# Full Text: EhrlichialInfection

> Extracted from `2015_EhrlichialInfection.pdf`

---

## Page 1

Could ehrlichial infection cause some of the changes associated
with leukemia, myelodysplastic diseases and autoimmune disorders,
and offer antibiotic treatment options?
Charles A. Kallick a,⇑, Daniel A. Friedman b, Mramba B.A. Nyindo c
a Rush University Medical Center, Chicago, IL, USA
b Department of Biology, Stanford University, Palo Alto, CA, USA
c Kilimanjaro Christian Medical University College, Moshi, Tanzania
a r t i c l e
i n f o
Article history:
Received 2 July 2014
Accepted 12 September 2015
a b s t r a c t
We hypothesize that a large group of medical conditions of unknown etiology including leukemia, mul-
tiple myeloma, myelodysplastic and autoimmune disorders, may be associated with or caused by an
obscure group of intracellular obligate parasitic bacteria named Ehrlichia/Anaplasma (EA). Ensconced
in the stem cells of the bone marrow, EA may disrupt the normal development and function of many
of the cells of immunity, manifesting itself as different syndromes. Recent studies of the activity of EA
suggest direct effects on the immune system consistent with the manifestations of leukemia. We refer-
ence here three leukemia patients with direct or indirect evidence of EA infection. Moreover, EA have
been shown to be most sensitive to rifamycins. Two moribund leukemia patients with levels of platelets
and white cells incompatible with life were treated with therapeutic doses of Rifampin. Though they did
not survive, their condition improved dramatically for a time, suggesting Rifampin provided some ther-
apeutic beneﬁt. We assert that these results warrant more extensive study.
 2015 Published by Elsevier Ltd.
Hypothesis: ehrlichial infection and leukemia
Diseases of the immune system broadly described by the term
leukemia include acute lymphoblastic leukemia (ALL), acute mye-
loid leukemia (AML), chronic lymphocytic leukemia (CLL) and
chronic myeloid leukemia (CML). The causes of these leukemic
syndromes are unknown though many genetic changes have been
associated with some forms.
In leukemia we observe the overproduction of cells needed for
immune system function, accompanied by large numbers of imma-
ture and dysfunctional cells of immunity (termed blasts) that are
inappropriately released into the circulation. Something unknown
is causing these blasts to fail to function as expected and accumu-
late in the system of the patient.
The Ehrlichia/Anaplasma (EA) are a family of obligate intracel-
lular parasitic bacteria that infect leukocytes. They have been rec-
ognized as human pathogens for a variety of medical conditions
[1,2]. EA can alter the DNA of their host cell during its division,
as discussed below. Interference with the normal progression of
marrow cell development may facilitate the survival of the bacteria
in their host leukocytes, by suppressing apoptosis and could also
cause a cascade of subsequent immune system failures.
The EA are a Chlamydia, which have different reproductive
methods than many other invasive bacterial pathogens. A study
of Ehrlichia Chaffeensis infection in a human monocyte cell line
demonstrated the ability of EA to alter host genes during transcrip-
tion (transcriptomic effects) [3]. These effects included suppression
of apoptosis, a primary defensive activity of intracellular pathogens
regulating cell differentiation, and others essential for survival of
the obligatory intracellular parasite. A culture of Anaplasma phago-
cytophilum was induced to grow in human immune system cells
and produced most of the changes seen in leukemia [4].
The ﬁrst reported association of EA with immune system dys-
function dates back to 1973, in a patient with aplastic anemia
[5]. Identiﬁcation of the EA infection required an unusual culture
method, as these obligate parasites do not grow in normal micro-
biologic media [6]. Subsequent reports also link EA with systemic
lupus erythematosus (SLE) and myelodysplastic syndromes [7].
Could immune system derangement by parasitic EA link all these
syndromes and leukemia as well?
Besides leukemia, the diseases of immunity also include
myelodysplastic disease, multiple myeloma, and aplastic anemia.
All of these disease states, which cause dysfunction of the immune
system,
are
known to
spontaneously
metamorphose
to
the
http://dx.doi.org/10.1016/j.mehy.2015.09.015
0306-9877/ 2015 Published by Elsevier Ltd.
⇑Corresponding author.
E-mail addresses: Cak0529@aol.com (C.A. Kallick), Danielarifriedman@gmail.
com (D.A. Friedman), Mnyindo2002@yahoo.co.uk (M.B.A. Nyindo).
Medical Hypotheses 85 (2015) 891–893
Contents lists available at ScienceDirect
Medical Hypotheses
journal homepage: www.elsevier.com/locate/mehy

