F1000Research 2019, 8(F1000 Faculty Rev):589 Last updated: 30 APR 2019

REVIEW
Combating Cholera [version 1; peer review: 2 approved]
Brian Y. Hsueh , Christopher M. Waters
Microbiology and Molecular Genetics, Michigan State University, East Lansing, MI, 48824, USA

First published: 30 Apr 2019, 8(F1000 Faculty Rev):589 ( Open Peer Review
v1 https://doi.org/10.12688/f1000research.18093.1)
Latest published: 30 Apr 2019, 8(F1000 Faculty Rev):589 (
https://doi.org/10.12688/f1000research.18093.1)
Referee Status:

Abstract Invited Referees

Cholera infections caused by the gamma-proteobacterium Vibrio cholerae have 1 2
ravaged human populations for centuries, and cholera pandemics have
afflicted every corner of the globe. Fortunately, interventions such as oral version 1
rehydration therapy, antibiotics/antimicrobials, and vaccines have saved published
countless people afflicted with cholera, and new interventions such as 30 Apr 2019
probiotics and phage therapy are being developed as promising approaches to
treat even more cholera infections. Although current therapies are mostly
effective and can reduce disease transmission, cholera outbreaks remain F1000 Faculty Reviews are commissioned
deadly, as was seen during recent outbreaks in Haiti, Ethiopia, and Yemen. from members of the prestigious F1000
This is due to significant underlying political and socioeconomic complications, Faculty. In order to make these reviews as
including shortages of vaccines and clean food and water and a lack of health
comprehensive and accessible as possible,
surveillance. In this review, we highlight the strengths and weaknesses of
current cholera therapies, discuss emerging technologies, and argue that a peer review takes place before publication; the
multi-pronged, flexible approach is needed to continue to reduce the worldwide referees are listed below, but their reports are
burden of cholera. not formally published.

Keywords
cholera, Vibrio cholera, antibiotics, oral-rehydration therapy, vaccine, phage 1 Jyl Matson, University of Toledo College
therapy, probiotics of Medicine and Life Sciences, USA

2 Jeffrey Withey, Wayne State University

School of Medicine, USA

Any comments on the article can be found at

the end of the article.

Corresponding author: Christopher M. Waters ([email protected])

Author roles: Hsueh BY: Conceptualization, Writing – Original Draft Preparation, Writing – Review & Editing; Waters CM: Conceptualization,
Funding Acquisition, Writing – Original Draft Preparation, Writing – Review & Editing
Competing interests: No competing interests were disclosed.
Grant information: This work was supported by National Institutes of Health grants GM109259 and AI130554 and National Science Foundation
grant MCB1714612 to CMW.
The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Copyright: © 2019 Hsueh BY and Waters CM. This is an open access article distributed under the terms of the Creative Commons Attribution
Licence, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
How to cite this article: Hsueh BY and Waters CM. Combating Cholera [version 1; peer review: 2 approved] F1000Research 2019, 8(F1000
Faculty Rev):589 (https://doi.org/10.12688/f1000research.18093.1)
First published: 30 Apr 2019, 8(F1000 Faculty Rev):589 (https://doi.org/10.12688/f1000research.18093.1)

Page 1 of 8
 F1000Research 2019, 8(F1000 Faculty Rev):589 Last updated: 30 APR 2019

