, Turn-
er MR, Eisen A, et al. (2011) Amyotrophic lateral sclerosis.
The Lancet 377: 942-955.
2. Steenblock D (2016) Hypothesis: Osseous spinal injury and
reinjury as a risk factor, biomarker, and etiological factor in
sporadic ALS. Townsend Letter 1-2.
3. Ikrar T (2017) Amyotrophic lateral sclerosis: New sugges-
tions of pathophysiology and treatments. Prog Comm Sci-
ence 3: 9-18.
4. Chio A, Logroscino G, Traynor BJ, Collins J, Simeone
JC, et al. (2013) Global epidemiology of amyotrophic lat-
eral sclerosis: A systematic review of the published litera-
ture. Neuroepidemiology 41: 118-130.
5. Arthur KC, Calvo A, Price TR, Geiger JT, Chio A, et al.
(2016) Projected increase in amyotrophic lateral sclerosis
from 2015 to 2040. Nature Communications 7: 12408.
6. Mitsumoto H, David A Chad, Pioro EP (1998) Amyotrophic
lateral sclerosis. FA Davis Co.