## Page 2

leukemic state. This indicates that they may be related to a similar
or singular infectious agent. Since EA are known to alter the func-
tion of immune system cells and have been implicated in many of
these diseases, this begs the question of whether EA may be the
culprit.
Assuming leukemia is associated with an EA infection of the
leukocytes, it would further be expected that treatments impacting
the bone marrow would be palliative. This in fact is the case; stan-
dard treatment for leukemia includes a large list of substances
related to the cells of immunity [8] as well as full bone marrow
replacement [9] and often induces a diminution of symptoms for
some time, though complete remissions are often rare.
It is further hypothesized, moreover, that treatment of leukemia
with antibiotics effective against EA would also result in beneﬁcial
impact. This has been tried. The results, cited below [10], hint at
proof of EA infection as a cause of leukemia as well as a potentially
important course of treatment.
Three illustrative cases
Two of three patients discussed here either presented with leu-
kemia; the other suffered from polycythemia vera, which some-
times progresses to leukemia. Two were treated with Rifampin
when all other treatments had been exhausted and the patients
were moribund and near death. Though both eventually expired,
the dramatic and unexpected improvement in their condition after
treatment with Rifampin (an available rifamycin) suggests that EA
may have been impacted by the antibiotic. Earlier intervention
may have been worthwhile. In the third case, PCR analysis indi-
cated the presence of a previously unknown EA. Taken together
these three cases, though not deﬁnitive, provide tantalizing evi-
dence of a connection between leukemia and EA [10].
Case number 1
Patient AM was a 64 year old man, who ﬁrst presented with
acute lymphocytic leukemia (ALL). The standard treatment, bone
marrow replacement, caused the patient to enter remission for a
period of approximately 2 years. Then the leukemia recurred. The
patient was sent to home care, being thrombocytopenic and with
leukocyte counts below those needed to counter infection. After
requesting and receiving treatment with Rifampin, he experienced
a rapid increase in platelets and leukocytes, coupled with a dra-
matic recovery of physical well-being, which lasted for several
months. Eventually he developed hearing loss and a rash resem-
bling that seen in cytomegalovirus infection and died suddenly,
of unknown causes [10].
Case number 2
MR was a 14 year old male who developed acute myelogenous
leukemia (AML). After the early failure of conventional chemother-
apy, he developed thrombocytopenia and leukopenia. His white
count decreased to less than 200 for 2 months, at which time his
mother requested therapeutic doses of Rifampin. Within a short
period of treatment, his leukocytes rose from 200 to 24,000, of
which 25% were mature. His platelets, however, rose considerably
slower. Due to a falling hematocrit, MR was prescribed blood
replacement, but he died of blood loss during transfer to a hospital
from hospice care [10].
Case number 3
This woman in her 50 s suffered from polycythemia vera, a
myelodysplastic disease, which occasionally progresses to leuke-
mia. The patient suddenly developed an acute, rapidly fatal AML.
A blood ﬁlm of her peripheral blood showed unexpected inclusions
in her platelets, white cells and red cells. These were morphologi-