Introduction current epidemics, and O1 V. cholerae is the major infectious

For centuries, cholera has wreaked havoc on developing agent2. Overall, owing to a lack of TCP or CT, non-O1/non-
countries with poor infrastructure, sanitation, and access to O139 biotypes do not cause cholera, but there are cases where
clean drinking water. Cholera also flourishes when normal they do instigate diarrheal symptoms6. The O1 and O139 strains
societal function is disrupted, such as during natural disasters are prevalent in several endemic regions, including Yemen,
like the 2010 earthquake in Haiti1 or the current refugee crisis parts of Africa, Southeast Asia, and Haiti2,7–12. Serogroups are
in Yemen2. Vibrio cholerae, the etiological agent of cholera, is a subclassified into two major biotypes. The first six pandemics
Gram-negative, rod-shaped pathogen that can cycle between of V. cholerae from the years 1817 to 1921 were caused
two distinct environments—persistence in brackish-water ponds by the classical biotype, whereas the seventh and current
and infection of the human gut—and it transmits from the pandemic that started in 1961 was caused by the El Tor
environmental reservoirs to the human host via contami- biotype13. Virtually all modern-day cholera is caused by El Tor,
nated food or water. V. cholerae is highly sensitive to the and environmental sampling identifies only El Tor, suggesting
low pH of the stomach and thus the infectious dose for this that classical biotypes are no longer prevalent. The El Tor
bacterium is high at greater than 108 organisms3. Those cells biotype can be grouped into the serotypes Ogawa and Inaba,
that survive the stomach acid eventually colonize the intestinal which are the most prevalent serotypes that are causing the
tract. The toxin co-regulated pilus (TCP) aids in coloniza- pandemics2,14,15, and these are used in contemporary vaccines
tion by promoting bacterial microcolony formation. V. cholerae like Shanchol and Euvichol16. Even with modern-day
then secretes cholera toxin (CT), which disrupts normal ion treatments, it is estimated that there are over 3 million cases of
transport of the gut epithelium, inducing the massive water cholera with more than 100,000 deaths annually16–18. The World
efflux into the intestine which leads to debilitating diarrhea Health Organization (WHO) public database of annual epidemic
and vomiting4. Transitioning between biofilm formation and cholera cases provides outbreak updates and a summary of
motility during infection is also a key component of V. cholerae worldwide infections19.
colonization5.
The objective of this review is to describe the current strategies
Out of more than 200 serogroups of V. cholerae, only the of oral rehydration therapy (ORT), antibiotics, and vaccination
serogroups O1 and O139 have been the causative agent of which are used to treat and prevent cholera. (See Figure 1 for

Figure 1. The five strategies to treat cholera. This diagram summarizes the strengths and weaknesses of five different cholera treatments
discussed in this review. By considering the strengths and weaknesses of the current therapies and leveraging the diversity of resources
and new technologies, a multi-pronged approach could well improve the chances of success in combating cholera infections worldwide and
potentially establish cost-effective, pre-emptive solutions more quickly than conventional methods of treatment.

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an overview.) We also highlight novel emerging approaches treat cholera and could supplant glucose-based ORT as the major
to treat and prevent cholera—such as probiotic treatment and treatment for cholera.
phage therapy—that have shown success in laboratory conditions
but are not yet used in human populations. Because cholera Although modifying the ORT ingredients to achieve optimal
outbreaks often are linked to poor infrastructure, lack of Na+ and water absorption in the intestines reduces symptoms,
access to clean water, or societal disruptions, our view is that a there remain cases where ORT could not keep the cholera
multi-pronged, flexible strategy is needed to combat these symptoms in check. For instance, severe dehydration requires
infections, and each of these treatment strategies can meet a intravenous rehydration. Because ORT is not 100% effective,
specific need to reduce the burden of cholera (Figure 1). concurrent treatments such as antibiotics/antimicrobials23–25
and vaccination26 may be necessary to sustain the reductions in
Oral rehydration therapy cholera symptoms, as will be discussed in the next two sections.
ORT has a long and interesting history in the field of medicine
as a therapeutic to treat acute diarrheal infection. Based on prior Antibiotics/Antimicrobials
knowledge that glucose was essential to facilitate absorption of The objective of antibiotics to treat cholera infections is to
water from the gut7,20, the idea of ORT was first attempted in 1964, reduce both (1) the time and severity of the illness and (2) the
when US Navy Capt. Robert Phillips used oral glucose saline to transmission to other individuals. Acute infection with severe
successfully treat cholera in two patients in the Philippines21. dehydration is treated with ORT and antibiotics to produce
ORT has since become the most widely used quintessential synergistic efficacy10–12,16. Effective antibiotics to treat cholera
cholera treatment. Prior to ORT, cholera infections had a are doxycycline, azithromycin, and tetracycline. Administration
mortality rate of more than 50%. However, ORT has treated of multiple doses of 12.5 mg tetracycline for 3 days can
infection in millions of individuals and saved millions of lives reduce the duration of symptoms in adults from 4 to 2 days
by replacing lost fluids and electrolytes during infection7,20. and average stool volume from 21 L to 8 L23. A single dose of
This treatment strategy relies on the fact that cholera is a self- doxycycline (300 mg for adults and 6 mg for children) is as
limiting infection. Thus, if the patient can survive the massive effective as multiple doses of tetracycline2,24. On the other
fluid loss elicited by CT, the infection ultimately will resolve hand, an analysis of significantly more trials with indirect
within a few days. ORT has reduced the mortality rate of comparisons of tetracycline to doxycycline found that patients
cholera by more than 97%, and more than 99% of patients on who received tetracycline had a shorter duration of diarrhea
ORT survive V. cholerae infections14,22. Because V. cholerae (by 1 day) while the stool volume reduction was significantly
infections cause the intestinal epithelial cells to lose copious higher17. Likewise, a single dose of 20 mg azithromycin can