cally indistinguishable from similar structures found in blood from
EA infections. Because she had been splenectomized, her red cells
were apparently infected at an observed 2% level. Because of the
presumed level of bacteria in her blood, a PCR analysis was done,
and showed evidence of a previously unreported Ehrlichia. The
details of that PCR are available from the original publication [10].
The rapidly evolving science of study of the EA
The presentation of the factors needed in the study of the rela-
tionship of the EA to leukemia includes knowledge of several disci-
plines, including oncology, infectious disease, immunology and
genetics. In Appendix A, we discuss EA-induced changes in the
transcriptome and epigenome of human cells, as well as the known
molecular similarities between the proliferative states of leukemia
and EA infection.
In order for these reports to be understood and become a part of
this report, and because these important descriptions of laboratory
investigation are unfamiliar to most clinicians engaged in the care
of these patients, we included a detailed but carefully presented
discussion presented as an Appendix A, Addendum Vida infra.
Consequences of the hypothesis
These data do not prove that these apparently unrelated syn-
dromes are caused by the same or similar microorganism, but
are so suggestive that much more study of serology and DNA must
be done to see if there are enough cases to declare that there is a
connection or not. Rifampin is now used for control of Ehrlichia
and other pathogens and could easily be the subject of further
investigations of its effect on leukemia [11]. The accumulating evi-
dence discussed above gives hope for a signiﬁcant change in mor-
tality rates from the scourge of leukemia, as well as other
autoimmune syndromes. Indeed, application of the antibiotics
therapy could be used in a compassionate manner for any leuke-
mia cases that are found to be untreatable with conventional ther-
apy and almost certain to eventually result in death. Any small risk
of harm is outweighed by the possibility of help.
Conclusions
Perhaps the mystery of leukemia can be explained as an attack
by an unsuspected, hard to detect pathogen, evading destruction
by the immune system by infecting, disabling and eliminating
the cells and mechanisms of immunity. This is not a new phe-
nomenon, as we know from AIDS and other diseases. The resulting
consequence, a ﬂood of dysfunctional immune system cells, pre-
sents as the disorder we call leukemia (or in other cases aplastic
anemia, polycythemia vera, lupus and similar autoimmune dis-
eases). Leukemia, under this conceptualization, may not be a
malignancy at all, but rather an infection that can be treated with
antibiotics and other means. Lacking any better explanation for
this terrible disease, nor any reliable curative treatments, the pos-
sibility that leukemia is caused by or associated with EA or similar
infection cries out for more extensive investigation than has taken
place to date.
Conﬂicts of interest
The authors Friedman and Nyindo have no conﬂict of interest.
Author C.A. Kallick has one patent which may have a conﬂict of
interest with some of the information in the paper. Author Kallick
has transferred that patent to a partnership composed of his chil-
dren. Author Kallick has retained only 1% of the possible value of
this patent.
892
C.A. Kallick et al. / Medical Hypotheses 85 (2015) 891–893