amounts of essential electrolytes, conventional ORT prescribed stop diarrheal symptoms in less than 48 hours—24 hours earlier
by the WHO contains several vital ions (sodium, chloride, and than with ciprofloxacin25—and decrease vomiting frequency
potassium) and a carbon source (glucose). while allowing passage of an average of 36 stools with volumes
averaging about 5 L2,24. Azithromycin is recommended for
Though effective, the constituents of ORT have been studied pregnant women and young children whereas tetracycline is
and modified since its inception. Glucose, one of the com- suitable for adults. They are both more advantageous than
ponents of ORT, can increase the production of CT, the main ciprofloxacin and erythromycin1,11,12.
cause of the severe symptoms associated with the disease7.
Kühn et al. established that a rice- or starch-based ORT would One drawback to antibiotic therapy is that V. cholerae O1 and
circumvent this dilemma7. Glucose stimulates Na+ absorption O139 strains have developed resistance to most of the antibi-
faster than rice starch in the small intestine, so it was possible otics that are used. For example, ciprofloxacin, a type of fluo-
that a glucose-based ORT would be shorter and more cost- roquinolone that was commonly used in the early 1990s because
effective. However, even with the slower Na+ absorption rate, the of its long half-life and high in vitro activity, was ineffective in
rice-based ORT reduced stool volumes by 36% compared with multiple countries with a high burden of cholera infection, such
glucose ORT23. Recently, the rice-based ORT was successfully as Haiti and Bangladesh24,27. This is because O1 and O139 are
field-tested to treat cholera in Haiti7. Concomitantly, starch- also resistant to nalidixic acid, which has a mechanism similar
based ORT is resistant to metabolic degradation in the small to that of ciprofloxacin, and this mechanism confers cross-
intestine, persisting longer than glucose20,23, and it does not resistance to ciprofloxacin24,27. Strains resistant to tetracycline
significantly induce CT production7. A starch-based ORT has were isolated in several developing countries like Bangladesh,
the additional benefit of stimulating the synthesis of short-chain India, Thailand, and Northern Vietnam2,14,25, and based on
fatty acids (SCFAs) that can lessen the occurrences of diarrhea sequencing analysis, the resistance to tetracycline is plasmid-
by activating the retention of ions by colonic epithelial cells. mediated, suggesting that it could continue to rapidly spread
These SCFAs are produced from starch through fermentation in V. cholerae populations25,27. To avoid the development of resist-
of carbohydrates which are not rapidly absorbed or degraded in ance to these agents, Khan et al. recommended taking these
the small intestine by the colonic microbiome20. Moreover, medications only when the resistant strains are not prevalent27.
since glucose increases ion absorption in the small intestine, Furthermore, V. cholerae is evolving new genetic mechanisms
these two additives could have a synergistic effect at lessening to confer resistance to these drugs. Models that predict the
cholera symptoms20. Though not yet endorsed by the WHO, emergence of new pandemic strains in heavy-burden, developing
these alternative carbon-based ORTs exhibit potential benefits to countries may be useful for planning future antibiotic treatment

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strategies, including proper drug allocation, and for elucidating the and Drug Administration–approved, single-dose vaccine that
epidemiology of drug-resistant outbreak strains10. protects against either the Inaba or Ogawa serotype and con-
tains CTB from both classical and El Tor biotype2,30,33. Owing
Although these classes of antibiotics can achieve positive to the robust, rapid cell-mediated protection of Vaxchora, its
therapeutic effects, it is important to consider the adverse side efficacy against cholera is estimated to be around 90% post-
effects of these treatments. Hypersensitivity reactions are the vaccination and 80% 3 months post-vaccination in travel-
most common life-threatening side effect of antibiotic treatment ers to high-risk cholera areas2,30,33. The next steps for Vaxchora
of cholera, whereas the irregular cardiac rhythm condition is development are to evaluate its safety and effectiveness in
common only in ciprofloxacin and azithromycin24. ORT and cholera-endemic populations and to optimize the preparation
antibiotic therapy function to treat cholera infections but do not of the vaccine since it relies on cold-chain shipping and water
prevent patients from acquiring cholera. Therefore, the next mixing, which are problematic for distribution in some endemic or
intervention that we will discuss that has the potential to limit disrupted regions33.
cholera infections in susceptible populations is vaccines.
Several alternative forms of vaccines are being developed and
Vaccines these include a combined outer membrane vesicle (OMV)
The WHO advocates the use of oral cholera vaccines (OCVs), vaccine against V. cholerae and Escherichia coli that has been
including both live-attenuated and inactivated oral whole cell shown to induce a strong immunogenic response12, a genetically
(WC) vaccines, in endemic areas or during outbreaks as a manipulated form of live V. cholerae without the diarrhea-
transient protection since they have been shown to be effec- genic factors to mediate probiotic-like protection from cholera34,
tive in combination with other correlative treatments, including and antimicrobial glycoconjugates, particularly the lipopoly-
antibiotics, ORT, and health management17,28. OCVs princi- saccharide epitopes across different serotypes (Ogawa and
pally stimulate mucosal immunity mediated by antibodies, Inaba)35. These alternative forms of vaccines are not yet in
particularly IgA, against the pathogen. These antibodies are clinical trials.
directed against antigens such as O1-specific polysaccha-
ride and CT28. Although IgA has limited systemic circulation Effectively using vaccines to prevent or curtail cholera outbreaks
(~6 months), the memory B cells that are responsible for relies heavily on epidemiological research as different endemic
preventing cholera infection persist and can quickly expand and regions need distinguishing vaccines to target the divergent
differentiate into plasmablasts and eventually the plasma cells, circulating strains. Moreover, ideal cholera vaccines will not be
which can reseed protective antibodies upon antigen-contact dependent on cold-chain shipping. In addition to these three
activation28. Moreover, OCVs could provide herd immunity to treatments, which are currently used, new approaches to prevent
unvaccinated adults, but the effect in unvaccinated children or treat cholera infections are emerging.
requires further study29.
Probiotics
One widely used WC strain vaccine is Dukoral (CTB-WC), An emerging concept in microbiology is the ability of the host
which contains inactivated/dead V. cholerae O1 (El Tor and microbiome to prevent or limit infections. A relatively new
classical biotypes) with the addition of recombinant B subunits concept for V. cholerae, this idea is beginning to be explored
of CT (CTB)12,16,30,31. The effectiveness of Dukoral is between as a treatment or prevention for cholera infections. Owing to
55% and 88%17, and it is intended for travelers but—owing the excessive fluid accumulation, V. cholerae elicits severe
to its short period of usability, high cost, and its requirements disruption to the gut microbiome during infection such that
for cold-chain circulation—not for populations in endemic the majority of bacteria found in the characteristic rice-water
regions9,25. Dukoral can provide protection from infection for stools are V. cholerae36. Furthermore, the type VI secretion
2 years in vaccinated individuals above the age of 5, but it is system of V. cholerae can deliver effector toxins to the gut
effective for only 6 months between the ages of 2 and 5 and microbiome or modulate host cells themselves, both of which
requires at least two doses to be effective16. Unlike Dukoral, alter the gut microbiota to facilitate colonization37,38. For these
Shanchol and Euvichol are WC vaccines composed of inac- reasons, restoration of the gut microbiome or prevention of
tivated O1 Inaba, O1 Ogawa, and O139 strains, but these colonization through probiotic treatment is a promising new
vaccines do not contain CTB16,32. The efficacy of Shanchol is approach to treat cholera infections.
about 65% protection based on a 5-year study17. In clinical
studies in the Philippines, Euvichol was effective in adults and Several bacterial species have been shown to dispel or suppress
children18,32. Shanchol and Euvichol are intended for all patients cholera infections. Ruminococcus obeum increases in its
who are at least 1 year old but not for pregnant women16,18. relative abundance after V. cholerae infection of mice and
Furthermore, there are two variations of Euvichol (with or restricts V. cholerae colonization by disrupting its quorum-
without the preservative thimerosal), both of which show no sensing system39. Interestingly, R. obeum is one of the species
significant difference in protection32. These WHO prequalified in the human gut microbiome whose abundance positively
inactivated vaccines can provide protection against cholera correlates with recovery from cholera39. Co-culture of V. cholerae
for at least 3 years and are not available in the US4. with Lactobacillus rhamnosus GG or Bifidobacterium longum
46 decreases CT production in vitro40. One study engineered a
Aside from the inactivated V. cholerae vaccines, the oral probiotic strain of Lactococcus lactis that increases the produc-
live-attenuated vaccine Vaxchora (CVD 103-HgR) is a US Food tion of lactic acid upon detecting the quorum-sensing signals