## Page 3

Acknowledgements
The authors express gratitude for the following work and assis-
tance; Steven Kallick, Marlyn Twitchell for manuscript and other
assistance. To Doctors Mark Lepper, Midrag Ristic, Carl Woese, Stu-
art Levin and Kathryn Iacono for knowledge, advice, and scientiﬁc
direction.
Appendix A
Transcriptomic comparison of a human monocyte cell line
(THP1) with or without an E. chaffeensis (EC) infection elucidated
genes that were differentially expressed between the infected
and uninfected states. EC infection induces changes in several cel-
lular systems of the monocyte, as evidenced by enrichment of
differentially-expressed genes in categories relating to differentia-
tion, apoptosis, and signal transduction. These transcriptomic
changes are consistent with a model in which EC inhibits key sig-
naling pathways of human cells, impairing the host cell’s internal
defensive capabilities and apoptotic potential. This is the predicted
high-ﬁtness transcriptomic strategy that a co-evolved intracellular
parasite would evolve towards – decrease the ability of the host
cell to respond to infection, prevent the host cell from undergoing
apoptosis, and encourage the host cell to proliferate at a high rate.
ETV6 is an oncogenic transcription factor involved in hemato-
poiesis, and mutations in ETV6 are strongly associated with leuke-
mia in children [13]. ETV5 is implicated in malignant cancers [14],
and in stem cell proliferation [15]. ETV5 and ETV6 expression
levels are altered by EC [12].
BCL proteins and other apoptosis inhibitors are common major
driver mutations in acute leukemia at all ages, as gain-of-function
mutations in apoptosis inhibitor genes allows a cell to escape inter-
nal and/or external regulators of the cell cycle [13]. EC infection in
monocytes upregulates the production of several known apoptosis
inhibitors, such as BCL2A1, NF-B, BIRC3, IER3, and MCL1 [12].
Akagi et al. [15] use 771 transcriptomic samples of AML to ﬁnd
networks of genes that are differentially utilized in AML. They ﬁnd
that CDK1, CDK2, CDK4, and CDK6 are the most dysregulated cen-
tral agents in the cell division gene regulatory network, and
encourage that these CDKs are the most promising drug available
for treatment targets. EC infection of monocytes downregulated
CDC2, CDK5, CDK8, and cyclin G1.
Taken together, these mechanistic studies suggest that EC may
hijack cellular signaling pathways relating to hematopoietic differ-
entiation, oncogenic malignancy, cell cycle, and pluripotency to
promote similar proliferative changes in blood cell phenotype as
some leukemias.
Immunohistochemical evidence supports the translocation of
the E. chaffeensis ankyrin repeat-containing protein p200 into the
nucleus of human monocytes during infection [16]. A similar ﬁnd-
ing has been reported for the translocation of the EC protein
TRP120 [17] and another ankyrin repeat-containing protein from
A. phagocytophilum into the nucleus of human granulocytes [18].
EC p200 proteins are the primary immunoreactive factors in infec-
tions of mammalian cells [19], and are involved in cell differentia-
tion decisions and host responses to infection. In the nucleus of
infected human cells, chromatin immunoprecipitation followed
by microarray quantiﬁcation (ChIP-chip) against EC p200 revealed
that p200 directly interacts with the promoter elements of 200
genes in the human genome. Genes that p200 bound to, and may
regulate, were enriched for Gene Ontology (GO) categories of tran-
scriptional regulation and apoptosis [19].
Yeast-two-hybrid screens revealed that EC proteins TRP47,
TRP120, and TRP32 physically interact with human proteins [20].
EC proteins interacted with several human genes known to play
roles in transcriptional regulation, cell differentiation, apoptosis,
and immune response to infection [20]. This evidence adds to a
body of literature showing that EC is able to use a variety of mech-
anisms to reprogram host cells towards uncontrolled proliferation
[21].
References
[1] Kallick CA, Levin S, Reddi KT, Landau WL. Systemic Lupus Erythematosus
associated
with
a
Haemobartonella-like
organism.
Nat
New
Biol
1972;236:145–6.
[2] Anaplasmosis and Ehrlichiosis. Morbidity and Mortality Weekly Report, vol. 58
No. 37; 2008 September 25. p. 1033–5.
[3] Zhang JZ, Luxon SM, Yu XJ. Survival strategies of obligately intracellular
Ehrlichia chaffeensis: novel modulation of immune response and host cell
cycles. Infect Immun 2004;72:4980507.
[4] Sukumaran B, Carluon JA, Berliner N, Fikrig E. Early transcriptomic response of
human neutrophils to Anaplasma phagocytophilum infection. Infect Immun
2005;73:8089–99.
[5] Kallick CA, Levin S, Reddi KT. Association of a Rickettsial agent identiﬁed in
human bone marrow failure with Ehrlichia canis and Tropical Canine
pancytopenia.
In:
Presented
at
the
13th
interscience
conference
on
antimicrobial agents and chemotherapy. Washington DC; 1973.
[6] Nyindo MBA, Ristic M, Huxholl DL, Smith AR. Tropical canine pancytopenia:
in vitro cultivation of the causative agent – Ehrlichia canis. Am J Vet Res
1971;32:1651–8.
[7] Kallick CA. Myelodysplastic diseases and Ehrlichia: consideration of a possible
etiologic connection and mechanisms of pathogenesis. In: 12th annual
symposium on myelodysplastic syndromes. (Abstract #238) Berlin; 2013.
[8] Wetzler M, Byrd JC, Bloomﬁeld CD. Acute and chronic leukemia in Harrison’s
principle of internal medicine. 15th Ed. McGraw-Hill; 2001. p. 706–27.
[9] Paveltic
S,
Khouri
IF,
Haagensen
M,
et
al.
Unrelated
bone
marrow
transplantation
for
B
cell
chronic
lymphocytic
leukemia
after
using
myeloablative conditioning Results from the Center for International Blood
and Bone marrow Transplant. J Clin Oncol 2005;23(24):5788–94.
[10] Kallick CA. Ehrlichia and bone marrow cells: could Ehrlichial infection explain
the unsuspected etiology of some diseases of the immune system? J Med
Hypotheses 2011;77:375–9.
[11] Klein MB, Nelson M, Goodman JL. Antibiotic sensitivity of the newly cultivated
agent of human granulocytic Ehrlichiosis. Antimicrob Agents Chemother
1997;41:76–9.
[12] Zhang J, Sinha M, Luxon BA, Yu X. Survival strategy of obligately intracellular
Ehrlichia Chaffeensis: novel modulation of immune response and host cell
cycles. Infect Immun 2004;72:498–507.
[13] Chiaretti S, Gianfelici V, Ceglie G, Foà R. Genomic characterization of acute
leukemias. Med Princ Pract 2014;23:487–506.
[14] Pedrola N, Devis L, Llauradó M, Campoy I, Martinez-Garcia E, Garcia M, et al.
Nidogen 1 and nuclear protein 1: novel targets of ETV5 transcription factor
involved in endometrial cancer invasion. Clin Exp Metastasis 2015;32:467–78.
[15] Akagi T, Kuure S, Uranishi K, Koide H, Costantini F, Yokota T. ETS-related
transcription factors ETV4 and ETV5 are involved in proliferation and
induction of differentiation-associated genes in ES cells. J Biol Chem 2015.
[16] Zhu B, Nethery KA, Kuriakose JA, Wakeel A, Zhang X, McBride JW. Nuclear
translocated ehrlichia chaffeensis ankyrin protein interacts with a speciﬁc
adenine-rich motif of host promoter and intronic Alu elements. Infect Immun
2009;77:4243–55.
[17] Zhu B, Kuriakose JA, Luo T, Ballesteros E, Gupta S, Fofanov Y, et al. Ehrlichia
chaffeensis TRP120 binds a G+C-rich motif in host cell DNA and exhibits
eukaryotic transcriptional activator function. Infect Immun 2011;79:4370–81.
[18] Park J, Kim KJ, Choi K, Grab DJ, Dumler JS. Anaplasma phagocytophilium AnkA
binds to granulocyte DNA and nuclear proteins. Cell Microbiol 2004;6:743–51.
[19] McBride JW, Comer JE, Walker DH. Novel immunoreactive glycoprotein
orthologs of Ehrlichia spp. Ann N Y Acad Sci 2003;990:678–84.
[20] Luo T, McBride JW. Ehrlichia chaffeensis TRP32 interacts with host cell targets
that inﬂuence intracellular survival. Infect Immun 2012;80:2297–306.
[21] Wakeel A, Zhu B, Yu X-J, McBride JW. New insights into molecular Ehrlichia
Chaffeensis-host interactions. Microbes Infect 2010;12:337–45.
C.A. Kallick et al. / Medical Hypotheses 85 (2015) 891–893
893


---
*Extraction method: pymupdf*