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of V. cholerae, thus decreasing the pH of the surrounding and thus the most likely application of phages would require a
medium to reduce V. cholerae during co-culture41. Another lytic phage cocktail that would necessitate multiple independent
experiment engineered an E. coli strain to mimic the CT binding mutations for resistance. Because of the intricate connection
ganglioside on its surface and this strain reduced the symp- between V. cholerae and lytic phages, this bacterium encodes
toms of a V. cholerae infection by decreasing the free CT42. The molecular defense mechanisms to limit phage infection49.
culture supernatant of a fecal Lactobacilli strain was exploited Owing to host specificity, which has a significant impact
to disturb a V. cholerae biofilm by increasing the pH to on treatment development and testing, mass production and
potentially reduce stress in the gut43. Lastly, E. coli was distribution of phage therapy, are not yet practical50. Before
demonstrated to decrease the colonization of V. cholerae when phage therapy is validated as a cholera therapeutic, there must
co-cultured with glucose in a zebrafish infection model by be an assessment of the immunological response to phages and
lowering the pH44. This finding is intriguing as it suggests a efficacy during cholera outbreaks.
possible synergistic effect between probiotics and glucose-based
ORT44. Conclusions
The biggest challenges to treat V. cholerae are the inherent
Probiotics often are taken with antibiotics and various other complications in endemic or disrupted regions, including
drugs, including anti-inflammatory adjuvants, but the adverse economics, natural disasters, wars, national security, and poor
side effect of the drugs on the probiotics warrants further infrastructure. Because of these challenges, a multi-pronged
testing43. Nevertheless, probiotics that reduce cholera may limit approach that is flexible to the specific demands of a current
antibiotic-resistant V. cholerae strains by reducing the quantity outbreak is needed to treat cholera. ORT has been and will
of antibiotics used41,42. Moreover, probiotics could serve as a continue to be a front-line defense to save patients already
better treatment in regions where cold-chain vaccine preparation infected with V. cholerae because it is cost-effective and easy
is not feasible and clean water supply is not available40. to use and combining antibiotics with ORT clearly reduces the
severity of disease. Additionally, the WHO has prequalified
Phage therapy Dukoral, Shanchol, and Euvichol as current cholera vaccines,
Another novel treatment for cholera involves the therapeu- although there is a worldwide deficiency of these vaccines2.
tic use of lytic bacteriophages. Phage therapy has been used for Increasing the availability of these vaccines could have a
decades in Eastern Europe and Russia and, with the emergence significant impact in reducing infections during an outbreak.
of antibiotic-resistant bacteria, has been developed and used Whether probiotics or phage therapy can work synergistically
to treat infections caused by Pseudomonas, Salmonella, and with ORT, antibiotics, or vaccines to treat or—preferably—to
Staphylococcus45. Phage therapy has many advantages over prevent cholera infections remains to be tested in the field.
antibiotics. For example, phages are able to kill antibiotic- Another approach to control cholera is the development
resistant bacteria, the amount of phages increases proportion- of anti-virulence compounds that inhibit the expression of
ally to the number of infecting bacteria, and the phages exert a virulence factors, thereby protecting the host from colonization
minimal impact on the resident microbiome45. This treatment by V. cholerae. Such compounds include virstatin51, a conjugated
strategy is inspired by the natural life cycle of V. cholerae in form of linoleic acid52, and synthetic compounds that resemble
which blooms of the bacteria during outbreaks are followed by folded fatty acids53. Further development of these anti-virulence
the expansion of lytic bacteriophages, primarily ICP1, ICP2, therapeutics requires testing these compounds during human
and ICP3, which ultimately reduce the population of viable infections and assessing their practicality to treat cholera
bacteria46,47. outbreaks. Ultimately, health surveillance plays a critical role
in preventing outbreaks by directing proactive countermeasures
A study in an infant mouse model has shown that a cocktail of during emergencies. A global commitment to reduce pandemic
the three ICP virulent bacteriophages could effectively reduce cholera requires devising better methods to quickly identify
the V. cholerae load after challenge in a dose-dependent manner outbreak strains, recognizing the best treatment option for the
because of the phage’s fast replication and ability to kill given strain, and developing new therapies that are not depend-
the bacteria48. Although it shows promising results in animal ent on cold-chain systems or clean water. The combination of
models, phage therapy for cholera requires more optimization current treatments with new therapies has significant potential
for its effectiveness and timing to be advanced to clinical to further combat the centuries-old human scourge of cholera.
trials48. Phage therapy could also be used to limit person-to-
person spread as even small numbers of these lytic phages could
rapidly expand during V. cholerae infections.
Grant information
Although phage therapy has many promising characteris- This work was supported by National Institutes of Health grants
tics, it also has potential drawbacks. During phage therapy, the GM109259 and AI130554 and National Science Foundation grant
host’s adaptive immune system can generate phage-neutralizing MCB1714612 to CMW.
antibodies that could inhibit their ability to lyse the targeted
bacteria in vivo or prevent subsequent treatments45. As with The funders had no role in study design, data collection and
antibiotics, V. cholerae will evolve resistance to phage infection analysis, decision to publish, or preparation of the manuscript.

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Version 1
1 Jeffrey Withey Department of Biochemistry, Microbiology and Immunology, Wayne State University School
of Medicine, Detroit, Michigan, USA
Competing Interests: No competing interests were disclosed.
2 Jyl Matson Department of Medical Microbiology and Immunology, University of Toledo College of Medicine
and Life Sciences, Toledo, OH, USA
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